Inhibiting aerobic glycolysis suppresses renal interstitial fibroblast activation and renal fibrosis.

PubMed

Ding, Hao; Jiang, Lei; Xu, Jing; Bai, Feng; Zhou, Yang; Yuan, Qi; Luo, Jing; Zen, Ke; Yang, Junwei

2017-09-01

Chronic kidney diseases generally lead to renal fibrosis. Despite great progress having been made in identifying molecular mediators of fibrosis, the mechanism that governs renal fibrosis remains unclear, and so far no effective therapeutic antifibrosis strategy is available. Here we demonstrated that a switch of metabolism from oxidative phosphorylation to aerobic glycolysis (Warburg effect) in renal fibroblasts was the primary feature of fibroblast activation during renal fibrosis and that suppressing renal fibroblast aerobic glycolysis could significantly reduce renal fibrosis. Both gene and protein assay showed that the expression of glycolysis enzymes was upregulated in mouse kidneys with unilateral ureter obstruction (UUO) surgery or in transforming growth factor-β1 (TGF-β1)-treated renal interstitial fibroblasts. Aerobic glycolysis flux, indicated by glucose uptake and lactate production, was increased in mouse kidney with UUO nephropathy or TGF-β1-treated renal interstitial fibroblasts and positively correlated with fibrosis process. In line with this, we found that increasing aerobic glycolysis can remarkably induce myofibroblast activation while aerobic glycolysis inhibitors shikonin and 2-deoxyglucose attenuate UUO-induced mouse renal fibrosis and TGF-β1-stimulated myofibroblast activation. Furthermore, mechanistic study indicated that shikonin inhibits renal aerobic glycolysis via reducing phosphorylation of pyruvate kinase type M2, a rate-limiting glycolytic enzyme associated with cell reliance on aerobic glycolysis. In conclusion, our findings demonstrate the critical role of aerobic glycolysis in renal fibrosis and support treatment with aerobic glycolysis inhibitors as a potential antifibrotic strategy. Copyright © 2017 the American Physiological Society.

High levels of the extracellular matrix proteoglycan decorin are associated with inhibition of testicular function

PubMed Central

Adam, Marion; Urbanski, Henryk F.; Garyfallou, Vasilios T.; Welsch, Ulrich; Köhn, Frank M.; Schwarzer, J. Ullrich; Strauss, Leena; Poutanen, Matti; Mayerhofer, Artur

2011-01-01

Decorin (DCN), a component of the extracellular matrix of the peritubular wall and the interstitial areas of the human testis, can interact with growth factor (GF) signaling, thereby blocking downstream actions of GFs. In the present study the expression and regulation of DCN using both human testes and two experimental animal models, namely the rhesus monkey and mouse, were examined. DCN protein was present in peritubular and interstitial areas of adult human and monkey testes, while it was almost undetectable in adult wild-type mice. Interestingly, the levels and sites of testicular DCN expression in the monkeys were inversely correlated with testicular maturation markers. A strong DCN expression associated with the abundant connective tissue of the interstitial areas in the postnatal through prepubertal phases was observed. In adult and old monkeys the DCN pattern was similar to the one in normal human testes, presenting strong expression at the peritubular region. In the testes of both infertile men and in a mouse model of inflammation associated infertility (aromatase-overexpressing transgenic mice), the fibrotic changes and increased numbers of Tumor necrosis factor (TNF)-α-producing immune cells were shown to be associated with increased production of DCN. Furthermore, studies with human testicular peritubular cells isolated from fibrotic testis indicated that TNF-α significantly increased DCN production. The data, thus, show that an increased DCN level is associated with impaired testicular function, supporting our hypothesis that DCN interferes with paracrine signaling of the testis in health and disease. PMID:22413766

Accumulation of BDCA1⁺ dendritic cells in interstitial fibrotic lung diseases and Th2-high asthma.

PubMed

Greer, Alexandra M; Matthay, Michael A; Kukreja, Jasleen; Bhakta, Nirav R; Nguyen, Christine P; Wolters, Paul J; Woodruff, Prescott G; Fahy, John V; Shin, Jeoung-Sook

2014-01-01

Dendritic cells (DCs) significantly contribute to the pathology of several mouse lung disease models. However, little is known of the contribution of DCs to human lung diseases. In this study, we examined infiltration with BDCA1⁺ DCs of human lungs in patients with interstitial lung diseases or asthma. Using flow cytometry, we found that these DCs increased by 5∼6 fold in the lungs of patients with idiopathic pulmonary fibrosis or hypersensitivity pneumonitis, which are both characterized by extensive fibrosis in parenchyma. The same DC subset also significantly increased in the lung parenchyma of patients with chronic obstructive pulmonary disease, although the degree of increase was relatively modest. By employing immunofluorescence microscopy using FcεRI and MHCII as the specific markers for BDCA1⁺ DCs, we found that the numbers of BDCA1⁺ DCs also significantly increased in the airway epithelium of Th2 inflammation-associated asthma. These findings suggest a potential contribution of BDCA1⁺ DCs in human lung diseases associated with interstitial fibrosis or Th2 airway inflammation.

Ulinastatin inhibits renal tubular epithelial apoptosis and interstitial fibrosis in rats with unilateral ureteral obstruction.

PubMed

Zhang, Qing-Fang

2017-12-01

The effect of ulinastatin (UTI) on renal tubular epithelial apoptosis and interstitial fibrosis in rats with unilateral ureteral obstruction (UUO) was investigated. A total of 18 male Wistar rats were randomly divided into the following 3 groups: The Sham group (n=6), the UUO group (n=6), and the UTI group (n=6). In the UUO and UTI groups, the left ureter was ligated to establish a UUO model. Starting from day 1 after surgery, an intervention treatment was performed using normal saline (1 ml/kg/d) and UTI (40,000 unit/kg/d). On day 7 after surgery, 6 rats from each group were sacrificed. In the Sham group, the left ureter was only freed, not ligated; after 7 days of abdominal closure, all of the rats were sacrificed. Blood samples were collected prior to sacrificing the animals to measure the blood urea nitrogen (BUN) and serum creatinine (Scr). The incidence of renal interstitial lesions on the obstruction side was observed by hematoxylin and eosin, and Masson staining. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, and immunohistochemical detection of apoptosis regulator Bax (Bax), apoptosis regulator Bcl‑2 (Bcl‑2) and caspase‑3 were performed to observe the presence of renal tubular epithelial cell apoptosis. The UTI did not have a significant influence on the mouse BUN and Scr levels in any of the groups (P>0.05). Compared with that in the Sham group, renal tissue injury in the UUO group was significantly aggravated with renal tubular dilation, epithelial cell atrophy, renal interstitial inflammatory cell infiltration and fibrous tissue hyperplasia (P<0.01). Furthermore, the renal tubular epithelial TUNEL+ cell number and Bax and caspase‑3 levels were increased, and the expression of Bcl‑2 was decreased (P<0.01). Following the UTI treatment, the renal interstitial injury at the obstruction side was significantly attenuated (P<0.05), the renal tubular epithelial TUNEL+ cell number, and Bax and caspase‑3 levels significantly decreased, and the expression of Bcl‑2 was restored (P<0.05). UTI inhibited renal tubular epithelial apoptosis and interstitial fibrosis in UUO rats.

Carbachol regulates pacemaker activities in cultured interstitial cells of Cajal from the mouse small intestine.

PubMed

So, Keum Young; Kim, Sang Hun; Sohn, Hong Moon; Choi, Soo Jin; Parajuli, Shankar Prasad; Choi, Seok; Yeum, Cheol Ho; Yoon, Pyung Jin; Jun, Jae Yeoul

2009-05-31

We studied the effect of carbachol on pacemaker currents in cultured interstitial cells of Cajal (ICC) from the mouse small intestine by muscarinic stimulation using a whole cell patch clamp technique and Ca2+-imaging. ICC generated periodic pacemaker potentials in the current-clamp mode and generated spontaneous inward pacemaker currents at a holding potential of-70 mV. Exposure to carbachol depolarized the membrane and produced tonic inward pacemaker currents with a decrease in the frequency and amplitude of the pacemaker currents. The effects of carbachol were blocked by 1-dimethyl-4-diphenylacetoxypiperidinium, a muscarinic M(3) receptor antagonist, but not by methotramine, a muscarinic M(2) receptor antagonist. Intracellular GDP-beta-S suppressed the carbachol-induced effects. Carbachol-induced effects were blocked by external Na+-free solution and by flufenamic acid, a non-selective cation channel blocker, and in the presence of thapsigargin, a Ca2+-ATPase inhibitor in the endoplasmic reticulum. However, carbachol still produced tonic inward pacemaker currents with the removal of external Ca2+. In recording of intracellular Ca2+ concentrations using fluo 3-AM dye, carbachol increased intracellular Ca2+ concentrations with increasing of Ca2+ oscillations. These results suggest that carbachol modulates the pacemaker activity of ICC through the activation of non-selective cation channels via muscarinic M(3) receptors by a G-protein dependent intracellular Ca2+ release mechanism.

Roles of CD34+ cells and ALK5 signaling in the reconstruction of seminiferous tubule-like structures in 3-D re-aggregate culture of dissociated cells from neonatal mouse testes.

PubMed

Abe, Shin-Ichi; Abe, Kazuko; Zhang, Jidong; Harada, Tomoaki; Mizumoto, Go; Oshikawa, Hiroki; Akiyama, Haruhiko; Shimamura, Kenji

2017-01-01

Tissue reconstruction in vitro can provide, if successful, a refined and simple system to analyze the underlying mechanisms that drive the morphogenesis and maintain the ordered structure. We have recently succeeded in reconstruction of seminiferous cord-like and tubule-like structures using 3-D re-aggregate culture of dissociated testicular cells. In testis formation, endothelial cells that migrated from mesonephroi to embryonic gonads have been shown to be critical for development of testis cords, but how endothelial cells contribute to testis cord formation remains unknown. To decipher the roles of endothelial and peritubular cells in the reconstruction of cord-like and tubule-like structures, we investigated the behavior of CD34+ endothelial and p75+ cells, and peritubular myoid cells (PTMCs) in 3-D re-aggregate cultures of testicular cells. The results showed that these 3 types of cells had the capacity of re-aggregation on their own and with each other, and of segregation into 3 layers in a re-aggregate, which were very similar to interstitial and peritubular tissues in vivo. Observation of behaviors of fluorescent Sertoli cells and other non-fluorescent types of cells using testes from Sox9-EGFP transgenic mice showed dynamic cell movement and segregation in re-aggregate cultures. Cultures of testicular cells deprived of interstitial and peritubular cells resulted in dysmorphic structures, but re-addition of them restored tubule-like structures. Purified CD34+ cells in culture differentiated into p75+ cells and PTMCs. These results indicate that CD34+ cells differentiate into p75+ cells, which then differentiate into PTMCs. TGFβ signaling inhibitors, SB431542 and ALK5i, disturbed the reconstruction of cord-like and tubule-like structures, and the latter compromised re-construction of interstitial-like and peritubular-like structures, as well as the proliferation of CD34+, p75+, PTMCs, and Sertoli cells, and their movement and differentiation. These results indicate that CD34+ cells and signaling through ALK5 play pivotal roles in the morphogenesis of interstitial-like, peritubular-like and cord-like structures.

Regional variation in contribution of myenteric and intramuscular interstitial cells of Cajal to generation of slow waves in mouse gastric antrum

PubMed Central

Hirst, G D S; Beckett, E A H; Sanders, K M; Ward, S M

2002-01-01

When intracellular recordings were made from the antral region of murine stomach, cells with three different patterns of electrical activity were detected. One group of cells generated follower potentials, the second group generated pacemaker potentials and the third group generated slow waves that consisted of primary and secondary components. Slow waves recorded in different regions of the gastric antrum had similar amplitudes but different characteristic shapes. At the greater curvature, slow waves had large initial components. Midway between the greater and lesser curvature, the amplitude of the initial component was reduced and at the lesser curvature an initial component was difficult to detect. When the distributions of myenteric (ICC-MY) and intramuscular interstitial cells of Cajal (ICC-IM) were determined, using an antibody to Kit, ICC-MY were found to be present at the greater curvature but were greatly reduced in density at the lesser curvature. In contrast, ICC-IM were found in the circular layer of each region. When recordings were made from the antrum of W/WV mice, which lack ICC-IM, incomplete slow waves were detected and their amplitudes fell from the greater to the lesser curvature. Again, a corresponding fall in the density of ICC-MY was detected. The observations indicate that the contribution of ICC-MY and ICC-IM to the generation of slow waves varies in different regions of the mouse gastric antrum. PMID:11986385

In vitro organogenesis of gut-like structures from mouse embryonic stem cells.

PubMed

Kuwahara, M; Ogaeri, T; Matsuura, R; Kogo, H; Fujimoto, T; Torihashi, S

2004-04-01

Embryonic stem (ES) cells have pluripotency and give rise to many cell types and tissues, including representatives of all three germ layers in the embryo. We have reported previously that mouse ES cells formed contracting gut-like organs from embryoid bodies (EBs). These gut-like structures contracted spontaneously, and had large lumens surrounded by three layers, i.e. epithelium, lamina propria and muscularis. Ganglia were scattered along the periphery, and interstitial cells of Cajal (ICC) were distributed among the smooth muscle cells. In the present study, to determine whether they can be a model of gut organogenesis, we investigated the formation process of the gut-like structures in comparison with embryonic gut development. As a result, we found that the fundamental process of formation in vitro was similar to embryonic gut development in vivo. The result indicates that the gut-like structure is a useful tool not only for developmental study to determine the factors that induce gut organogenesis, but also for studies of enteric neurone and ICC development.

Extracellular Cl- regulates electrical slow waves and setting of smooth muscle membrane potential by interstitial cells of Cajal in mouse jejunum.

PubMed

Saravanaperumal, Siva Arumugam; Gibbons, Simon J; Malysz, John; Sha, Lei; Linden, David R; Szurszewski, Joseph H; Farrugia, Gianrico

2018-01-01

What is the central question of this study? The aim was to investigate the roles of extracellular chloride in electrical slow waves and resting membrane potential of mouse jejunal smooth muscle by replacing chloride with the impermeant anions gluconate and isethionate. What is the main finding and its importance? The main finding was that in smooth muscle cells, the resting Cl - conductance is low, whereas transmembrane Cl - movement in interstitial cells of Cajal (ICCs) is a major contributor to the shape of electrical slow waves. Furthermore, the data confirm that ICCs set the smooth muscle membrane potential and that altering Cl - homeostasis in ICCs can alter the smooth muscle membrane potential. Intracellular Cl - homeostasis is regulated by anion-permeable channels and transporters and contributes to excitability of many cell types, including smooth muscle and interstitial cells of Cajal (ICCs). Our aims were to investigate the effects on electrical activity in mouse jejunal muscle strips of replacing extracellular Cl - (Cl - o ) with the impermeant anions gluconate and isethionate. On reducing Cl - o , effects were observed on electrical slow waves, with small effects on smooth muscle membrane voltage (E m ). Restoration of Cl - hyperpolarized smooth muscle E m proportional to the change in Cl - o concentration. Replacement of 90% of Cl - o with gluconate reversibly abolished slow waves in five of nine preparations. Slow waves were maintained in isethionate. Gluconate and isethionate substitution had similar concentration-dependent effects on peak amplitude, frequency, width at half peak amplitude, rise time and decay time of residual slow waves. Gluconate reduced free ionized Ca 2+ in Krebs solutions to 0.13 mm. In Krebs solutions containing normal Cl - and 0.13 mm free Ca 2+ , slow wave frequency was lower, width at half peak amplitude was smaller, and decay time was faster. The transient hyperpolarization following restoration of Cl - o was not observed in W/W v mice, which lack pacemaker ICCs in the small intestine. We conclude that in smooth muscle cells, the resting Cl - conductance is low, whereas transmembrane Cl - movement in ICCs plays a major role in generation or propagation of slow waves. Furthermore, these data support a role for ICCs in setting smooth muscle E m and that altering Cl - homeostasis in ICCs can alter smooth muscle E m . © 2017 Mayo Clinic. Experimental Physiology © 2017 The Physiological Society.

Muscle Weakness and Fibrosis Due to Cell Autonomous and Non-cell Autonomous Events in Collagen VI Deficient Congenital Muscular Dystrophy.

PubMed

Noguchi, Satoru; Ogawa, Megumu; Malicdan, May Christine; Nonaka, Ikuya; Nishino, Ichizo

2017-02-01

Congenital muscular dystrophies with collagen VI deficiency are inherited muscle disorders with a broad spectrum of clinical presentation and are caused by mutations in one of COL6A1-3 genes. Muscle pathology is characterized by fiber size variation and increased interstitial fibrosis and adipogenesis. In this study, we define critical events that contribute to muscle weakness and fibrosis in a mouse model with collagen VI deficiency. The Col6a1 GT/GT mice develop non-progressive weakness from younger age, accompanied by stunted muscle growth due to reduced IGF-1 signaling activity. In addition, the Col6a1 GT/GT mice have high numbers of interstitial skeletal muscle mesenchymal progenitor cells, which dramatically increase with repeated myofiber necrosis/regeneration. Our results suggest that impaired neonatal muscle growth and the activation of the mesenchymal cells in skeletal muscles contribute to the pathology of collagen VI deficient muscular dystrophy, and more importantly, provide the insights on the therapeutic strategies for collagen VI deficiency. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Involvement of mitochondrial Na+–Ca2+ exchange in intestinal pacemaking activity

PubMed Central

Kim, Byung Joo; Jun, Jae Yeoul; So, Insuk; Kim, Ki Whan

2006-01-01

AIM: Interstitial cells of Cajal (ICCs) are the pacemaker cells that generate slow waves in the gastrointestinal (GI) tract. We have aimed to investigate the involvement of mitochondrial Na+-Ca2+ exchange in intestinal pacemaking activity in cultured interstitial cells of Cajal. METHODS: Enzymatic digestions were used to dissociate ICCs from the small intestine of a mouse. The whole-cell patch-clamp configuration was used to record membrane currents (voltage clamp) and potentials (current clamp) from cultured ICCs. RESULTS: Clonazepam and CGP37157 inhibited the pacemaking activity of ICCs in a dose-dependent manner. Clonazepam from 20 to 60 µmol/L and CGP37157 from 10 to 30 µmol/L effectively inhibited Ca2+ efflux from mitochondria in pacemaking activity of ICCs. The IC50s of clonazepam and CGP37157 were 37.1 and 18.2 µmol/L, respectively. The addition of 20 µmol/L NiCl2 to the internal solution caused a “wax and wane” phenomenon of pacemaking activity of ICCs. CONCLUSION: These results suggest that mitochondrial Na+-Ca2+ exchange has an important role in intestinal pacemaking activity. PMID:16521198

A role for MCP-1/CCR2 in interstitial lung disease in children

PubMed Central

Hartl, Dominik; Griese, Matthias; Nicolai, Thomas; Zissel, Gernot; Prell, Christine; Reinhardt, Dietrich; Schendel, Dolores J; Krauss-Etschmann, Susanne

2005-01-01

Background Interstitial lung diseases (ILD) are chronic inflammatory disorders leading to pulmonary fibrosis. Monocyte chemotactic protein 1 (MCP-1) promotes collagen synthesis and deletion of the MCP-1 receptor CCR2 protects from pulmonary fibrosis in ILD mouse models. We hypothesized that pulmonary MCP-1 and CCR2+ T cells accumulate in pediatric ILD and are related to disease severity. Methods Bronchoalveolar lavage fluid was obtained from 25 children with ILD and 10 healthy children. Levels of pulmonary MCP-1 and Th1/Th2-associated cytokines were quantified at the protein and the mRNA levels. Pulmonary CCR2+, CCR4+, CCR3+, CCR5+ and CXCR3+ T cells were quantified by flow-cytometry. Results CCR2+ T cells and MCP-1 levels were significantly elevated in children with ILD and correlated with forced vital capacity, total lung capacity and ILD disease severity scores. Children with lung fibrosis had significantly higher MCP-1 levels and CCR2+ T cells in bronchoalveolar lavage fluid compared to non-fibrotic children. Conclusion The results indicate that pulmonary CCR2+ T cells and MCP-1 contribute to the pathogenesis of pediatric ILD and might provide a novel target for therapeutic strategies. PMID:16095529

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ohori, N.P.; Sciurba, F.C.; Owens, G.R.

We report four cases of giant-cell interstitial pneumonia that occurred in association with exposure to hard metals. All patients presented with chronic interstitial lung disease and had open-lung biopsies that revealed marked interstitial fibrosis, cellular interstitial infiltrates, and prominent intraalveolar macrophages as well as giant cells displaying cellular cannibalism. We also review the literature to determine the sensitivity and specificity of giant-cell interstitial pneumonia for hard-metal pneumoconiosis. Although hard-metal pneumoconiosis may take the form of usual interstitial pneumonia, desquamative interstitial pneumonia, and giant-cell interstitial pneumonia, the finding of giant-cell interstitial pneumonia is almost pathognomonic of hard-metal disease and should provokemore » an investigation of occupational exposure. 25 references.« less

Inhibitory effect of tributyltin on expression of steroidogenic enzymes in mouse testis.

PubMed

Kim, Suel-Kee; Kim, Jong-Hoon; Han, Jung Ho; Yoon, Yong-Dal

2008-01-01

Tributyltin (TBT) is known to disrupt the development of reproductive organs, thereby reducing fertility. The aim of this study was to evaluate the acute toxicity of TBT on the testicular development and steroid hormone production. Immature (3-week-old) male mice were given a single administration of 25, 50, or 100 mg/kg of TBT by oral gavage. Lumen formation in seminiferous tubule was remarkably delayed, and the number of apoptotic germ cells found inside the tubules was increased in the TBT-exposed animals, whereas no apoptotic signal was observed in interstitial Leydig cells. Reduced serum testosterone concentration and down-regulated expressions of the mRNAs for cholesterol side-chain cleavage enzyme (P450scc), 17alpha -hydroxylase/C(17-20) lyase (P450(17alpha)), 3beta -hydroxysteroid-dehydrogenase (3beta -HSD), and 17beta -hydroxysteroid-dehydrogenase (17beta -HSD) were also observed after TBT exposure. Altogether, these findings demonstrate that exposure to TBT is associated with induced apoptosis of testicular germ cells and inhibition of steroidogenesis by reduction in the expression of steroidogenic enzymes in interstitial Leydig cells. These adverse effects of TBT would cause serious defects in testicular development and function.

Muc1 deficiency exacerbates pulmonary fibrosis in a mouse model of silicosis.

PubMed

Kato, Kosuke; Zemskova, Marina A; Hanss, Alec D; Kim, Marianne M; Summer, Ross; Kim, Kwang Chul

2017-11-25

MUC1 (MUC in human and Muc in animals) is a membrane-tethered mucin expressed on the apical surface of lung epithelial cells. However, in the lungs of patients with interstitial lung disease, MUC1 is aberrantly expressed in hyperplastic alveolar type II epithelial (ATII) cells and alveolar macrophages (AM), and elevated levels of extracellular MUC1 are found in bronchoalveolar lavage (BAL) fluid and the serum of these patients. While pro-fibrotic effects of extracellular MUC1 have recently been described in cultured fibroblasts, the contribution of MUC1 to the pathobiology of pulmonary fibrosis is unknown. In this study, we hypothesized that MUC1 deficiency would reduce susceptibility to pulmonary fibrosis in a mouse model of silicosis. We employed human MUC1 transgenic mice, Muc1 deficient mice and wild-type mice on C57BL/6 background in these studies. Some mice received a one-time dose of crystalline silica instilled into their oropharynx in order to induce pulmonary fibrosis and assess the effects of Muc1 deficiency on fibrotic and inflammatory responses in the lung. As previously described in other mouse models of pulmonary fibrosis, we found that extracellular MUC1 levels were markedly increased in whole lung tissues, BALF and serum of human MUC1 transgenic mice after silica. We also detected an increase in total MUC1 levels in the lungs of these mice, indicating that production as well as release contributed to elevated levels after lung injury. Immunohistochemical staining revealed that increased MUC1 expression was mostly confined to ATII cells and AMs in areas of fibrotic remodeling, illustrating a pattern similar to the expression of MUC1 in human fibrotic lung tissues. However, contrary to our hypothesis, we found that Muc1 deficiency resulted in a worsening of fibrotic remodeling in the mouse lung as judged by an increase in number of silicotic nodules, an increase in lung collagen deposition and an increase in the severity of pulmonary inflammation. Altogether, our results indicate that Muc1 has anti-fibrotic properties in the mouse lung and suggest that elevated levels of MUC1 in patients with interstitial lung disease may serve a protective role, which aims to limit the severity of tissue remodeling in the lung. Copyright © 2017. Published by Elsevier Inc.

Nonclinical safety of the sodium-glucose cotransporter 2 inhibitor empagliflozin.

PubMed

Bogdanffy, Matthew S; Stachlewitz, Robert F; van Tongeren, Susan; Knight, Brian; Sharp, Dale E; Ku, Warren; Hart, Susan Emeigh; Blanchard, Kerry

2014-01-01

Empagliflozin, a selective inhibitor of the renal tubular sodium-glucose cotransporter 2, was developed for treatment of type 2 diabetes mellitus. Nonclinical safety of empagliflozin was studied in a battery of tests to support global market authorization. Safety pharmacology studies indicated no effect of empagliflozin on measures of respiratory or central nervous system function in rats or cardiovascular safety in telemeterized dogs. In CD-1 mouse, Wistar Han rat, or beagle dogs up to 13, 26, or 52 weeks of treatment, respectively, empagliflozin exhibited a toxicity profile consistent with secondary supratherapeutic pharmacology related to glucose loss and included decreased body weight and body fat, increased food consumption, diarrhea, dehydration, decreased serum glucose and increases in other serum parameters reflective of increased protein catabolism, gluconeogenesis, and electrolyte imbalances, and urinary changes such as polyuria and glucosuria. Microscopic changes were consistently observed in kidney and included tubular nephropathy and interstitial nephritis (dog), renal mineralization (rat) and tubular epithelial cell karyomegaly, single cell necrosis, cystic hyperplasia, and hypertrophy (mouse). Empagliflozin was not genotoxic. Empagliflozin was not carcinogenic in female mice or female rats. Renal adenoma and carcinoma were induced in male mice only at exposures 45 times the maximum clinical dose. These tumors were associated with a spectrum of nonneoplastic changes suggestive of a nongenotoxic, cytotoxic, and cellular proliferation-driven mechanism. In male rats, testicular interstitial cell tumors and hemangiomas of the mesenteric lymph node were observed; both tumors are common in rats and are unlikely to be relevant to humans. These studies demonstrate the nonclinical safety of empagliflozin. © The Author(s) 2014.

Alveolar Macrophages Drive Hepatocellular Carcinoma Lung Metastasis by Generating Leukotriene B4.

PubMed

Nosaka, Takuto; Baba, Tomohisa; Tanabe, Yamato; Sasaki, Soichiro; Nishimura, Tatsunori; Imamura, Yoshiaki; Yurino, Hideaki; Hashimoto, Shinichi; Arita, Makoto; Nakamoto, Yasunari; Mukaida, Naofumi

2018-03-01

Macrophages in lungs can be classified into two subpopulations, alveolar macrophages (AMs) and interstitial macrophages (IMs), which reside in the alveolar and interstitial spaces, respectively. Accumulating evidence indicates the involvement of IMs in lung metastasis, but the roles of AMs in lung metastasis still remain elusive. An i.v. injection of a mouse hepatocellular carcinoma (HCC) cell line, BNL, caused lung metastasis foci with infiltration of AMs and IMs. Comprehensive determination of arachidonic acid metabolite levels revealed increases in leukotrienes and PGs in lungs in this metastasis model. A 5-lipoxygenase (LOX) inhibitor but not a cyclooxygenase inhibitor reduced the numbers of metastatic foci, particularly those of a larger size. A major 5-LOX metabolite, LTB 4 , augmented in vitro cell proliferation of human HCC cell lines as well as BNL cells. Moreover, in this lung metastasis course, AMs exhibited higher expression levels of the 5-LOX and LTB 4 than IMs. Consistently, 5-LOX-expressing AMs increased in the lungs of human HCC patients with lung metastasis, compared with those without lung metastasis. Furthermore, intratracheal clodronate liposome injection selectively depleted AMs but not IMs, together with reduced LTB 4 content and metastatic foci numbers in this lung metastasis process. Finally, IMs in mouse metastatic foci produced CCL2, thereby recruiting blood-borne, CCR2-expressing AMs into lungs. Thus, AMs can be recruited under the guidance of IM-derived CCL2 into metastatic lungs and can eventually contribute to the progression of lung metastasis by providing a potent arachidonic acid-derived tumor growth promoting mediator, LTB 4 . Copyright © 2018 by The American Association of Immunologists, Inc.

Effects of Dai-kenchu-to on spontaneous activity in the mouse small intestine.

PubMed

Kito, Yoshihiko; Suzuki, Hikaru

2006-12-01

The effects of Dai-kenchu-to (DKT), a Chinese medicine, on spontaneous activity of mouse small intestine were investigated. Experiments were carried out with tension recording and intracellular recording. DKT contracted mouse longitudinal smooth muscles in a dose dependent manner (0.1-10 mg/ml). Low concentration of DKT (0.1 mg/ml) did not contract the longitudinal muscles of mouse small intestine. DKT (0.1 mg/ml) inhibited contraction elicited by transmural nerve stimulation (TNS). DKT (1 mg/ml) evoked relaxation before contraction. The initial relaxation was abolished by Nomega-nitro-L-arginine (L-NNA). DKT (10 mg/ml)-induced contraction had two components: a transient rapid contraction and a following slow contraction. Atropine inhibited DKT (1 mg/ml)-induced contraction to about 50% of control. In the presence of atropine, tetrodotoxin (TTX) inhibited the contraction elicited by DKT (1 mg/ml) to about 80%. DKT depolarized the membrane and decreased the amplitude of pacemaker potentials recorded from in situ myenteric interstitial cells of Cajal (ICC-MY) with no alteration to the frequency, duration and maximum rates of rise in the presence of nifedipine and TTX. The same results were obtained in slow waves recorded from circular smooth muscle cells. These results indicate that DKT evoked both contraction and relaxation by releasing acetylcholine, nitric oxide and other excitatory neurotransmitters in mouse small intestine. DKT had no effects on pacemaker mechanisms and electrical coupling between ICC-MY and smooth muscle cells in mouse small intestine. The results also suggest that DKT may contract smooth muscles by depolarizing the membrane directly.

Mouse, but Not Human, ApoB-100 Lipoprotein Cholesterol Is a Potent Innate Inhibitor of Streptococcus pneumoniae Pneumolysin

PubMed Central

Wade, Kristin R.; Hotze, Eileen M.; Briles, David E.; Tweten, Rodney K.

2014-01-01

Streptococcus pneumoniae produces the pore-forming toxin pneumolysin (PLY), which is a member of the cholesterol-dependent cytolysin (CDC) family of toxins. The CDCs recognize and bind the 3β-hydroxyl group of cholesterol at the cell surface, which initiates membrane pore formation. The cholesterol transport lipoproteins, which carry cholesterol in their outer monolayer, are potential off-pathway binding targets for the CDCs and are present at significant levels in the serum and the interstitial spaces of cells. Herein we show that cholesterol carried specifically by the ApoB-100-containing lipoprotein particles (CH-ApoB-100) in the mouse, but not that carried by human or guinea pig particles, is a potent inhibitor of the PLY pore-forming mechanism. Cholesterol present in the outer monolayer of mouse ApoB-100 particles is recognized and bound by PLY, which stimulates premature assembly of the PLY oligomeric complex thereby inactivating PLY. These studies further suggest that the vast difference in the inhibitory capacity of mouse CH-ApoB-100 and that of the human and the guinea pig is due to differences in the presentation of cholesterol in the outer monolayer of their ApoB-100 particles. Therefore mouse CH-ApoB-100 represents a significant innate CDC inhibitor that is absent in humans, which may underestimate the contribution of CDCs to human disease when utilizing mouse models of disease. PMID:25188225

Characterization of the liver tissue interstitial fluid (TIF) proteome indicates potential for application in liver disease biomarker discovery.

PubMed

Sun, Wei; Ma, Jie; Wu, Songfeng; Yang, Dong; Yan, Yujuan; Liu, Kehui; Wang, Jinglan; Sun, Longqin; Chen, Ning; Wei, Handong; Zhu, Yunping; Xing, Baocai; Zhao, Xiaohang; Qian, Xiaohong; Jiang, Ying; He, Fuchu

2010-02-05

Tissue interstitial fluid (TIF) forms the interface between circulating body fluids and intracellular fluid. Pathological alterations of liver cells could be reflected in TIF, making it a promising source of liver disease biomarkers. Mouse liver TIF was extracted, separated by SDS-PAGE, analyzed by linear ion trap mass spectrometer, and 1450 proteins were identified. These proteins may be secreted, shed from membrane vesicles, or represent cellular breakdown products. They show different profiling patterns, quantities, and possibly modification/cleavage of intracellular proteins. The high solubility and even distribution of liver TIF supports its suitability for proteome analysis. Comparison of mouse liver TIF data with liver tissue and plasma proteome data identified major proteins that might be released from liver to plasma and serve as blood biomarkers of liver origin. This result was partially supported by comparison of human liver TIF data with human liver and plasma proteome data. Paired TIFs from tumor and nontumor liver tissues of a hepatocellular carcinoma patient were analyzed and the profile of subtracted differential proteins supports the potential for biomarker discovery in TIF. This study is the first analysis of the liver TIF proteome and provides a foundation for further application of TIF in liver disease biomarker discovery.

The multiple faces of leukocyte interstitial migration

PubMed Central

Lämmermann, Tim; Germain, Ronald N.

2014-01-01

Spatiotemporal control of leukocyte dynamics within tissues is critical for successful innate and adaptive immune responses. Homeostatic trafficking and coordinated infiltration into and within sites of inflammation and infection rely on signaling in response to extracellular cues that in turn controls a variety of intracellular protein networks regulating leukocyte motility, migration, chemotaxis, positioning, and cell–cell interaction. In contrast to mesenchymal cells, leukocytes migrate in an amoeboid fashion by rapid cycles of actin polymerization and actomyosin contraction, and their migration in tissues is generally referred to as low adhesive and nonproteolytic. The interplay of actin network expansion, contraction, and adhesion shapes the exact mode of amoeboid migration, and in this review, we explore how leukocyte subsets potentially harness the same basic biomechanical mechanisms in a cell-type-specific manner. Most of our detailed understanding of these processes derives from in vitro migration studies in three-dimensional gels and confined spaces that mimic geometrical aspects of physiological tissues. We summarize these in vitro results and then critically compare them to data from intravital imaging of leukocyte interstitial migration in mouse tissues. We outline the technical challenges of obtaining conclusive mechanistic results from intravital studies, discuss leukocyte migration strategies in vivo, and present examples of mode switching during physiological interstitial migration. These findings are also placed in the context of leukocyte migration defects in primary immunodeficiencies. This overview of both in vitro and in vivo studies highlights recent progress in understanding the molecular and biophysical mechanisms that shape robust leukocyte migration responses in physiologically complex and heterogeneous environments. PMID:24573488

Nitrergic neuromuscular transmission in the mouse internal anal sphincter is accomplished by multiple pathways and postjunctional effector cells

PubMed Central

Sotherton, A. G.; Peri, L. E.; Sanders, K. M.; Ward, S. M.; Keef, K. D.

2014-01-01

The effector cells and second messengers participating in nitrergic neuromuscular transmission (NMT) were investigated in the mouse internal anal sphincter (IAS). Protein expression of guanylate cyclase (GCα, GCβ) and cyclic GMP-dependent protein kinase I (cGKI) were examined in cryostat sections with dual-labeling immunohistochemical techniques in PDGFRα+ cells, interstitial cells of Cajal (ICC), and smooth muscle cells (SMC). Gene expression levels were determined with quantitative PCR of dispersed cells from Pdgfrαegfp/+, KitcopGFP/+, and smMHCCre-egfp mice sorted with FACS. The relative gene and protein expression levels of GCα and GCβ were PDGFRα+ cells > ICC ≫ SMC. In contrast, cGKI gene expression sequence was SMC = ICC > PDGFRα+ cells whereas cGKI protein expression sequence was neurons > SMC ≫ ICC = PDGFRα+ cells. The functional role of cGKI was investigated in cGKI−/− mice. Relaxation with 8-bromo (8-Br)-cGMP was greatly reduced in cGKI−/− mice whereas responses to sodium nitroprusside (SNP) were partially reduced and forskolin responses were unchanged. A nitrergic relaxation occurred with nerve stimulation (NS, 5 Hz, 60 s) in cGKI+/+ and cGKI−/− mice although there was a small reduction in the cGKI−/− mouse. Nω-nitro-l-arginine (l-NNA) abolished responses during the first 20–30 s of NS in both animals. The GC inhibitor ODQ greatly reduced or abolished SNP and nitrergic NS responses in both animals. These data confirm an essential role for GC in NO-induced relaxation in the IAS. However, the expression of GC and cGKI by all three cell types suggests that each may participate in coordinating muscular responses to NO. The persistence of nitrergic NMT in the cGKI−/− mouse suggests the presence of a significant GC-dependent, cGKI-independent pathway. PMID:25301187

Characterization of Epicardial-Derived Cardiac Interstitial Cells: Differentiation and Mobilization of Heart Fibroblast Progenitors

PubMed Central

Ehrbar, Martin; Pérez-Pomares, José M.

2013-01-01

The non-muscular cells that populate the space found between cardiomyocyte fibers are known as ‘cardiac interstitial cells’ (CICs). CICs are heterogeneous in nature and include different cardiac progenitor/stem cells, cardiac fibroblasts and other cell types. Upon heart damage CICs soon respond by initiating a reparative response that transforms with time into extensive fibrosis and heart failure. Despite the biomedical relevance of CICs, controversy remains on the ontogenetic relationship existing between the different cell kinds homing at the cardiac interstitium, as well as on the molecular signals that regulate their differentiation, maturation, mutual interaction and role in adult cardiac homeostasis and disease. Our work focuses on the analysis of epicardial-derived cells, the first cell type that colonizes the cardiac interstitium. We present here a characterization and an experimental analysis of the differentiation potential and mobilization properties of a new cell line derived from mouse embryonic epicardium (EPIC). Our results indicate that these cells express some markers associated with cardiovascular stemness and retain part of the multipotent properties of embryonic epicardial derivatives, spontaneously differentiating into smooth muscle, and fibroblast/myofibroblast-like cells. Epicardium-derived cells are also shown to initiate a characteristic response to different growth factors, to display a characteristic proteolytic expression profile and to degrade biological matrices in 3D in vitro assays. Taken together, these data indicate that EPICs are relevant to the analysis of epicardial-derived CICs, and are a god model for the research on cardiac fibroblasts and the role these cells play in ventricular remodeling in both ischemic or non/ischemic myocardial disease. PMID:23349729

Deficiency of interstitial cells of Cajal in the small intestine of patients with Crohn's disease.

PubMed

Porcher, Christophe; Baldo, Marjolaine; Henry, Monique; Orsoni, Pierre; Julé, Yvon; Ward, Sean M

2002-01-01

Interstitial cells of Cajal are critical for the generation of electrical slow waves that regulate the phasic contractile activity of the tunica muscularis of the GI tract. Under certain pathophysiological conditions loss of interstitial cells of Cajal may play a role in the generation of certain motility disorders. The aim of the present study was to determine if there is an abnormality in the density or distribution of interstitial cells of Cajal from patients with Crohn's disease. Small intestines from control subjects and patients with Crohn's disease were examined using immunohistochemistry and antibodies against the Kit receptor, which is expressed in interstitial cells of Cajal within the tunica muscularis of the GI tract. The density and distribution of interstitial cells of Cajal were assessed in the longitudinal and circular muscle layers and in the myenteric and deep muscular plexus regions of Crohn's and control tissues. Tissues from Crohn's disease patients showed an almost complete abolition of interstitial cells of Cajal within the longitudinal and circular muscle layers and a significant reduction in numbers at the level of the myenteric and deep muscular plexuses. In tissues from Crohn's disease patients, the density of interstitial cells of Cajal was reduced throughout the tunica muscularis in comparison to control small intestines. The disturbance of intestinal motility that occurs in patients with Crohn's disease may be a consequence of the loss of or defects in specific populations of interstitial cells of Cajal within the tunica muscularis.

Interstitial Cells: Regulators of Smooth Muscle Function

PubMed Central

Sanders, Kenton M.; Ward, Sean M.; Koh, Sang Don

2014-01-01

Smooth muscles are complex tissues containing a variety of cells in addition to muscle cells. Interstitial cells of mesenchymal origin interact with and form electrical connectivity with smooth muscle cells in many organs, and these cells provide important regulatory functions. For example, in the gastrointestinal tract, interstitial cells of Cajal (ICC) and PDGFRα+ cells have been described, in detail, and represent distinct classes of cells with unique ultrastructure, molecular phenotypes, and functions. Smooth muscle cells are electrically coupled to ICC and PDGFRα+ cells, forming an integrated unit called the SIP syncytium. SIP cells express a variety of receptors and ion channels, and conductance changes in any type of SIP cell affect the excitability and responses of the syncytium. SIP cells are known to provide pacemaker activity, propagation pathways for slow waves, transduction of inputs from motor neurons, and mechanosensitivity. Loss of interstitial cells has been associated with motor disorders of the gut. Interstitial cells are also found in a variety of other smooth muscles; however, in most cases, the physiological and pathophysiological roles for these cells have not been clearly defined. This review describes structural, functional, and molecular features of interstitial cells and discusses their contributions in determining the behaviors of smooth muscle tissues. PMID:24987007

Depolarizing Effects of Daikenchuto on Interstitial Cells of Cajal from Mouse Small Intestine

PubMed Central

Kim, Hyungwoo; Kim, Hyun Jung; Yang, Dongki; Jung, Myeong Ho; Kim, Byung Joo

2017-01-01

Background: Daikenchuto (DKT; TJ-100, TU-100), a traditional herbal medicineis used in modern medicine to treat gastrointestinal (GI) functional disorders. Interstitial cells of Cajal (ICCs) are the pacemaker cells of the GI tract and play important roles in the regulation of GI motility. Objective: The objective of this study was to investigate the effects of DKT on the pacemaker potentials (PPs) of cultured ICCs from murine small intestine. Materials and Methods: Enzymatic digestions were used to dissociate ICCs from mouse small intestine tissues. All experiments on ICCs were performed after 12 h of culture. The whole-cell patch-clamp configuration was used to record ICC PPs (current clamp mode). All experiments were performed at 30-32°C. Results: In current-clamp modeDKT depolarized and concentration-dependently decreased the amplitudes of PPs. Y25130 (a 5-HT3 receptor antagonist) or SB269970 (a 5-HT7 receptor antagonist) did not block DKT-induced PP depolarization, but RS39604 (a 5-HT4 receptor antagonist) did. Methoctramine (a muscarinic M2 receptor antagonist) failed to block DKT-induced PP depolarization, but pretreating 4-diphenylacetoxy-N-methylpiperidine methiodide (a muscarinic M3 receptor antagonist) facilitated blockade of DKT-induced PP depolarization. Pretreatment with an external Ca2+-free solution or thapsigargin abolished PPsand under these conditions, DKT did not induce PP depolarization. Furthermore Ginseng radix and Zingiberis rhizomes depolarized PPs, whereas Zanthoxyli fructus fruit (the third component of DKT) hyperpolarized PPs. Conclusion: These results suggest that DKT depolarizes ICC PPs in an internal or external Ca2+-dependent manner by stimulating 5-HT4 and M3 receptors. Furthermore, the authors suspect that the component in DKT largely responsible for depolarization is probably also a component of Ginseng radix and Zingiberis rhizomes. SUMMARY Daikenchuto (DKT) depolarized and concentration-dependently decreased the amplitudes of pacemaker potentials (PPs)Y25130 (a 5-HT3 receptor antagonist) or SB269970 (a 5-HT7 receptor antagonist) did not block DKT-induced PP depolarization, but RS39604 (a 5-HT4 receptor antagonist) didMethoctramine (a muscarinic M2 receptor antagonist) failed to block DKT-induced PP depolarization, but pretreating 4-DAMP (a muscarinic M3 receptor antagonist) facilitated blockade of DKT-induced PP depolarizationGinseng radix and Zingiberis rhizomes depolarized PPswhereas Zanthoxyli fructus fruit (the third component of DKT) hyperpolarized PPs. Abbreviation used: DKT: Daikenchuto, GI: Gastrointestinal, ICCs: Interstitial cells of Cajal, PPs: Pacemaker Potentials. PMID:28216898

Depolarizing Effects of Daikenchuto on Interstitial Cells of Cajal from Mouse Small Intestine.

PubMed

Kim, Hyungwoo; Kim, Hyun Jung; Yang, Dongki; Jung, Myeong Ho; Kim, Byung Joo

2017-01-01

Daikenchuto (DKT; TJ-100, TU-100), a traditional herbal medicineis used in modern medicine to treat gastrointestinal (GI) functional disorders. Interstitial cells of Cajal (ICCs) are the pacemaker cells of the GI tract and play important roles in the regulation of GI motility. The objective of this study was to investigate the effects of DKT on the pacemaker potentials (PPs) of cultured ICCs from murine small intestine. Enzymatic digestions were used to dissociate ICCs from mouse small intestine tissues. All experiments on ICCs were performed after 12 h of culture. The whole-cell patch-clamp configuration was used to record ICC PPs (current clamp mode). All experiments were performed at 30-32°C. In current-clamp modeDKT depolarized and concentration-dependently decreased the amplitudes of PPs. Y25130 (a 5-HT 3 receptor antagonist) or SB269970 (a 5-HT 7 receptor antagonist) did not block DKT-induced PP depolarization, but RS39604 (a 5-HT 4 receptor antagonist) did. Methoctramine (a muscarinic M 2 receptor antagonist) failed to block DKT-induced PP depolarization, but pretreating 4-diphenylacetoxy-N-methylpiperidine methiodide (a muscarinic M 3 receptor antagonist) facilitated blockade of DKT-induced PP depolarization. Pretreatment with an external Ca 2+ -free solution or thapsigargin abolished PPsand under these conditions, DKT did not induce PP depolarization. Furthermore Ginseng radix and Zingiberis rhizomes depolarized PPs, whereas Zanthoxyli fructus fruit (the third component of DKT) hyperpolarized PPs. These results suggest that DKT depolarizes ICC PPs in an internal or external Ca 2+ -dependent manner by stimulating 5-HT 4 and M 3 receptors. Furthermore, the authors suspect that the component in DKT largely responsible for depolarization is probably also a component of Ginseng radix and Zingiberis rhizomes. Daikenchuto (DKT) depolarized and concentration-dependently decreased the amplitudes of pacemaker potentials (PPs)Y25130 (a 5-HT 3 receptor antagonist) or SB269970 (a 5-HT 7 receptor antagonist) did not block DKT-induced PP depolarization, but RS39604 (a 5-HT 4 receptor antagonist) didMethoctramine (a muscarinic M 2 receptor antagonist) failed to block DKT-induced PP depolarization, but pretreating 4-DAMP (a muscarinic M 3 receptor antagonist) facilitated blockade of DKT-induced PP depolarizationGinseng radix and Zingiberis rhizomes depolarized PPswhereas Zanthoxyli fructus fruit (the third component of DKT) hyperpolarized PPs. Abbreviation used: DKT: Daikenchuto, GI: Gastrointestinal, ICCs: Interstitial cells of Cajal, PPs: Pacemaker Potentials.

Expression of profibrotic growth factors and their receptors by mouse lung macrophages and fibroblasts under conditions of acute viral inflammation in influenza A/H5N1 virus.

PubMed

Anikina, A G; Shkurupii, V A; Potapova, O V; Kovner, A V; Shestopalov, A M

2014-04-01

Morphological signs of early interstitial fibrosis, developing under conditions of acute viral inflammation (postinfection days 1-14), were observed in C57Bl/6 mice infected with influenza A/H5N1 A/goose/Krasnoozerskoye/627/05 virus. The development of fibrosis was confirmed by an increase in the number of lung cells expressing TNF-α. These changes were recorded in the presence of a many-fold increase in the counts of macrophages and fibroblasts expressing FGF, EGF, and their receptors.

Temporal Transcriptional Profiling of Somatic and Germ Cells Reveals Biased Lineage Priming of Sexual Fate in the Fetal Mouse Gonad

PubMed Central

Jameson, Samantha A.; Natarajan, Anirudh; Cool, Jonah; DeFalco, Tony; Maatouk, Danielle M.; Mork, Lindsey; Munger, Steven C.; Capel, Blanche

2012-01-01

The divergence of distinct cell populations from multipotent progenitors is poorly understood, particularly in vivo. The gonad is an ideal place to study this process, because it originates as a bipotential primordium where multiple distinct lineages acquire sex-specific fates as the organ differentiates as a testis or an ovary. To gain a more detailed understanding of the process of gonadal differentiation at the level of the individual cell populations, we conducted microarrays on sorted cells from XX and XY mouse gonads at three time points spanning the period when the gonadal cells transition from sexually undifferentiated progenitors to their respective sex-specific fates. We analyzed supporting cells, interstitial/stromal cells, germ cells, and endothelial cells. This work identified genes specifically depleted and enriched in each lineage as it underwent sex-specific differentiation. We determined that the sexually undifferentiated germ cell and supporting cell progenitors showed lineage priming. We found that germ cell progenitors were primed with a bias toward the male fate. In contrast, supporting cells were primed with a female bias, indicative of the robust repression program involved in the commitment to XY supporting cell fate. This study provides a molecular explanation reconciling the female default and balanced models of sex determination and represents a rich resource for the field. More importantly, it yields new insights into the mechanisms by which different cell types in a single organ adopt their respective fates. PMID:22438826

Maintenance of airway epithelium in acutely rejected orthotopic vascularized mouse lung transplants.

PubMed

Okazaki, Mikio; Gelman, Andrew E; Tietjens, Jeremy R; Ibricevic, Aida; Kornfeld, Christopher G; Huang, Howard J; Richardson, Steven B; Lai, Jiaming; Garbow, Joel R; Patterson, G Alexander; Krupnick, Alexander S; Brody, Steven L; Kreisel, Daniel

2007-12-01

Lung transplantation remains the only therapeutic option for many patients suffering from end-stage pulmonary disease. Long-term success after lung transplantation is severely limited by the development of bronchiolitis obliterans. The murine heterotopic tracheal transplantation model has been widely used for studies investigating pathogenesis of obliterative airway disease and immunosuppressive strategies to prevent its development. Despite its utility, this model employs proximal airway that lacks airflow and is not vascularized. We have developed a novel model of orthotopic vascularized lung transplantation in the mouse, which leads to severe vascular rejection in allogeneic strain combinations. Here we characterize differences in the fate of airway epithelial cells in nonimmunosuppressed heterotopic tracheal and vascularized lung allograft models over 28 days. Up-regulation of growth factors that are thought to be critical for the development of airway fibrosis and interstitial collagen deposition were similar in both models. However, while loss of airway epithelial cells occurred in the tracheal model, airway epithelium remained intact and fully differentiated in lung allografts, despite profound vascular rejection. Moreover, we demonstrate expression of the anti-apoptotic protein Bcl-2 in airway epithelial cells of acutely rejected lung allografts. These findings suggest that in addition to alloimmune responses, other stimuli may be required for the destruction of airway epithelial cells. Thus, the model of vascularized mouse lung transplantation may provide a new and more physiologic experimental tool to study the interaction between immune and nonimmune mechanisms affecting airway pathology in lung allografts.

Network properties of interstitial cells of Cajal affect intestinal pacemaker activity and motor patterns, according to a mathematical model of weakly coupled oscillators.

PubMed

Wei, Ruihan; Parsons, Sean P; Huizinga, Jan D

2017-03-01

What is the central question of this study? What are the effects of interstitial cells of Cajal (ICC) network perturbations on intestinal pacemaker activity and motor patterns? What is the main finding and its importance? Two-dimensional modelling of the ICC pacemaker activity according to a phase model of weakly coupled oscillators showed that network properties (coupling strength between oscillators, frequency gradient and frequency noise) strongly influence pacemaker network activity and subsequent motor patterns. The model explains motor patterns observed in physiological conditions and provides predictions and testable hypotheses for effects of ICC loss and frequency modulation on the motor patterns. Interstitial cells of Cajal (ICC) are the pacemaker cells of gut motility and are associated with motility disorders. Interstitial cells of Cajal form a network, but the contributions of its network properties to gut physiology and dysfunction are poorly understood. We modelled an ICC network as a two-dimensional network of weakly coupled oscillators with a frequency gradient and showed changes over time in video and graphical formats. Model parameters were obtained from slow-wave-driven contraction patterns in the mouse intestine and pacemaker slow-wave activities from the cat intestine. Marked changes in propagating oscillation patterns (including changes from propagation to non-propagating) were observed by changing network parameters (coupling strength between oscillators, the frequency gradient and frequency noise), which affected synchronization, propagation velocity and occurrence of dislocations (termination of an oscillation). Complete uncoupling of a circumferential ring of oscillators caused the proximal and distal section to desynchronize, but complete synchronization was maintained with only a single oscillator connecting the sections with high enough coupling. The network of oscillators could withstand loss; even with 40% of oscillators lost randomly within the network, significant synchronization and anterograde propagation remained. A local increase in pacemaker frequency diminished anterograde propagation; the effects were strongly dependent on location, frequency gradient and coupling strength. In summary, the model puts forth the hypothesis that fundamental changes in oscillation patterns (ICC slow-wave activity or circular muscle contractions) can occur through physiological modulation of network properties. Strong evidence is provided to accept the ICC network as a system of coupled oscillators. © 2016 The Authors. Experimental Physiology © 2016 The Physiological Society.

The Effects of Lactobacillus acidophilus on the Intestinal Smooth Muscle Contraction through PKC/MLCK/MLC Signaling Pathway in TBI Mouse Model

PubMed Central

Fang, Huan; Zhu, Lina; Gao, Ning; Zhu, Jingci

2015-01-01

Clinical studies have shown that probiotics influence gastrointestinal motility. However, the molecular mechanisms by which probiotic Lactobacillus modulates intestinal motility in traumatic brain injury (TBI) mouse model have not been explored. In the present study, we provided evidence showing that treatment of TBI mice with Lactobacillus acidophilus significantly improved the terminal ileum villus morphology, restored the impaired interstitial cells of Cajal (ICC) and the disrupted ICC networks after TBI, and prevented TBI-mediated inhibition of contractile activity in intestinal smooth muscle. Mechanistically, the decreased concentration of MLCK, phospho-MLC20 and phospho-MYPT1 and increased concentration of MLCP and PKC were observed after TBI, and these events mediated by TBI were efficiently prevented by Lactobacillus acidophilus application. These findings may provide a novel mechanistic basis for the application of Lactobacillus acidophilus in the treatment of TBI. PMID:26030918

The Effects of Lactobacillus acidophilus on the Intestinal Smooth Muscle Contraction through PKC/MLCK/MLC Signaling Pathway in TBI Mouse Model.

PubMed

Sun, Bo; Hu, Chen; Fang, Huan; Zhu, Lina; Gao, Ning; Zhu, Jingci

2015-01-01

Clinical studies have shown that probiotics influence gastrointestinal motility. However, the molecular mechanisms by which probiotic Lactobacillus modulates intestinal motility in traumatic brain injury (TBI) mouse model have not been explored. In the present study, we provided evidence showing that treatment of TBI mice with Lactobacillus acidophilus significantly improved the terminal ileum villus morphology, restored the impaired interstitial cells of Cajal (ICC) and the disrupted ICC networks after TBI, and prevented TBI-mediated inhibition of contractile activity in intestinal smooth muscle. Mechanistically, the decreased concentration of MLCK, phospho-MLC20 and phospho-MYPT1 and increased concentration of MLCP and PKC were observed after TBI, and these events mediated by TBI were efficiently prevented by Lactobacillus acidophilus application. These findings may provide a novel mechanistic basis for the application of Lactobacillus acidophilus in the treatment of TBI.

Spatial expression of components of a calcitonin receptor-like receptor (CRL) signalling system (CRL, calcitonin gene-related peptide, adrenomedullin, adrenomedullin-2/intermedin) in mouse and human heart valves.

PubMed

Pfeil, Uwe; Bharathala, Subhashini; Murtaza, Ghulam; Mermer, Petra; Papadakis, Tamara; Boening, Andreas; Kummer, Wolfgang

2016-12-01

Heart valves are highly organized structures determining the direction of blood flow through the heart. Smooth muscle cells within the valve are thought to play an active role during the heart cycle, rather than being just passive flaps. The mature heart valve is composed of extracellular matrix (ECM), various differentiations of valvular interstitial cells (VIC), smooth muscle cells and overlying endothelium. VIC are important for maintaining the structural integrity of the valve, thereby affecting valve function and ECM remodelling. Accumulating evidence suggests an important role of calcitonin receptor-like receptor (CRL) signalling in preventing heart damage under several pathological conditions. Thus we investigate the existence of a putative CRL signalling system in mouse and human heart valves by real-time RT-PCR, laser-assisted microdissection, immunofluorescence and NADPH-diaphorase histochemistry. Mouse and human heart valves expressed mRNAs for the CRL ligands adrenomedullin (AM), adrenomedullin-2 (AM-2) and calcitonin gene-related peptide (CGRP) and for their receptor components, i.e., CRL and receptor-activity-modifying proteins 1-3. Immunofluorescence analysis revealed AM-, AM-2- and CRL-immunolabelling in endothelial cells and VIC, whereas CGRP immunoreactivity was restricted to nerve fibres and some endothelial cells. Nitric oxide synthase activity, as demonstrated by NADPH-diaphorase histochemistry, was shown mainly in valvular endothelial cells in mice, whereas in human aortic valves, VIC and smooth muscle cells were positive. Our results showed the presence of an intrinsic AM/AM-2/CGRP signalling system in murine and human heart valves with distinct cellular localization, suggesting its involvement in the regulation of valve stiffness and ECM production and turnover.

Effects of aqueous extracts of "Betel quid" and its constituents on testosterone production by dispersed mouse interstitial cells.

PubMed

Yang, Nai-Yen Jack; Kaphle, Krishna; Wang, Pei-Hwa; Jong, De-Shien; Wu, Leang-Shin; Lin, Jen-Hsou

2004-01-01

Betel quid (BQ) is a favorite chewing item among many communities in different parts of Asia where it is popular by different names. BQ is a unique combination of nut or fruit from the Areca catechu Linn. (AN) tree, leaf from the Piper betle Linn. (BL) vine, slaked lime, paste of bark from the Acacia catechu tree and other spices. AN has been used successfully in various traditional medicines by different civilizations over several ages. Initially condemned by the medical communities for its health hazards, identification and application of potent pharmacologically bioactive compounds from different constituents of BQ have rekindled growing interest in related investigations. Curious about the stimulating role of BQ, we investigated the potential steroidogenic activity of hot water extract from BQ and its constituents and arecoline on testosterone producing ability in an in vitro experiment. Enzyme dissociated interstitial cells from adult mouse testes (ICR strain) were cultured with/without different doses of the extracts and the level of testosterone produced was assayed by an enzyme immunoassay (EIA) technique. It was found that at lower doses of arecoline, AN and BL extracts had significantly stimulated testosterone production over the basal level (p < 0.05). BQ extract, on the other hand, did not show any significant effect on testosterone production. Combinations of arecoline at low doses with 10 ng/ml ovine leutinizing hormone (oLH) showed increases in testosterone produced, while cyclic adenosine monophosphate (cAMP) co-culture showed dose-related inhibition. Our current finding hints at the possible dose-dependent dualistic role of AN and BL extracts and arecoline for testosterone production employing possible non-cAMP-dependent pathway of steroidogenesis. However, the identity of the active compounds besides arecoline and the exact mechanism involved remains to be further investigated.

Reduced interstitial cells of Cajal and increased intraepithelial lymphocytes are associated with development of small intestinal bacterial overgrowth in post-infectious IBS mouse model.

PubMed

Chen, Binrui; Zhu, Shuwen; Du, Lijun; He, Huiqin; Kim, John J; Dai, Ning

2017-10-01

Intestinal dysmotility and immune activation are likely involved in the pathogenesis of small intestinal bacteria overgrowth (SIBO) in irritable bowel syndrome (IBS). We aimed at investigating the role of interstitial cells of Cajal (ICC) and intestinal inflammation in the development of SIBO using a post-infectious IBS (PI-IBS) mouse model. NIH mice were randomly infected with Trichinella spiralis. Visceral sensitivity and stool pattern were assessed at 8-weeks post-infection (PI). Intestinal bacteria counts from jejunum and ileum were measured by quantitative real-time PCR to evaluate the presence of SIBO. ICC density, intraepithelial lymphocytes (IELs) counts, and intestinal cytokine levels (IL1-β, IL-6, toll-like receptor-4 (TLR-4), IL-10) in the ileum were examined. PI-IBS mice demonstrated increased visceral sensitivity compared with the control group. One-third of the PI-IBS mice developed SIBO (SIBO+/PI-IBS) and was more likely to have abnormal stool form compared with SIBO negative PI-IBS (SIBO-/PI-IBS) mice but without difference in visceral sensitivity. SIBO+/PI-IBS mice had decreased ICC density and increased IELs counts in the ileum compared with SIBO-/PI-IBS mice. No difference in inflammatory cytokine expression levels were detected among the groups except for increased TLR-4 in PI-IBS mice compared with the control group. Development of SIBO in PI-IBS mice was associated with reduced ICC density and increased IELs counts in the ileum. Our findings support the role of intestinal dysmotility and inflammation in the pathogenesis of SIBO in IBS and may provide potential therapeutic targets.

Telomeres and mechanisms of Robertsonian fusion.

PubMed

Slijepcevic, P

1998-05-01

The Robertsonian (Rb) fusion, a chromosome rearrangement involving centric fusion of two acro-(telo)centric chromosomes to form a single metacentric, is one of the most frequent events in mammalian karyotype evolution. Since one of the functions of telomeres is to preserve chromosome integrity, a prerequisite for the formation of Rb fusions should be either telomere loss or telomere inactivation. Possible mechanisms underlying the formation of various types of Rb fusion are discussed here. For example, Rb fusion in wild mice involves complete loss of p-arm telomeres by chromosome breakage within minor satellite sequences. By contrast, interstitial telomeric sites are found in the pericentromeric regions of chromosomes originating from a number of vertebrate species, suggesting the occurrence of Rb-like fusion without loss of telomeres, a possibility consistent with some form of telomere inactivation. Finally, a recent study suggests that telomere shortening induced by the deletion of the telomerase RNA gene in the mouse germ-line leads to telomere loss and high frequencies of Rb fusion in mouse somatic cells. Thus, at least three mechanisms in mammalian cells lead to the formation of Rb fusions.

Induced Autologous Stem Cell Transplantation for Treatment of Rabbit Renal Interstitial Fibrosis

PubMed Central

Ruan, Guang-Ping; Xu, Fan; Li, Zi-An; Zhu, Guang-Xu; Pang, Rong-Qing; Wang, Jin-Xiang; Cai, Xue-Min; He, Jie; Yao, Xiang; Ruan, Guang-Hong; Xu, Xin-Ming; Pan, Xing-Hua

2013-01-01

Introduction Renal interstitial fibrosis (RIF) is a significant cause of end-stage renal failure. The goal of this study was to characterize the distribution of transplanted induced autologous stem cells in a rabbit model of renal interstitial fibrosis and evaluate its therapeutic efficacy for treatment of renal interstitial fibrosis. Methods A rabbit model of renal interstitial fibrosis was established. Autologous fibroblasts were cultured, induced and labeled with green fluorescent protein (GFP). These labeled stem cells were transplanted into the renal artery of model animals at 8 weeks. Results Eight weeks following transplantation of induced autologous stem cells, significant reductions (P < 0.05) were observed in serum creatinine (SCr) (14.8 ± 1.9 mmol/L to 10.1 ± 2.1 mmol/L) and blood urea nitrogen (BUN) (119 ± 22 µmol/L to 97 ± 13 µmol/L), indicating improvement in renal function. Conclusions We successfully established a rabbit model of renal interstitial fibrosis and demonstrated that transplantation of induced autologous stem cells can repair kidney damage within 8 weeks. The repair occurred by both inhibition of further development of renal interstitial fibrosis and partial reversal of pre-existing renal interstitial fibrosis. These beneficial effects lead to the development of normal tissue structure and improved renal function. PMID:24367598

[Effect of rapamycin on proliferation of rat heart valve interstitial cells in vitro].

PubMed

Tan, Yan; Wang, Ji-Ye; Yi, Ren-Liang; Qiu, Jian

2016-04-01

To investigate the effect of rapamycin on the proliferation of rat valvular interstitial cells in primary culture. The interstitial cells isolated from rat aortic valves were cultured and treated with rapamycin, and the cell growth and cell cycle changes were analyzed using MTT assay and flow cytometry, respectively. RT-PCR was used to detect mRNA expression levels of S6 and P70S6K in cells, and the protein expressions level of S6, P70S6K, P-S6, and P-P70S6K were detected using Western blotting. Rat aortic valvular interstitial cells was isolated successfully. The rapamycin-treated cells showed a suppressed proliferative activity (P<0.05), but the cell cycle distribution remained unaffected. Rapamycin treatment resulted in significantly decreased S6 and P70S6K protein phosphorylation level in the cells (P<0.05). The mechanism by which rapamycin inhibits the proliferation of valvular interstitial cells probably involves suppression of mTOR to lower S6 and P70S6K phosphorylation level but not direct regulation of the cell cycle.

Mast cells play no role in the pathogenesis of postoperative ileus induced by intestinal manipulation.

PubMed

Gomez-Pinilla, Pedro J; Farro, Giovanna; Di Giovangiulio, Martina; Stakenborg, Nathalie; Némethova, Andrea; de Vries, Annick; Liston, Adrian; Feyerabend, Thorsten B; Rodewald, Hans-Reimer; Rodewald, Hans-Reimwer; Boeckxstaens, Guy E; Matteoli, Gianluca

2014-01-01

Intestinal manipulation (IM) during abdominal surgery results in intestinal inflammation leading to hypomotility or ileus. Mast cell activation is thought to play a crucial role in the pathophysiology of postoperative ileus (POI). However, this conclusion was mainly drawn using mast cell-deficient mouse models with abnormal Kit signaling. These mice also lack interstitial cells of Cajal (ICC) resulting in aberrant gastrointestinal motility even prior to surgery, compromising their use as model to study POI. To avoid these experimental weaknesses we took advantage of a newly developed knock-in mouse model, Cpa3(Cre/+) , devoid of mast cells but with intact Kit signaling. The role of mast cells in the development of POI and intestinal inflammation was evaluated assessing gastrointestinal transit and muscularis externa inflammation after IM in two strains of mice lacking mast cells, i.e. Kit(W-sh/W-sh) and Cpa3(Cre/+) mice, and by use of the mast cell stabilizer cromolyn. Kit(W-sh/W-sh) mice lack ICC networks and already revealed significantly delayed gastrointestinal transit even before surgery. IM did not further delay intestinal transit, but induced infiltration of myeloperoxidase positive cells, expression of inflammatory cytokines and recruitment of monocytes and neutrophils into the muscularis externa. On the contrary, Cpa3(Cre/+) mice have a normal network of ICC and normal gastrointestinal. Surprisingly, IM in Cpa3(Cre/+) mice caused delay in gut motility and intestinal inflammation as in wild type littermates mice (Cpa3(+/+) ). Furthermore, treatment with the mast cell inhibitor cromolyn resulted in an inhibition of mast cells without preventing POI. Here, we confirm that IM induced mast cell degranulation. However, our data demonstrate that mast cells are not required for the pathogenesis of POI in mice. Although there might be species differences between mouse and human, our results argue against mast cell inhibitors as a therapeutic approach to shorten POI.

Mast Cells Play No Role in the Pathogenesis of Postoperative Ileus Induced by Intestinal Manipulation

PubMed Central

Gomez-Pinilla, Pedro J.; Farro, Giovanna; Di Giovangiulio, Martina; Stakenborg, Nathalie; Némethova, Andrea; de Vries, Annick; Liston, Adrian; Feyerabend, Thorsten B.; Rodewald, Hans-Reimwer; Boeckxstaens, Guy E.; Matteoli, Gianluca

2014-01-01

Introduction Intestinal manipulation (IM) during abdominal surgery results in intestinal inflammation leading to hypomotility or ileus. Mast cell activation is thought to play a crucial role in the pathophysiology of postoperative ileus (POI). However, this conclusion was mainly drawn using mast cell-deficient mouse models with abnormal Kit signaling. These mice also lack interstitial cells of Cajal (ICC) resulting in aberrant gastrointestinal motility even prior to surgery, compromising their use as model to study POI. To avoid these experimental weaknesses we took advantage of a newly developed knock-in mouse model, Cpa3Cre/+, devoid of mast cells but with intact Kit signaling. Design The role of mast cells in the development of POI and intestinal inflammation was evaluated assessing gastrointestinal transit and muscularis externa inflammation after IM in two strains of mice lacking mast cells, i.e. KitW-sh/W-sh and Cpa3Cre/+ mice, and by use of the mast cell stabilizer cromolyn. Results KitW-sh/W-sh mice lack ICC networks and already revealed significantly delayed gastrointestinal transit even before surgery. IM did not further delay intestinal transit, but induced infiltration of myeloperoxidase positive cells, expression of inflammatory cytokines and recruitment of monocytes and neutrophils into the muscularis externa. On the contrary, Cpa3Cre/+ mice have a normal network of ICC and normal gastrointestinal. Surprisingly, IM in Cpa3Cre/+ mice caused delay in gut motility and intestinal inflammation as in wild type littermates mice (Cpa3+/+). Furthermore, treatment with the mast cell inhibitor cromolyn resulted in an inhibition of mast cells without preventing POI. Conclusions Here, we confirm that IM induced mast cell degranulation. However, our data demonstrate that mast cells are not required for the pathogenesis of POI in mice. Although there might be species differences between mouse and human, our results argue against mast cell inhibitors as a therapeutic approach to shorten POI. PMID:24416383

The Significance of Interstitial Cells in Neurogastroenterology

PubMed Central

Blair, Peter J; Rhee, Poong-Lyul; Sanders, Kenton M; Ward, Sean M

2014-01-01

Smooth muscle layers of the gastrointestinal tract consist of a heterogeneous population of cells that include enteric neurons, several classes of interstitial cells of mesenchymal origin, a variety of immune cells and smooth muscle cells (SMCs). Over the last number of years the complexity of the interactions between these cell types has begun to emerge. For example, interstitial cells, consisting of both interstitial cells of Cajal (ICC) and platelet-derived growth factor receptor alpha-positive (PDGFRα+) cells generate pacemaker activity throughout the gastrointestinal (GI) tract and also transduce enteric motor nerve signals and mechanosensitivity to adjacent SMCs. ICC and PDGFRα+ cells are electrically coupled to SMCs possibly via gap junctions forming a multicellular functional syncytium termed the SIP syncytium. Cells that make up the SIP syncytium are highly specialized containing unique receptors, ion channels and intracellular signaling pathways that regulate the excitability of GI muscles. The unique role of these cells in coordinating GI motility is evident by the altered motility patterns in animal models where interstitial cell networks are disrupted. Although considerable advances have been made in recent years on our understanding of the roles of these cells within the SIP syncytium, the full physiological functions of these cells and the consequences of their disruption in GI muscles have not been clearly defined. This review gives a synopsis of the history of interstitial cell discovery and highlights recent advances in structural, molecular expression and functional roles of these cells in the GI tract. PMID:24948131

Renal Impairment with Sublethal Tubular Cell Injury in a Chronic Liver Disease Mouse Model

PubMed Central

Ishida, Tokiko; Kotani, Hirokazu; Miyao, Masashi; Kawai, Chihiro; Jemail, Leila; Abiru, Hitoshi; Tamaki, Keiji

2016-01-01

The pathogenesis of renal impairment in chronic liver diseases (CLDs) has been primarily studied in the advanced stages of hepatic injury. Meanwhile, the pathology of renal impairment in the early phase of CLDs is poorly understood, and animal models to elucidate its mechanisms are needed. Thus, we investigated whether an existing mouse model of CLD induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) shows renal impairment in the early phase. Renal injury markers, renal histology (including immunohistochemistry for tubular injury markers and transmission electron microscopy), autophagy, and oxidative stress were studied longitudinally in DDC- and standard diet–fed BALB/c mice. Slight but significant renal dysfunction was evident in DDC-fed mice from the early phase. Meanwhile, histological examinations of the kidneys with routine light microscopy did not show definitive morphological findings, and electron microscopic analyses were required to detect limited injuries such as loss of brush border microvilli and mitochondrial deformities. Limited injuries have been recently designated as sublethal tubular cell injury. As humans with renal impairment, either with or without CLD, often show almost normal tubules, sublethal injury has been of particular interest. In this study, the injuries were associated with mitochondrial aberrations and oxidative stress, a possible mechanism for sublethal injury. Intriguingly, two defense mechanisms were associated with this injury that prevent it from progressing to apparent cell death: autophagy and single-cell extrusion with regeneration. Furthermore, the renal impairment of this model progressed to chronic kidney disease with interstitial fibrosis after long-term DDC feeding. These findings indicated that DDC induces renal impairment with sublethal tubular cell injury from the early phase, leading to chronic kidney disease. Importantly, this CLD mouse model could be useful for studying the pathophysiological mechanisms of sublethal tubular cell injury. PMID:26752420

The deep muscular plexus of the pig duodenum: a histochemical and ultrastructural study with special reference to the interstitial cells.

PubMed

Henry, M; Porcher, C; Julé, Y

1998-06-10

The aim of the present study was to describe the deep muscular plexus of the pig duodenum and to characterize its cellular components. Numerous nerve varicosities have been detected in the deep muscular plexus using anti-synaptophysin antibodies. Nerve fibres were also detected here in the outer circular muscle layer, whereas no nerve fibres were observed in the inner circular muscle layer. In the deep muscular plexus, nerve fibres projected to interstitial cells which were characterized at the ultrastructural level. The interstitial cells were of two kinds: the interstitial fibroblastic-like cells (FLC) and the interstitial dense cells (IDC), both of which were interposed between nerve fibres and smooth muscle cells. The FLC were characterized by their elongated bipolar shape, the lack of basal lamina, a well-developed endoplasmic reticulum, a Golgi apparatus, and intermediate filaments. They were closely apposed to axon terminals containing small clear synaptic vesicles and/or dense-cored vesicles. They were frequently connected to each other and to smooth muscle cells of the inner and outer circular layer by desmosomes and more rarely by gap junctions. The IDC are myoid-like cells. They had a stellate appearance and were characterized by a dense cell body, numerous caveolae, and a discontinuous basal lamina. The IDC were always closely apposed to nerve fibres and were connected to smooth muscle cells by desmosomes and small gap junctions. The present results show the unique pattern of cellular organization of the deep muscular plexus of the pig small intestine. They suggest that the interstitial cells in the deep muscular plexus are involved in the integration and transmission of nervous inputs from myenteric neurons to the inner and outer circular muscle layers. The clear-cut distinction observed here between the two types of interstitial cells (fibroblastic and myoid-like) suggests that the interstitial cells of each type may also be involved in some other specific activity, which still remains to be determined.

Distinct subpopulations of FOXD1 stroma-derived cells regulate renal erythropoietin

PubMed Central

Liu, Qingdu; Binns, Thomas C.; Davidoff, Olena; Kapitsinou, Pinelopi P.; Pfaff, Andrew S.; Olauson, Hannes; Fogo, Agnes B.; Fong, Guo-Hua; Gross, Kenneth W.

2016-01-01

Renal peritubular interstitial fibroblast-like cells are critical for adult erythropoiesis, as they are the main source of erythropoietin (EPO). Hypoxia-inducible factor 2 (HIF-2) controls EPO synthesis in the kidney and liver and is regulated by prolyl-4-hydroxylase domain (PHD) dioxygenases PHD1, PHD2, and PHD3, which function as cellular oxygen sensors. Renal interstitial cells with EPO-producing capacity are poorly characterized, and the role of the PHD/HIF-2 axis in renal EPO-producing cell (REPC) plasticity is unclear. Here we targeted the PHD/HIF-2/EPO axis in FOXD1 stroma-derived renal interstitial cells and examined the role of individual PHDs in REPC pool size regulation and renal EPO output. Renal interstitial cells with EPO-producing capacity were entirely derived from FOXD1-expressing stroma, and Phd2 inactivation alone induced renal Epo in a limited number of renal interstitial cells. EPO induction was submaximal, as hypoxia or pharmacologic PHD inhibition further increased the REPC fraction among Phd2–/– renal interstitial cells. Moreover, Phd1 and Phd3 were differentially expressed in renal interstitium, and heterozygous deficiency for Phd1 and Phd3 increased REPC numbers in Phd2–/– mice. We propose that FOXD1 lineage renal interstitial cells consist of distinct subpopulations that differ in their responsiveness to Phd2 inactivation and thus regulation of HIF-2 activity and EPO production under hypoxia or conditions of pharmacologic or genetic PHD inactivation. PMID:27088801

Channel-mediated lactate release by K⁺-stimulated astrocytes.

PubMed

Sotelo-Hitschfeld, Tamara; Niemeyer, María I; Mächler, Philipp; Ruminot, Iván; Lerchundi, Rodrigo; Wyss, Matthias T; Stobart, Jillian; Fernández-Moncada, Ignacio; Valdebenito, Rocío; Garrido-Gerter, Pamela; Contreras-Baeza, Yasna; Schneider, Bernard L; Aebischer, Patrick; Lengacher, Sylvain; San Martín, Alejandro; Le Douce, Juliette; Bonvento, Gilles; Magistretti, Pierre J; Sepúlveda, Francisco V; Weber, Bruno; Barros, L Felipe

2015-03-11

Excitatory synaptic transmission is accompanied by a local surge in interstitial lactate that occurs despite adequate oxygen availability, a puzzling phenomenon termed aerobic glycolysis. In addition to its role as an energy substrate, recent studies have shown that lactate modulates neuronal excitability acting through various targets, including NMDA receptors and G-protein-coupled receptors specific for lactate, but little is known about the cellular and molecular mechanisms responsible for the increase in interstitial lactate. Using a panel of genetically encoded fluorescence nanosensors for energy metabolites, we show here that mouse astrocytes in culture, in cortical slices, and in vivo maintain a steady-state reservoir of lactate. The reservoir was released to the extracellular space immediately after exposure of astrocytes to a physiological rise in extracellular K(+) or cell depolarization. Cell-attached patch-clamp analysis of cultured astrocytes revealed a 37 pS lactate-permeable ion channel activated by cell depolarization. The channel was modulated by lactate itself, resulting in a positive feedback loop for lactate release. A rapid fall in intracellular lactate levels was also observed in cortical astrocytes of anesthetized mice in response to local field stimulation. The existence of an astrocytic lactate reservoir and its quick mobilization via an ion channel in response to a neuronal cue provides fresh support to lactate roles in neuronal fueling and in gliotransmission. Copyright © 2015 the authors 0270-6474/15/354168-11$15.00/0.

A Knock-in Mouse Model of Human PHD2 Gene-associated Erythrocytosis Establishes a Haploinsufficiency Mechanism*

PubMed Central

Arsenault, Patrick R.; Pei, Fei; Lee, Rebecca; Kerestes, Heddy; Percy, Melanie J.; Keith, Brian; Simon, M. Celeste; Lappin, Terence R. J.; Khurana, Tejvir S.; Lee, Frank S.

2013-01-01

The central pathway for controlling red cell mass is the PHD (prolyl hydroxylase domain protein):hypoxia-inducible factor (HIF) pathway. HIF, which is negatively regulated by PHD, activates numerous genes, including ones involved in erythropoiesis, such as the ERYTHROPOIETIN (EPO) gene. Recent studies have implicated PHD2 as the key PHD isoform regulating red cell mass. Studies of humans have identified erythrocytosis-associated, heterozygous point mutations in the PHD2 gene. A key question concerns the mechanism by which human mutations lead to phenotypes. In the present report, we generated and characterized a mouse line in which a P294R knock-in mutation has been introduced into the mouse Phd2 locus to model the first reported human PHD2 mutation (P317R). Phd2P294R/+ mice display a degree of erythrocytosis equivalent to that seen in Phd2+/− mice. The Phd2P294R/+-associated erythrocytosis is reversed in a Hif2a+/−, but not a Hif1a+/− background. Additional studies using various conditional knock-outs of Phd2 reveal that erythrocytosis can be induced by homozygous and heterozygous knock-out of Phd2 in renal cortical interstitial cells using a Pax3-Cre transgene or by homozygous knock-out of Phd2 in hematopoietic progenitors driven by a Vav1-Cre transgene. These studies formally prove that a missense mutation in PHD2 is the cause of the erythrocytosis, show that this occurs through haploinsufficiency, and point to multifactorial control of red cell mass by PHD2. PMID:24121508

Telocytes and stem cells in limbus and uvea of mouse eye

PubMed Central

Luesma, María José; Gherghiceanu, Mihaela; Popescu, Laurenţiu M

2013-01-01

The potential of stem cell (SC) therapies for eye diseases is well-recognized. However, the results remain only encouraging as little is known about the mechanisms responsible for eye renewal, regeneration and/or repair. Therefore, it is critical to gain knowledge about the specific tissue environment (niches) where the stem/progenitor cells reside in eye. A new type of interstitial cell–telocyte (TC) (http://www.telocytes.com) was recently identified by electron microscopy (EM). TCs have very long (tens of micrometres) and thin (below 200 nm) prolongations named telopodes (Tp) that form heterocellular networks in which SCs are embedded. We found TCs by EM and electron tomography in sclera, limbus and uvea of the mouse eye. Furthermore, EM showed that SCs were present in the anterior layer of the iris and limbus. Adhaerens and gap junctions were found to connect TCs within a network in uvea and sclera. Nanocontacts (electron-dense structures) were observed between TCs and other cells: SCs, melanocytes, nerve endings and macrophages. These intercellular ‘feet’ bridged the intercellular clefts (about 10 nm wide). Moreover, exosomes (extracellular vesicles with a diameter up to 100 nm) were delivered by TCs to other cells of the iris stroma. The ultrastructural nanocontacts of TCs with SCs and the TCs paracrine influence via exosomes in the epithelial and stromal SC niches suggest an important participation of TCs in eye regeneration. PMID:23991685

Histochemical and cellular changes accompanying the appearance of lung fibrosis in an experimental mouse model for Hermansky Pudlak syndrome

PubMed Central

Lyerla, Timothy

2010-01-01

Hermansky Pudlak syndrome (HPS) is a heterogeneous recessive genetic disease with a tendency to develop lung fibrosis with aging. A mouse strain with two mutant HPS genes affecting separate vesicle trafficking pathways, C57BL/6-Hps1ep-Ap3b1pe, exhibits severe lung abnormalities at young ages, including enlarged alveolar type II (ATII) cells with giant lamellar bodies and foamy alveolar macrophages (AMs), which are readily identified histologically. In this study, the appearance of lung fibrosis in older animals was studied using classical histological and biochemical methods. The HPS double mutant mice, but not Chediak Higashi syndrome (C57BL/6-Lystbg-J-J, CHS) or C57BL/6J black control (WT) mice, were found to develop lung fibrosis at about 17 months of age using Masson trichrome staining, which was confirmed by hydroxyproline analysis. TGF β1 levels were elevated in bronchial alveolar lavage samples at all ages tested in the double mutant, but not WT or CHS mice, indicative of a prefibrotic condition in this experimental strain; and AMs were highly positive for this cytokine using immunohistochemistry staining. Prosurfactant protein C staining for ATII cells showed redistribution and dysmorphism of these cells with aging, but there was no evidence for epithelial-mesenchymal transition of ATII cells by dual staining for prosurfactant C protein and α-smooth muscle actin. This investigation showed that the HPS double mutant mouse strain develops interstitial pneumonia (HPSIP) past 1 year of age, which may be initiated by abnormal ATII cells and exacerbated by AM activation. With prominent prefibrotic abnormalities, this double mutant may serve as a model for interventive therapy in HPS. PMID:20603711

Innervation of the anterior byssal retractor muscle in Mytilus edulis L. II. Ultrastructure of the glio-interstitial cells.

PubMed

Gilloteaux, J

1975-08-27

Studies on the intrinsic innervation of the anterior byssal retractor muscle (ABRM) in Mytilus edulis L. were continued at the ultrastructural level. Electron micrographs show nerve processes ensheathed by glio-interstitial cells running between muscle fibers. The glio-interstitial cells may represent all the types of osmiophilic cells previously described by the light microscopic ZIO technique in the anterior byssal retractor muscle.

Dermal Collagen and Lipid Deposition Correlate with Tissue Swelling and Hydraulic Conductivity in Murine Primary Lymphedema

PubMed Central

Rutkowski, Joseph M.; Markhus, Carl Erik; Gyenge, Christina C.; Alitalo, Kari; Wiig, Helge; Swartz, Melody A.

2010-01-01

Primary lymphedema is a congenital pathology of dysfunctional lymphatic drainage characterized by swelling of the limbs, thickening of the dermis, and fluid and lipid accumulation in the underlying tissue. Two mouse models of primary lymphedema, the Chy mouse and the K14-VEGFR-3-Ig mouse, both lack dermal lymphatic capillaries and exhibit a lymphedematous phenotype attributable to disrupted VEGFR-3 signaling. Here we show that the differences in edematous tissue composition between these two models correlated with drastic differences in hydraulic conductivity. The skin of Chy mice possessed significantly higher levels of collagen and fat, whereas K14-VEGFR-3-Ig mouse skin composition was relatively normal, as compared with their respective wild-type controls. Functionally, this resulted in a greatly increased dermal hydraulic conductivity in K14-VEGFR3-Ig, but not Chy, mice. Our data suggest that lymphedema associated with increased collagen and lipid accumulation counteracts an increased hydraulic conductivity associated with dermal swelling, which in turn further limits interstitial transport and swelling. Without lipid and collagen accumulation, hydraulic conductivity is increased and overall swelling is minimized. These opposing tissue responses to primary lymphedema imply that tissue remodeling—predominantly collagen and fat deposition—may dictate tissue swelling and govern interstitial transport in lymphedema. PMID:20110415

Dermal collagen and lipid deposition correlate with tissue swelling and hydraulic conductivity in murine primary lymphedema.

PubMed

Rutkowski, Joseph M; Markhus, Carl Erik; Gyenge, Christina C; Alitalo, Kari; Wiig, Helge; Swartz, Melody A

2010-03-01

Primary lymphedema is a congenital pathology of dysfunctional lymphatic drainage characterized by swelling of the limbs, thickening of the dermis, and fluid and lipid accumulation in the underlying tissue. Two mouse models of primary lymphedema, the Chy mouse and the K14-VEGFR-3-Ig mouse, both lack dermal lymphatic capillaries and exhibit a lymphedematous phenotype attributable to disrupted VEGFR-3 signaling. Here we show that the differences in edematous tissue composition between these two models correlated with drastic differences in hydraulic conductivity. The skin of Chy mice possessed significantly higher levels of collagen and fat, whereas K14-VEGFR-3-Ig mouse skin composition was relatively normal, as compared with their respective wild-type controls. Functionally, this resulted in a greatly increased dermal hydraulic conductivity in K14-VEGFR3-Ig, but not Chy, mice. Our data suggest that lymphedema associated with increased collagen and lipid accumulation counteracts an increased hydraulic conductivity associated with dermal swelling, which in turn further limits interstitial transport and swelling. Without lipid and collagen accumulation, hydraulic conductivity is increased and overall swelling is minimized. These opposing tissue responses to primary lymphedema imply that tissue remodeling--predominantly collagen and fat deposition--may dictate tissue swelling and govern interstitial transport in lymphedema.

Immunephenotype of glomerular and interstitial infiltrating cells in protocol renal allograft biopsies and histological diagnosis.

PubMed

Moreso, F; Seron, D; O'Valle, F; Ibernon, M; Gomà, M; Hueso, M; Cruzado, J M; Bestard, O; Duarte, V; del Moral, R García; Grinyó, J M

2007-12-01

Patients with a protocol renal allograft biopsy simultaneously displaying interstitial fibrosis/tubular atrophy (IF/TA) and subclinical rejection (SCR) have a shortened graft survival than patients with a normal biopsy, or with a biopsy only displaying IF/TA or SCR. The poor outcome of these patients could be related with a more severe inflammation. We evaluate the immunophenotype of infiltrating cells in these diagnostic categories. Nonexhausted paraffin blocks from protocol biopsies done during the first year were stained with anti-CD45, CD3, CD20, CD68 and CD15 monoclonal antibodies. Glomerular and interstitial positive cells were counted. C4d deposition in peritubular capillaries was evaluated. Histological diagnoses were: normal (n = 80), SCR (n = 17), IF/TA (n = 42) and IF/TA + SCR (n = 17). Only interstitial CD20 positive cells were significantly increased in patients displaying IF/TA + SCR; normal (137 +/- 117), SCR (202 +/- 145), IF/TA (208 +/- 151) and IF/TA + SCR (307 +/- 180 cells/mm(2)), p < 0.01. The proportion of biopsies displaying C4d deposition was not different among groups. The upper tertile of CD20 positive interstitial cells was associated with a decreased death-censored graft survival (relative risk: 3.01, 95% confidence interval: 1.23-7.35; p = 0.015). These data suggest that B-cell interstitial infiltrates are associated with histological damage and outcome, but do not distinguish whether these infiltrates were the cause or the consequence of chronic tubulo-interstitial damage.

Alveolar epithelial cells undergo epithelial-mesenchymal transition in acute interstitial pneumonia: a case report

PubMed Central

2014-01-01

Background Acute interstitial pneumonia is a rare interstitial lung disease that rapidly progresses to respiratory failure or death. Several studies showed that myofibroblast plays an important role in the evolution of diffuse alveolar damage, which is the typical feature of acute interstitial pneumonia. However, no evidence exists whether alveolar epithelial cells are an additional source of myofibroblasts via epithelial-mesenchymal transition in acute interstitial pneumonia. Case presentation In this report, we present a case of acute interstitial pneumonia in a previously healthy 28-year-old non-smoking woman. Chest high-resolution computed tomography scan showed bilateral and diffusely ground-glass opacification. The biopsy was performed on the fifth day of her hospitalization, and results showed manifestation of acute exudative phase of diffuse alveolar damage characterized by hyaline membrane formation. On the basis of the preliminary diagnosis of acute interstitial pneumonia, high-dose glucocorticoid was used. However, this drug showed poor clinical response and could improve the patient’s symptoms only during the early phase. The patient eventually died of respiratory dysfunction. Histological findings in autopsy were consistent with the late form of acute interstitial pneumonia. Conclusions The results in this study revealed that alveolar epithelial cells underwent epithelial-mesenchymal transition and may be an important origin of myofibroblasts in the progression of acute interstitial pneumonia. Conducting research on the transformation of alveolar epithelial cells into myofibroblasts in the lung tissue of patients with acute interstitial pneumonia may be beneficial for the treatment of this disease. However, to our knowledge, no research has been conducted on this topic. PMID:24755111

Mechanics of Fluid-Filled Interstitial Gaps. II. Gap Characteristics in Xenopus Embryonic Ectoderm.

PubMed

Barua, Debanjan; Parent, Serge E; Winklbauer, Rudolf

2017-08-22

The ectoderm of the Xenopus embryo is permeated by a network of channels that appear in histological sections as interstitial gaps. We characterized this interstitial space by measuring gap sizes, angles formed between adjacent cells, and curvatures of cell surfaces at gaps. From these parameters, and from surface-tension values measured previously, we estimated the values of critical mechanical variables that determine gap sizes and shapes in the ectoderm, using a general model of interstitial gap mechanics. We concluded that gaps of 1-4 μm side length can be formed by the insertion of extracellular matrix fluid at three-cell junctions such that cell adhesion is locally disrupted and a tension difference between cell-cell contacts and the free cell surface at gaps of 0.003 mJ/m 2 is generated. Furthermore, a cell hydrostatic pressure of 16.8 ± 1.7 Pa and an interstitial pressure of 3.9 ± 3.6 Pa, relative to the central blastocoel cavity of the embryo, was found to be consistent with the observed gap size and shape distribution. Reduction of cell adhesion by the knockdown of C-cadherin increased gap volume while leaving intracellular and interstitial pressures essentially unchanged. In both normal and adhesion-reduced ectoderm, cortical tension of the free cell surfaces at gaps does not return to the high values characteristic of the free surface of the whole tissue. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Patterns of interstitial lung disease during everolimus treatment in patients with metastatic renal cell carcinoma.

PubMed

Mizuno, Ryuichi; Asano, Koichiro; Mikami, Shuji; Nagata, Hirohiko; Kaneko, Gou; Oya, Mototsugu

2012-05-01

To elucidate the patterns of interstitial lung disease during everolimus treatment in patients with metastatic renal cell carcinoma, we reviewed seven cases of everolimus-induced interstitial lung disease. Seven patients with metastatic renal cell carcinoma, which continued to progress despite treatment with sunitinib or sorafenib, developed interstitial lung disease after treatment with everolimus. Chest X-ray demonstrated diffuse infiltrates in lung fields, and chest computed tomography showed bilateral reticular and ground-glass opacities. Serum levels of lactate dehydrogenase (7/7), C-reactive protein (6/7), pulmonary surfactant associated protein D (1/7) and Krebs von den Lungen 6 (5/7) were elevated. The bronchoalveolar lavage fluid obtained from four patients with Grade 3 interstitial lung disease showed lymphocytosis. The transbronchial lung biopsy specimens showed interstitial lymphocytic infiltration and septal thickening of alveolar walls. In two cases with mild interstitial lung disease, the everolimus therapy was successfully continued. In four cases with Grade 3 interstitial lung disease, the drug was discontinued and steroid therapy was initiated. Pulmonary symptoms and radiological abnormalities resolved within 2 months. Serum Krebs von den Lungen 6 was elevated compared with baseline in all cases with interstitial lung disease. Some patients who developed mild interstitial lung disease during everolimus treatment could continue to receive the treatment. Even when severe interstitial lung disease developed, withdrawal of the drug and short-term use of high-dose steroids resulted in rapid recovery. Prompt recognition of interstitial lung disease exacerbation as well as exclusion of progressive disease or infection is of primary importance.

Role of the testis interstitial compartment in spermatogonial stem cell function

PubMed Central

Potter, Sarah J.; DeFalco, Tony

2017-01-01

Male fertility is maintained through intricate cellular and molecular interactions that ensure spermatogonial stem cells (SSCs) proceed in a step-wise differentiation process through spermatogenesis and spermiogenesis to produce sperm. SSCs lie within the seminiferous tubule compartment, which provides a nurturing environment for the development of sperm. Cells outside of the tubules, such as interstitial and peritubular cells, also help direct SSC activity. This review focuses on interstitial (interstitial macrophages, Leydig cells, and vasculature) and peritubular (peritubular macrophages, peritubular myoid cells) cells and their role in regulating SSC self-renewal and differentiation in mammals. Leydig cells, the major steroidogenic cells in the testis, influence SSCs through secreted factors, such as insulin growth factor 1 (IGF1) and colony stimulating factor 1 (CSF1). Macrophages interact with SSCs through various potential mechanisms, such as CSF1 and retinoic acid (RA), to induce proliferation or differentiation of SSCs, respectively. Vasculature influences SSC dynamics through CSF1, vascular endothelial growth factor (VEGF), and regulating oxygen levels. Lastly, peritubular myoid cells produce one of the most well-known factors that is required for SSC self-renewal, glial cell line derived neurotrophic factor (GDNF), as well as CSF1. Overall, SSC interactions with interstitial and peritubular cells are critical for SSC function and are an important underlying factor promoting male fertility. PMID:28115580

ATRX has a critical and conserved role in mammalian sexual differentiation

PubMed Central

2011-01-01

Background X-linked alpha thalassemia, mental retardation syndrome in humans is a rare recessive disorder caused by mutations in the ATRX gene. The disease is characterised by severe mental retardation, mild alpha-thalassemia, microcephaly, short stature, facial, skeletal, genital and gonadal abnormalities. Results We examined the expression of ATRX and ATRY during early development and gonadogenesis in two distantly related mammals: the tammar wallaby (a marsupial) and the mouse (a eutherian). This is the first examination of ATRX and ATRY in the developing mammalian gonad and fetus. ATRX and ATRY were strongly expressed in the developing male and female gonad respectively, of both species. In testes, ATRY expression was detected in the Sertoli cells, germ cells and some interstitial cells. In the developing ovaries, ATRX was initially restricted to the germ cells, but was present in the granulosa cells of mature ovaries from the primary follicle stage onwards and in the corpus luteum. ATRX mRNA expression was also examined outside the gonad in both mouse and tammar wallaby whole embryos. ATRX was detected in the developing limbs, craniofacial elements, neural tissues, tail and phallus. These sites correspond with developmental deficiencies displayed by ATR-X patients. Conclusions There is a complex expression pattern throughout development in both mammals, consistent with many of the observed ATR-X syndrome phenotypes in humans. The distribution of ATRX mRNA and protein in the gonads was highly conserved between the tammar and the mouse. The expression profile within the germ cells and somatic cells strikingly overlaps with that of DMRT1, suggesting a possible link between these two genes in gonadal development. Taken together, these data suggest that ATRX has a critical and conserved role in normal development of the testis and ovary in both the somatic and germ cells, and that its broad roles in early mammalian development and gonadal function have remained unchanged for over 148 million years of mammalian evolution. PMID:21672208

ATRX has a critical and conserved role in mammalian sexual differentiation.

PubMed

Huyhn, Kim; Renfree, Marilyn B; Graves, Jennifer A; Pask, Andrew J

2011-06-14

X-linked alpha thalassemia, mental retardation syndrome in humans is a rare recessive disorder caused by mutations in the ATRX gene. The disease is characterised by severe mental retardation, mild alpha-thalassemia, microcephaly, short stature, facial, skeletal, genital and gonadal abnormalities. We examined the expression of ATRX and ATRY during early development and gonadogenesis in two distantly related mammals: the tammar wallaby (a marsupial) and the mouse (a eutherian). This is the first examination of ATRX and ATRY in the developing mammalian gonad and fetus. ATRX and ATRY were strongly expressed in the developing male and female gonad respectively, of both species. In testes, ATRY expression was detected in the Sertoli cells, germ cells and some interstitial cells. In the developing ovaries, ATRX was initially restricted to the germ cells, but was present in the granulosa cells of mature ovaries from the primary follicle stage onwards and in the corpus luteum. ATRX mRNA expression was also examined outside the gonad in both mouse and tammar wallaby whole embryos. ATRX was detected in the developing limbs, craniofacial elements, neural tissues, tail and phallus. These sites correspond with developmental deficiencies displayed by ATR-X patients. There is a complex expression pattern throughout development in both mammals, consistent with many of the observed ATR-X syndrome phenotypes in humans. The distribution of ATRX mRNA and protein in the gonads was highly conserved between the tammar and the mouse. The expression profile within the germ cells and somatic cells strikingly overlaps with that of DMRT1, suggesting a possible link between these two genes in gonadal development. Taken together, these data suggest that ATRX has a critical and conserved role in normal development of the testis and ovary in both the somatic and germ cells, and that its broad roles in early mammalian development and gonadal function have remained unchanged for over 148 million years of mammalian evolution.

Telocytes and stem cells in limbus and uvea of mouse eye.

PubMed

Luesma, María José; Gherghiceanu, Mihaela; Popescu, Laurenţiu M

2013-08-01

The potential of stem cell (SC) therapies for eye diseases is well-recognized. However, the results remain only encouraging as little is known about the mechanisms responsible for eye renewal, regeneration and/or repair. Therefore, it is critical to gain knowledge about the specific tissue environment (niches) where the stem/progenitor cells reside in eye. A new type of interstitial cell-telocyte (TC) (www.telocytes.com) was recently identified by electron microscopy (EM). TCs have very long (tens of micrometres) and thin (below 200 nm) prolongations named telopodes (Tp) that form heterocellular networks in which SCs are embedded. We found TCs by EM and electron tomography in sclera, limbus and uvea of the mouse eye. Furthermore, EM showed that SCs were present in the anterior layer of the iris and limbus. Adhaerens and gap junctions were found to connect TCs within a network in uvea and sclera. Nanocontacts (electron-dense structures) were observed between TCs and other cells: SCs, melanocytes, nerve endings and macrophages. These intercellular 'feet' bridged the intercellular clefts (about 10 nm wide). Moreover, exosomes (extracellular vesicles with a diameter up to 100 nm) were delivered by TCs to other cells of the iris stroma. The ultrastructural nanocontacts of TCs with SCs and the TCs paracrine influence via exosomes in the epithelial and stromal SC niches suggest an important participation of TCs in eye regeneration. © 2013 The Authors. Journal of Cellular and Molecular Medicine Published by Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.

Mechanics of Fluid-Filled Interstitial Gaps. I. Modeling Gaps in a Compact Tissue.

PubMed

Parent, Serge E; Barua, Debanjan; Winklbauer, Rudolf

2017-08-22

Fluid-filled interstitial gaps are a common feature of compact tissues held together by cell-cell adhesion. Although such gaps can in principle be the result of weak, incomplete cell attachment, adhesion is usually too strong for this to occur. Using a mechanical model of tissue cohesion, we show that, instead, a combination of local prevention of cell adhesion at three-cell junctions by fluidlike extracellular material and a reduction of cortical tension at the gap surface are sufficient to generate stable gaps. The size and shape of these interstitial gaps depends on the mechanical tensions between cells and at gap surfaces, and on the difference between intracellular and interstitial pressures that is related to the volume of the interstitial fluid. As a consequence of the dependence on tension/tension ratios, the presence of gaps does not depend on the absolute strength of cell adhesion, and similar gaps are predicted to occur in tissues of widely differing cohesion. Tissue mechanical parameters can also vary within and between cells of a given tissue, generating asymmetrical gaps. Within limits, these can be approximated by symmetrical gaps. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Imaging CD4 T Cell Interstitial Migration in the Inflamed Dermis

PubMed Central

Gaylo, Alison; Overstreet, Michael G.; Fowell, Deborah J.

2016-01-01

The ability of CD4 T cells to carry out effector functions is dependent upon the rapid and efficient migration of these cells in inflamed peripheral tissues through an as-yet undefined mechanism. The application of multiphoton microscopy to the study of the immune system provides a tool to measure the dynamics of immune responses within intact tissues. Here we present a protocol for non-invasive intravital multiphoton imaging of CD4 T cells in the inflamed mouse ear dermis. Use of a custom imaging platform and a venous catheter allows for the visualization of CD4 T cell dynamics in the dermal interstitium, with the ability to interrogate these cells in real-time via the addition of blocking antibodies to key molecular components involved in motility. This system provides advantages over both in vitro models and surgically invasive imaging procedures. Understanding the pathways used by CD4 T cells for motility may ultimately provide insight into the basic function of CD4 T cells as well as the pathogenesis of both autoimmune diseases and pathology from chronic infections. PMID:27078264

Therapeutic Effects of Human Amniotic Fluid-Derived Stem Cells on Renal Interstitial Fibrosis in a Murine Model of Unilateral Ureteral Obstruction

PubMed Central

Yin, Zhongcheng; Zhou, Xudong; Li, Xiaoju; Xiao, Aiguo

2013-01-01

Interstitial fibrosis is regarded as the main pathway for the progression of chronic kidney disease (CKD) and is often associated with severe renal dysfunction. Stem cell-based therapies may provide alternative approaches for the treatment of CKD. Human amniotic fluid-derived stem cells (hAFSCs) are a novel stem cell population, which exhibit both embryonic and mesenchymal stem cell characteristics. Herein, the present study investigated whether the transplantation of hAFSCs into renal tissues could improve renal interstitial fibrosis in a murine model of unilateral ureteral obstruction (UUO). We showed that hAFSCs provided a protective effect and alleviated interstitial fibrosis as reflected by an increase in microvascular density; additionally, hAFSCs treatment beneficially modulated protein levels of vascular endothelial growth factor (VEGF), hypoxia inducible factor-1α (HIF-1α) and transforming growth factor-β1 (TGF-β1). Therefore, we hypothesize that hAFSCs could represent an alternative, readily available source of stem cells that can be applied for the treatment of renal interstitial fibrosis. PMID:23724119

The dynamic of FUS-induced BBB Opening in Mouse Brain assessed by contrast enhanced MRI

NASA Astrophysics Data System (ADS)

Jenne, Jürgen W.; Krafft, Axel J.; Maier, Florian; Krause, Marie N.; Kleber, Susanne; Huber, Peter E.; Martin-Villalba, Ana; Bock, Michael

2010-03-01

Focused ultrasound (FUS) in combination with the administration of gas-filled microbubbles, can induce a localized and reversible opening of the blood brain barrier (BBB). Contrast enhanced magnetic resonance imaging (MRI) has been demonstrated as a precise tool to monitor such a local BBB disruption. However, the opening/closing mechanisms of the BBB with FUS are still largely unknown. In this ongoing project, we study the BBB opening dynamics in mouse brain comparing an interstitial and an intravascular MR contrast agent (CA). FUS in mouse brain was performed with an MRI compatible treatment setup (1.7 MHz fix-focus US transducer, f' = 68 mm, NA = 0.44; focus: 8.1 mm length; O/ = 1.1 mm) in a 1.5 T whole body MRI system. For BBB opening, forty 10 ms-long FUS-pulses were applied at a repetition rate of 1 Hz at 1 MPa. The i.v. administration of the micro bubbles (50 μl SonoVue®) was started simultaneously with FUS exposure. To analyze the BBB opening process, short-term and long-term MRI signal dynamics of the interstitial MR contrast agent Magnevist® and the intravascular CA Vasovist® (Bayer-Schering) were studied. To assess short-term signal dynamics, T1-weighted inversion recovery turbo FLASH images (1s) were repeatedly acquired. Repeated 3D FLASH acquisitions (90 s) were used to assess long-term MRI signal dynamics. The short-term MRI signal enhancements showed comparable time constants for both types of MR contrast agents: 1.1 s (interstitial) vs. 0.8 s (intravascular). This time constant may serve as a time constant of the BBB opening process with the given FUS exposure parameters. For the long-term signal dynamics the intravascular CA (62±10 min) showed a fife times greater time constant as the interstitial contrast agent (12±10 min). This might be explained by the high molecular weight (˜60 kDa) of the intravascular Vasovist due to its reversible binding to blood serum albumin resulting in a prolonged half-life in the blood stream compared to the interstitial CA. As the intravascular CA offers a much longer time window for therapy assessment, FUS-BBB therapy control with an intravascular CA might be favorable.

Development of a Model of Chronic Kidney Disease in the C57BL/6 Mouse with Properties of Progressive Human CKD

PubMed Central

Cruz, Gaile L.; Lu, Chao; Carlisle, Rachel E.; Werner, Kaitlyn E.; Ask, Kjetil; Dickhout, Jeffrey G.

2015-01-01

Chronic kidney disease (CKD) is a major healthcare problem with increasing prevalence in the population. CKD leads to end stage renal disease and increases the risk of cardiovascular disease. As such, it is important to study the mechanisms underlying CKD progression. To this end, an animal model was developed to allow the testing of new treatment strategies or molecular targets for CKD prevention. Many underlying risk factors result in CKD but the disease itself has common features, including renal interstitial fibrosis, tubular epithelial cell loss through apoptosis, glomerular damage, and renal inflammation. Further, CKD shows differences in prevalence between the genders with premenopausal women being relatively resistant to CKD. We sought to develop and characterize an animal model with these common features of human CKD in the C57BL/6 mouse. Mice of this genetic background have been used to produce transgenic strains that are commercially available. Thus, a CKD model in this strain would allow the testing of the effects of numerous genes on the severity or progression of CKD with minimal cost. This paper describes such a mouse model of CKD utilizing angiotensin II and deoxycorticosterone acetate as inducers. PMID:26064882

Robust G2 pausing of adult stem cells in Hydra.

PubMed

Buzgariu, Wanda; Crescenzi, Marco; Galliot, Brigitte

2014-01-01

Hydra is a freshwater hydrozoan polyp that constantly renews its two tissue layers thanks to three distinct stem cell populations that cannot replace each other, epithelial ectodermal, epithelial endodermal, and multipotent interstitial. These adult stem cells, located in the central body column, exhibit different cycling paces, slow for the epithelial, fast for the interstitial. To monitor the changes in cell cycling in Hydra, we established a fast and efficient flow cytometry procedure, which we validated by confirming previous findings, as the Nocodazole-induced reversible arrest of cell cycling in G2/M, and the mitogenic signal provided by feeding. Then to dissect the cycling and differentiation behaviors of the interstitial stem cells, we used the AEP_cnnos1 and AEP_Icy1 transgenic lines that constitutively express GFP in this lineage. For the epithelial lineages we used the sf-1 strain that rapidly eliminates the fast cycling cells upon heat-shock and progressively becomes epithelial. This study evidences similar cycling patterns for the interstitial and epithelial stem cells, which all alternate between the G2 and S-phases traversing a minimal G1-phase. We also found interstitial progenitors with a shorter G2 that pause in G1/G0. At the animal extremities, most cells no longer cycle, the epithelial cells terminally differentiate in G2 and the interstitial progenitors in G1/G0. At the apical pole ~80% cells are post-mitotic differentiated cells, reflecting the higher density of neurons and nematocytes in this region. We discuss how the robust G2 pausing of stem cells, maintained over weeks of starvation, may contribute to regeneration. Copyright © 2014 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

Modulation of Pacemaker Potentials in Murine Small Intestinal Interstitial Cells of Cajal by Gamisoyo-San, a Traditional Chinese Herbal Medicine.

PubMed

Kim, Doeun; Kim, Jung Nam; Nam, Joo Hyun; Lee, Jong Rok; Kim, Sang Chan; Kim, Byung Joo

2018-04-19

The Gamisoyo-san (GSS) has been used for -improving the gastrointestinal (GI) symptoms. The purpose of this study was to investigate the effects of GSS, a traditional Chinese herbal medicine, on the pacemaker potentials of mouse small intestinal interstitial cells of Cajal (ICCs). ICCs from the small intestines were dissociated and cultured. Whole-cell patch-clamp configuration was used to record pacemaker potentials and membrane currents. GSS depolarized ICC pacemaker potentials in a dose-dependent manner. Pretreatment with 4-diphenylacetoxypiperidinium iodide completely inhibited GSS-induced pacemaker potential depolarizations. Intracellular GDP-β-S inhibited GSS-induced effects, and in the presence of U-73122, GSS-induced effects were inhibited. Also, GSS in the presence of a Ca2+-free solution or thapsigargin did not depolarize pacemaker potentials. However, in the presence of calphostin C, GSS slightly depolarized pacemaker potentials. Furthermore, GSS inhibited both transient receptor potential melastatin7 and Ca2+-activated Cl- channel (anoctamin1) currents. GSS depolarized pacemaker potentials of ICCs via G protein and muscarinic M3 receptor signaling pathways and through internal or external Ca2+-, phospholipase C-, and protein kinase C-dependent and transient receptor potential melastatin 7-, and anoctamin 1-independent pathways. The study shows that GSS may regulate GI tract motility, suggesting that GSS could be a basis for developing novel prokinetic agents for treating GI motility dysfunctions. © 2018 S. Karger AG, Basel.

Mixed germ cell-sex cord-stromal tumor with a concurrent interstitial cell tumor in a ferret

PubMed Central

INOUE, Saki; YONEMARU, Kayoko; YANAI, Tokuma; SAKAI, Hiroki

2014-01-01

A 5-year-old male ferret presented with an enlarged canalicular testis in the left inguinal region. Microscopically, the enlarged testis consisted of a diffuse intimately admixed proliferation of c-kit-positive germ cell-like and Wilms tumor-1 protein-positive Sertoli cell-like components, but no Call-Exner body was detected. In addition, the compact proliferation of steroidogenic acute regulatory protein-intense positive interstitial cells was identified in a separate peripheral area of the mass. Based on histopathological and immunohistochemical findings, the tumor was diagnosed as a mixed germ cell-sex cord-stromal tumor with a concurrent interstitial cell tumor. PMID:25311985

Effects of lubiprostone on pacemaker activity of interstitial cells of cajal from the mouse colon.

PubMed

Jiao, Han-Yi; Kim, Dong Hyun; Ki, Jung Suk; Ryu, Kwon Ho; Choi, Seok; Jun, Jae Yeoul

2014-08-01

Lubiprostone is a chloride (Cl(-)) channel activator derived from prostaglandin E1 and used for managing constipation. In addition, lubiprostone affects the activity of gastrointestinal smooth muscles. Interstitial cells of Cajal (ICCs) are pacemaker cells that generate slow-wave activity in smooth muscles. We studied the effects of lubiprostone on the pacemaker potentials of colonic ICCs. We used the whole-cell patch-clamp technique to determine the pacemaker activity in cultured colonic ICCs obtained from mice. Lubiprostone hyperpolarized the membrane and inhibited the generation of pacemaker potentials. Prostanoid EP1, EP2, EP3, and EP4 antagonists (SC-19220, PF-04418948, 6-methoxypyridine-2-boronc acid N-phenyldiethanolamine ester, and GW627368, respectively) did not block the response to lubiprostone. L-NG-nitroarginine methyl ester (L-NAME, an inhibitor of nitric oxide synthase) and 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ, an inhibitor of guanylate cyclase) did not block the response to lubiprostone. In addition, tetraethylammonium (TEA, a voltage-dependent potassium [K(+)] channel blocker) and apamin (a calcium [Ca(2+)]-dependent K(+) channel blocker) did not block the response to lubiprostone. However, glibenclamide (an ATP-sensitive K(+) channel blocker) blocked the response to lubiprostone. Similar to lubiprostone, pinacidil (an opener of ATP-sensitive K(+) channel) hyperpolarized the membrane and inhibited the generation of pacemaker potentials, and these effects were inhibited by glibenclamide. These results suggest that lubiprostone can modulate the pacemaker potentials of colonic ICCs via activation of ATP-sensitive K(+) channel through a prostanoid EP receptor-independent mechanism.

Effects of Lubiprostone on Pacemaker Activity of Interstitial Cells of Cajal from the Mouse Colon

PubMed Central

Jiao, Han-Yi; Kim, Dong Hyun; Ki, Jung Suk; Ryu, Kwon Ho; Choi, Seok

2014-01-01

Lubiprostone is a chloride (Cl-) channel activator derived from prostaglandin E1 and used for managing constipation. In addition, lubiprostone affects the activity of gastrointestinal smooth muscles. Interstitial cells of Cajal (ICCs) are pacemaker cells that generate slow-wave activity in smooth muscles. We studied the effects of lubiprostone on the pacemaker potentials of colonic ICCs. We used the whole-cell patch-clamp technique to determine the pacemaker activity in cultured colonic ICCs obtained from mice. Lubiprostone hyperpolarized the membrane and inhibited the generation of pacemaker potentials. Prostanoid EP1, EP2, EP3, and EP4 antagonists (SC-19220, PF-04418948, 6-methoxypyridine-2-boronc acid N-phenyldiethanolamine ester, and GW627368, respectively) did not block the response to lubiprostone. L-NG-nitroarginine methyl ester (L-NAME, an inhibitor of nitric oxide synthase) and 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ, an inhibitor of guanylate cyclase) did not block the response to lubiprostone. In addition, tetraethylammonium (TEA, a voltage-dependent potassium [K+] channel blocker) and apamin (a calcium [Ca2+]-dependent K+ channel blocker) did not block the response to lubiprostone. However, glibenclamide (an ATP-sensitive K+ channel blocker) blocked the response to lubiprostone. Similar to lubiprostone, pinacidil (an opener of ATP-sensitive K+ channel) hyperpolarized the membrane and inhibited the generation of pacemaker potentials, and these effects were inhibited by glibenclamide. These results suggest that lubiprostone can modulate the pacemaker potentials of colonic ICCs via activation of ATP-sensitive K+ channel through a prostanoid EP receptor-independent mechanism. PMID:25177167

Interstitial flows promote an amoeboid cell phenotype and motility of breast cancer cells

NASA Astrophysics Data System (ADS)

Tung, Chih-Kuan; Huang, Yu Ling; Zheng, Angela; Wu, Mingming

2015-03-01

Lymph nodes, the drainage systems for interstitial flows, are clinically known to be the first metastatic sites of many cancer types including breast and prostate cancers. Here, we demonstrate that breast cancer cell morphology and motility is modulated by interstitial flows in a cell-ECM adhesion dependent manner. The average aspect ratios of the cells are significantly lower (or are more amoeboid like) in the presence of the flow in comparison to the case when the flow is absent. The addition of exogenous adhesion molecules within the extracellular matrix (type I collagen) enhances the overall aspect ratio (or are more mesenchymal like) of the cell population. Using measured cell trajectories, we find that the persistence of the amoeboid cells (aspect ratio less than 2.0) is shorter than that of mesenchymal cells. However, the maximum speed of the amoeboid cells is larger than that of mesenchymal cells. Together these findings provide the novel insight that interstitial flows promote amoeboid cell morphology and motility and highlight the plasticity of tumor cell motility in response to its biophysical environment. Supported by NIH Grant R21CA138366.

Abcg2 expression marks tissue-specific stem cells in multiple organs in a mouse progeny tracking model.

PubMed

Fatima, Soghra; Zhou, Sheng; Sorrentino, Brian P

2012-02-01

The side population phenotype is associated with the Hoechst dye efflux activity of the Abcg2 transporter and identifies hematopoietic stem cells (HSCs) in the bone marrow. This association suggests the direct use of Abcg2 expression to identify adult stem cells in various other organs. We have generated a lineage tracing mouse model based on an allele that coexpresses both Abcg2 and a CreERT2 expression cassette. By crossing these mice with lox-STOP-lox reporter lines (LacZ or YFP), cells that express Abcg2 and their progeny were identified following treatment with tamoxifen (Tam). In the liver and kidney, in which mature cells express Abcg2, reporter gene expression verified the expected physiologic expression pattern of the recombinant allele. Long-term marking of HSCs was seen in multiple peripheral blood lineages from adult mice, demonstrating that Abcg2(+) bone marrow HSCs contribute to steady-state hematopoiesis. Stem cell tracing patterns were seen in the small intestine and in seminiferous tubules in the testis 20 months after Tam treatment, proving that stem cells from these organs express Abcg2. Interstitial cells from skeletal and cardiac muscle were labeled, and some cells were costained with endothelial markers, raising the possibility that these cells may function in the repair response to muscle injury. Altogether, these studies prove that Abcg2 is a stem cell marker for blood, small intestine, testicular germ cells, and possibly for injured skeletal and/or cardiac muscle and provide a new model for studying stem cell activity that does not require transplant-based assays. Copyright © 2011 AlphaMed Press.

The safety and efficacy of carboplatin plus nanoparticle albumin-bound paclitaxel in the treatment of non-small cell lung cancer patients with interstitial lung disease.

PubMed

Yasuda, Yuichiro; Hattori, Yoshihiro; Tohnai, Rie; Ito, Shoichi; Kawa, Yoshitaka; Kono, Yuko; Urata, Yoshiko; Nogami, Munenobu; Takenaka, Daisuke; Negoro, Shunichi; Satouchi, Miyako

2018-01-01

The optimal chemotherapy regimen for non-small cell lung cancer patients with interstitial lung disease is unclear. We therefore investigated the safety and efficacy of carboplatin plus nab-paclitaxel as a first-line regimen for non-small cell lung cancer in patients with interstitial lung disease. We retrospectively reviewed advanced non-small cell lung cancer patients with interstitial lung disease who received carboplatin plus nab-paclitaxel as a first-line chemotherapy regimen at Hyogo Cancer Center between February 2013 and August 2016. interstitial lung disease was diagnosed according to the findings of pretreatment chest high-resolution computed tomography. Twelve patients were included (male, n = 11; female, n = 1). The overall response rate was 67% and the disease control rate was 100%. The median progression free survival was 5.1 months (95% CI: 2.9-8.3 months) and the median overall survival was 14.9 months (95% CI: 4.8-not reached). A chemotherapy-related acute exacerbation of interstitial lung disease was observed in one patient; the extent of this event was Grade 2. There were no treatment-related deaths. Carboplatin plus nab-paclitaxel, as a first-line chemotherapy regimen for non-small cell lung cancer, showed favorable efficacy and safety in patients with preexisting interstitial lung disease. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com

Nitrergic signalling via interstitial cells of Cajal regulates motor activity in murine colon.

PubMed

Lies, Barbara; Beck, Katharina; Keppler, Jonas; Saur, Dieter; Groneberg, Dieter; Friebe, Andreas

2015-10-15

In the enteric nervous systems, NO is released from nitrergic neurons as a major inhibitory neurotransmitter. NO acts via NO-sensitive guanylyl cyclase (NO-GC), which is found in different gastrointestinal (GI) cell types including smooth muscle cells (SMCs) and interstitial cells of Cajal (ICC). The precise mechanism of nitrergic signalling through these two cell types to regulate colonic spontaneous contractions is not fully understood yet. In the present study we investigated the impact of endogenous and exogenous NO on colonic contractile motor activity using mice lacking nitric oxide-sensitive guanylyl cyclase (NO-GC) globally and specifically in SMCs and ICC. Longitudinal smooth muscle of proximal colon from wild-type (WT) and knockout (KO) mouse strains exhibited spontaneous contractile activity ex vivo. WT and smooth muscle-specific guanylyl cyclase knockout (SMC-GCKO) colon showed an arrhythmic contractile activity with varying amplitudes and frequencies. In contrast, colon from global and ICC-specific guanylyl cyclase knockout (ICC-GCKO) animals showed a regular contractile rhythm with constant duration and amplitude of the rhythmic contractions. Nerve blockade (tetrodotoxin) or specific blockade of NO signalling (L-NAME, ODQ) did not significantly affect contractions of GCKO and ICC-GCKO colon whereas the arrhythmic contractile patterns of WT and SMC-GCKO colon were transformed into uniform motor patterns. In contrast, the response to electric field-stimulated neuronal NO release was similar in SMC-GCKO and global GCKO. In conclusion, our results indicate that basal enteric NO release acts via myenteric ICC to influence the generation of spontaneous contractions whereas the effects of elevated endogenous NO are mediated by SMCs in the murine proximal colon. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

Modulation of invasive phenotype by interstitial pressure-driven convection in aggregates of human breast cancer cells.

PubMed

Tien, Joe; Truslow, James G; Nelson, Celeste M

2012-01-01

This paper reports the effect of elevated pressure on the invasive phenotype of patterned three-dimensional (3D) aggregates of MDA-MB-231 human breast cancer cells. We found that the directionality of the interstitial pressure profile altered the frequency of invasion by cells located at the surface of an aggregate. In particular, application of pressure at one end of an aggregate suppressed invasion at the opposite end. Experimental alteration of the configuration of cell aggregates and computational modeling of the resulting flow and solute concentration profiles revealed that elevated pressure inhibited invasion by altering the chemical composition of the interstitial fluid near the surface of the aggregate. Our data reveal a link between hydrostatic pressure, interstitial convection, and invasion.

Anti-tau antibody administration increases plasma tau in transgenic mice and patients with tauopathy

PubMed Central

Yanamandra, Kiran; Patel, Tirth K.; Jiang, Hong; Schindler, Suzanne; Ulrich, Jason D.; Boxer, Adam L.; Miller, Bruce L.; Kerwin, Diana R.; Gallardo, Gilbert; Stewart, Floy; Finn, Mary Beth; Cairns, Nigel J.; Verghese, Philip B.; Fogelman, Ilana; West, Tim; Braunstein, Joel; Robinson, Grace; Keyser, Jennifer; Roh, Joseph; Knapik, Stephanie S.; Hu, Yan; Holtzman, David M.

2017-01-01

Tauopathies are a group of disorders in which the cytosolic protein tau aggregates and accumulates in cells within the brain, resulting in neurodegeneration. A promising treatment being explored for tauopathies is passive immunization with anti-tau antibodies. We previously found that administration of an anti-tau antibody to human tau transgenic mice increased the concentration of plasma tau. We further explored the effects of administering an anti-tau antibody on plasma tau. After peripheral administration of an anti-tau antibody to human patients with tauopathy and to mice expressing human tau in the central nervous system, there was a dose-dependent increase in plasma tau. In mouse plasma, we found that tau had a short half-life of 8 min that increased to more than 3 hours after administration of anti-tau antibody. As tau transgenic mice accumulated insoluble tau in the brain, brain soluble and interstitial fluid tau decreased. Administration of anti-tau antibody to tau transgenic mice that had decreased brain soluble tau and interstitial fluid tau resulted in an increase in plasma tau, but this increase was less than that observed in tau transgenic mice without these brain changes. Tau transgenic mice subjected to acute neuronal injury using 3-nitropropionic acid showed increased interstitial fluid tau and plasma tau. These data suggest that peripheral administration of an anti-tau antibody results in increased plasma tau, which correlates with the concentration of extracellular and soluble tau in the brain. PMID:28424326

Alpha-particle radiotherapy: For large solid tumors diffusion trumps targeting.

PubMed

Zhu, Charles; Sempkowski, Michelle; Holleran, Timothy; Linz, Thomas; Bertalan, Thomas; Josefsson, Anders; Bruchertseifer, Frank; Morgenstern, Alfred; Sofou, Stavroula

2017-06-01

Diffusion limitations on the penetration of nanocarriers in solid tumors hamper their therapeutic use when labeled with α-particle emitters. This is mostly due to the α-particles' relatively short range (≤100 μm) resulting in partial tumor irradiation and limited killing. To utilize the high therapeutic potential of α-particles against solid tumors, we designed non-targeted, non-internalizing nanometer-sized tunable carriers (pH-tunable liposomes) that are triggered to release, within the slightly acidic tumor interstitium, highly-diffusive forms of the encapsulated α-particle generator Actinium-225 ( 225 Ac) resulting in more homogeneous distributions of the α-particle emitters, improving uniformity in tumor irradiation and increasing killing efficacies. On large multicellular spheroids (400 μm-in-diameter), used as surrogates of the avascular areas of solid tumors, interstitially-releasing liposomes resulted in best growth control independent of HER2 expression followed in performance by (a) the HER2-targeting radiolabeled antibody or (b) the non-responsive liposomes. In an orthotopic human HER2-negative mouse model, interstitially-releasing 225 Ac-loaded liposomes resulted in the longest overall and median survival. This study demonstrates the therapeutic potential of a general strategy to bypass the diffusion-limited transport of radionuclide carriers in solid tumors enabling interstitial release from non-internalizing nanocarriers of highly-diffusing and deeper tumor-penetrating molecular forms of α-particle emitters, independent of cell-targeting. Copyright © 2017 Elsevier Ltd. All rights reserved.

Autosomal mutations in mouse kidney epithelial cells exposed to high-energy protons in vivo or in culture.

PubMed

Turker, Mitchell S; Grygoryev, Dmytro; Dan, Cristian; Eckelmann, Bradley; Lasarev, Michael; Gauny, Stacey; Kwoh, Ely; Kronenberg, Amy

2013-05-01

Proton exposure induces mutations and cancer, which are presumably linked. Because protons are abundant in the space environment and significant uncertainties exist for the effects of space travel on human health, the purpose of this study was to identify the types of mutations induced by exposure of mammalian cells to 4-5 Gy of 1 GeV protons. We used an assay that selects for mutations affecting the chromosome 8-encoded Aprt locus in mouse kidney cells and selected mutants after proton exposure both in vivo and in cell culture. A loss of heterozygosity (LOH) assay for DNA preparations from the in vivo-derived kidney mutants revealed that protons readily induced large mutational events. Fluorescent in situ hybridization painting for chromosome 8 showed that >70% of proton-induced LOH patterns resembling mitotic recombination were in fact the result of nonreciprocal chromosome translocations, thereby demonstrating an important role for DNA double-strand breaks in proton mutagenesis. Large interstitial deletions, which also require the formation and resolution of double-strand breaks, were significantly induced in the cell culture environment (14% of all mutants), but to a lesser extend in vivo (2% of all mutants) suggesting that the resolution of proton-induced double-strand breaks can differ between the intact tissue and cell culture microenvironments. In total, the results demonstrate that double-strand break formation is a primary determinant for proton mutagenesis in epithelial cell types and suggest that resultant LOH for significant genomic regions play a critical role in proton-induced cancers.

Interstitial Fluid Flow Increases Hepatocellular Carcinoma Cell Invasion through CXCR4/CXCL12 and MEK/ERK Signaling

PubMed Central

2015-01-01

Hepatocellular carcinoma (HCC) is the most common form of liver cancer (~80%), and it is one of the few cancer types with rising incidence in the United States. This highly invasive cancer is very difficult to detect until its later stages, resulting in limited treatment options and low survival rates. There is a dearth of knowledge regarding the mechanisms associated with the effects of biomechanical forces such as interstitial fluid flow (IFF) on hepatocellular carcinoma invasion. We hypothesized that interstitial fluid flow enhanced hepatocellular carcinoma cell invasion through chemokine-mediated autologous chemotaxis. Utilizing a 3D in vitro invasion assay, we demonstrated that interstitial fluid flow promoted invasion of hepatocellular carcinoma derived cell lines. Furthermore, we showed that autologous chemotaxis influences this interstitial fluid flow-induced invasion of hepatocellular carcinoma derived cell lines via the C-X-C chemokine receptor type 4 (CXCR4)/C-X-C motif chemokine 12 (CXCL12) signaling axis. We also demonstrated that mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK) signaling affects interstitial fluid flow-induced invasion; however, this pathway was separate from CXCR4/CXCL12 signaling. This study demonstrates, for the first time, the potential role of interstitial fluid flow in hepatocellular carcinoma invasion. Uncovering the mechanisms that control hepatocellular carcinoma invasion will aid in enhancing current liver cancer therapies and provide better treatment options for patients. PMID:26560447

PURIFICATION OF RAT LEYDIG CELLS: INCREASED YIELDS AFTER UNIT-GRAVITY SEDIMENTATION OF COLLAGENASE-DISPERSED INTERSTITIAL CELLS

EPA Science Inventory

Abstract

Procedures for purification of Leydig cells have facilitated studies of their regulatory biology. A multistep procedure, that includes a filtration with nylon mesh (100 micron pore size) to separate interstitial cells from the seminiferous tubules, combining centr...

Chromosomal rearrangements involving telomeric DNA sequences in Balb/3T3 cells transfected with the Ha-ras oncogene.

PubMed

Peitl, Paulo; Mello, Stephano S; Camparoto, Marjori L; Passos, Geraldo A S; Hande, Manoor P; Cardoso, Renato S; Sakamoto-Hojo, Elza T

2002-01-01

Chromosomal instability involving telomeric DNA sequences was studied in mouse Balb/3T3 fibroblasts transfected with a mutated human c-Ha-ras-1 gene (B61 cells) and spontaneously immortalized normal parental cells (A31 cells), using fluorescence in situ hybridization (FISH). FISH analysis with a telomeric probe revealed high frequencies of chromosome alterations involving telomeric regions, mainly stable and unstable Robertsonian fusion-like configurations (RLC) (0.25 and 1.95/cell in A31 and B61 cells, respectively) and chromosome ends lacking telomeric signals in one (LTS') or both chromatids (LTS") (5.9 and 17.5/cell for A31 and B61 cells, respectively). Interstitial telomeric sequences (ITS) were also detected at both non-telomeric sites and in the centromeres of RLC. The frequencies of RLCs with ITS located in the centromeres were 3-fold higher in B61 compared with A31 cells. We demonstrated a high level of chromosome instability involving telomeric DNA sequences in ras-transfected cells overexpressing ras mRNA, which could be a consequence of rapid cell cycle progression associated with a deficient telomere capping mechanism.

An essential role of intestinal cell kinase in lung development is linked to the perinatal lethality of human ECO syndrome

PubMed Central

Tong, Yixin; Park, So Hyun; Wu, Di; Xu, Wenhao; Guillot, Stacey J.; Jin, Li; Li, Xudong; Wang, Yalin; Lin, Chyuan-Sheng; Fu, Zheng

2017-01-01

Human endocrine-cerebro-osteodysplasia (ECO) syndrome, caused by the loss-of-function mutation R272Q in the ICK (intestinal cell kinase) gene, is a neonatal-lethal developmental disorder. To elucidate the molecular basis of ECO syndrome, we constructed an Ick R272Q knock-in mouse model that recapitulates ECO pathological phenotypes. Newborns bearing Ick R272Q homozygous mutations die at birth due to respiratory distress. Ick mutant lungs exhibit not only impaired branching morphogenesis associated with reduced mesenchymal proliferation, but also significant airspace deficiency in primitive alveoli concomitant with abnormal interstitial mesenchymal differentiation. ICK dysfunction induces elongated primary cilia and perturbs ciliary Hedgehog signaling and autophagy during lung sacculation. Our study identifies an essential role for ICK in lung development and advances the mechanistic understanding of ECO syndrome. PMID:28380258

An essential role of intestinal cell kinase in lung development is linked to the perinatal lethality of human ECO syndrome.

PubMed

Tong, Yixin; Park, So Hyun; Wu, Di; Xu, Wenhao; Guillot, Stacey J; Jin, Li; Li, Xudong; Wang, Yalin; Lin, Chyuan-Sheng; Fu, Zheng

2017-05-01

Human endocrine-cerebro-osteodysplasia (ECO) syndrome, caused by the loss-of-function mutation R272Q in the intestinal cell kinase (ICK) gene, is a neonatal-lethal developmental disorder. To elucidate the molecular basis of ECO syndrome, we constructed an Ick R272Q knock-in mouse model that recapitulates ECO pathological phenotypes. Newborns bearing Ick R272Q homozygous mutations die at birth due to respiratory distress. Ick mutant lungs exhibit not only impaired branching morphogenesis associated with reduced mesenchymal proliferation but also significant airspace deficiency in primitive alveoli concomitant with abnormal interstitial mesenchymal differentiation. ICK dysfunction induces elongated primary cilia and perturbs ciliary Hedgehog signaling and autophagy during lung sacculation. Our study identifies an essential role for ICK in lung development and advances the mechanistic understanding of ECO syndrome. © 2017 Federation of European Biochemical Societies.

Radiation damage and defect behavior in proton irradiated lithium-counterdoped n+p silicon solar cells

NASA Technical Reports Server (NTRS)

Stupica, John; Goradia, Chandra; Swartz, Clifford K.; Weinberg, Irving

1987-01-01

Two lithium-counterdoped n+p silicon solar cells with different lithium concentrations were irradiated by 10-MeV protons. Cell performance was measured as a function of fluence, and it was found that the cell with the highest concentration of lithium had the highest radiation resistance. Deep level transient spectroscopy which showed two deep level defects that were lithium related. Relating the defect energy levels obtained from this study with those from earlier work using 1-MeV electron irradiation shows no correlation of the defect energy levels. There is one marked similarity: the absence of the boron-interstitial-oxygen-interstitial defect. This consistency strengthens the belief that lithium interacts with oxygen to prevent the formation of the boron interstitial-oxygen interstitial defect. The results indicate that, in general, addition of lithium in small amounts to the p-base of a boron doped silicon solar cell such that the base remains p-type, tends to increase the radiation resistance of the cell.

Interstitial flow influences direction of tumor cell migration through competing mechanisms

PubMed Central

Polacheck, William J.; Charest, Joseph L.; Kamm, Roger D.

2011-01-01

Interstitial flow is the convective transport of fluid through tissue extracellular matrix. This creeping fluid flow has been shown to affect the morphology and migration of cells such as fibroblasts, cancer cells, endothelial cells, and mesenchymal stem cells. A microfluidic cell culture system was designed to apply stable pressure gradients and fluid flow and allow direct visualization of transient responses of cells seeded in a 3D collagen type I scaffold. We used this system to examine the effects of interstitial flow on cancer cell morphology and migration and to extend previous studies showing that interstitial flow increases the metastatic potential of MDA-MB-435S melanoma cells [Shields J, et al. (2007) Cancer Cell 11:526–538]. Using a breast carcinoma line (MDA-MB-231) we also observed cell migration along streamlines in the presence of flow; however, we further demonstrated that the strength of the flow as well as the cell density determined directional bias of migration along the streamline. In particular, we found that cells either at high seeding density or with the CCR-7 receptor inhibited migration against, rather than with the flow. We provide further evidence that CCR7-dependent autologous chemotaxis is the mechanism that leads to migration with the flow, but also demonstrate a competing CCR7-independent mechanism that causes migration against the flow. Data from experiments investigating the effects of cell concentration, interstitial flow rate, receptor activity, and focal adhesion kinase phosphorylation support our hypothesis that the competing stimulus is integrin mediated. This mechanism may play an important role in development of metastatic disease. PMID:21690404

The evaluation of interstitial Cajal cells distribution in non-tumoral colon disorders.

PubMed

Becheanu, G; Manuc, M; Dumbravă, Mona; Herlea, V; Hortopan, Monica; Costache, Mariana

2008-01-01

Interstitial cells of Cajal (ICC) are pacemakers that generate electric waves recorded from the gut and are important for intestinal motility. The aim of the study was to evaluate the distribution of interstitial cells of Cajal in colon specimens from patients with idiopathic chronic pseudo-obstruction and other non-tumoral colon disorders as compared with samples from normal colon. The distribution pattern of ICC in the normal and pathological human colon was evaluated by immunohistochemistry using antibodies for CD117, CD34, and S-100. In two cases with intestinal chronic idiopathic pseudo-obstruction we found a diffuse or focal reducing number of Cajal cells, the loss of immunoreactivity for CD117 being correlated with loss of immunoreactivity for CD34 marker. Our study revealed that the number of interstitial cells of Cajal also decrease in colonic diverticular disease and Crohn disease (p<0.05), whereas the number of enteric neurones appears to be normal. These findings might explain some of the large bowel motor abnormalities known to occur in these disorders. Interstitial Cajal cells may play an important role in pathogenesis and staining for CD117 on transmural intestinal surgical biopsies could allow a more extensive diagnosis in evaluation of chronic intestinal pseudo-obstruction.

Diazepam administration prevents testosterone decrease and lipofuscin accumulation in testis of mouse exposed to chronic noise stress.

PubMed

Ruffoli, R; Carpi, A; Giambelluca, M A; Grasso, L; Scavuzzo, M C; Giannessi F, F

2006-10-01

Lipofuscin is an autofluorescent and undegradable material, which accumulates in tissues during ageing and under different types of stress. Among these, oxidative stress represents a major trigger for lipofuscin formation. However, prolonged noise exposure is also an effective stressful stimuli. Diazepam may inhibit lipofuscinogenesis in liver and prevent the noise-induced reduction of the steroidogenesis in the adrenal gland. The aim of the study was to ascertain whether chronic noise exposure causes lipofuscin accumulation in mouse testis, and to evaluate the effects of diazepam administration. Eight-week old mice were either exposed for 6 weeks (6 h day(-1)) to white-noise (group A), or received diazepam (3 mg kg(-1), i.p.) before noise exposures (group B), while a further group was used as control (group C). Light fluorescence and transmission electron microscopy revealed lipofuscin in large amounts in the Leydig cells in mice of group A, which concomitantly had low serum testosterone levels; pre-treatment with diazepam occluded both effects. The present study indicates that: (i) chronic noise exposure causes lipofuscin accumulation at the level of the Leydig cells and a decrease in testosterone; (ii) all these effects are suppressed by pre-treatment with diazepam. As the Leydig cells represent the only cellular type of the interstitial testicular tissue having peripheral benzodiazepine receptors, these results could be explained by the capacity of the peripheral benzodiazepine receptors to prevent reactive oxygen species damage and to increase the resistance of these cells to oxidative stress.

Bevacizumab-induced chronic interstitial pneumonia during maintenance therapy in non-small cell lung cancer.

PubMed

Sekimoto, Yasuhito; Kato, Motoyasu; Shukuya, Takehiko; Koyama, Ryo; Nagaoka, Tetsutaro; Takahashi, Kazuhisa

2016-04-01

Bevacizumab is a monoclonal antibody targeting the vascular endothelial growth factor receptor and a key drug for advanced non-small cell lung cancer. There are few reports describing bevacizumab-induced chronic interstitial pneumonia. A 62-year-old man with advanced non-small cell lung cancer was admitted to our hospital with dyspnea. He previously received four courses of carboplatin plus paclitaxel with bevacizumab combination therapy and thereafter received four courses of maintenance bevacizumab monotherapy. A chest-computed tomography scan on admission revealed diffuse ground glass opacity. He had not received any other drugs and did not have pneumonia. Thus, he was diagnosed with bevacizumab-induced chronic interstitial pneumonia and was treated with a high dose of corticosteroids. After steroid treatment, his dyspnea and radiological findings improved. This case report is the first description of bevacizumab-induced chronic interstitial pneumonia during maintenance therapy in a patient with non-small cell lung cancer.

Longitudinal assessment of mouse renal injury using high-resolution anatomic and magnetization transfer MR imaging.

PubMed

Wang, Feng; Jiang, Rosie; Takahashi, Keiko; Gore, John; Harris, Raymond C; Takahashi, Takamune; Quarles, C Chad

2014-11-01

The purpose of this study is to evaluate the utility of high-resolution non-invasive endogenous high-field MRI methods for the longitudinal structural and quantitative assessments of mouse kidney disease using the model of unilateral ureter obstruction (UUO). T1-weighted, T2-weighted and magnetization transfer (MT) imaging protocols were optimized to improve the regional contrast in mouse kidney. Conventional T1 and T2 weighted images were collected in UUO mice on day 0 (~3h), day 1, day 3 and day 6 after injury, on a 7 T small animal MRI system. Cortical and medullary thickness, corticomedullary contrast and Magnetization Transfer Ratio (MTR) were assessed longitudinally. Masson trichrome staining was used to histologically assess changes in tissue microstructure. Over the course of UUO progression there were significant (p<0.05) changes in thickness of cortex and outer medulla, and regional changes in T2 signal intensity and MTR values. Histological changes included tubular cell death, tubular dilation, urine retention, and interstitial fibrosis, assessed by histology. The MRI measures of renal cortical and medullary atrophy, cortical-medullary differentiation and MTR changes provide an endogenous, non-invasive and quantitative evaluation of renal morphology and tissue composition during UUO progression. Copyright © 2014 Elsevier Inc. All rights reserved.

Proliferation and differentiation of brown adipocytes from interstitial cells during cold acclimation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bukowiecki, L.J.; Geloeen, A.; Collet, A.J.

1986-06-01

The mechanisms of brown adipocyte proliferation and differentiation during cold acclimation (and/or adaptation to hyperphagia) have been studied by quantitative photonic radioautography. (/sup 3/H)thymidine was injected to warm-acclimated (25/sup 0/C) rats and to animals exposed to 5/sup 0/C for 2 days. Samples of interscapular brown adipose tissue were collected for quantitative analysis of mitotic frequencies at various periods of time (4 h-15 days) after the injection of (/sup 3/H)thymidine, the rats being maintained at the temperatures to which they were initially exposed. It was found that cold exposure for 2 days markedly enhanced mitotic activity in endothelial cells, interstitial cells,more » and brown preadipocytes rather than in fully differentiated brown adipocytes. The total tissue labeling index (percent of labeled nuclei) increased approx.70 times over control values. The authors now report that cellular labeling progressively increased in mature brown adipocytes during cold acclimation, whereas it correspondingly decreased in interstitial cells and brown preadipocytes. This indicates that the sequence of events for cellular differentiation is interstitial cells ..-->.. brown preadipocytes ..-->.. mature brown adipocytes. Remarkable, labeling frequency did not change in endothelial cells during cold acclimation demonstrating that these cells cannot be considered as progenitors of brown adipocytes. It is suggested that brown adipocyte proliferation and differentiation from interstitial cells represent the fundamental phenomena explaining the enhanced capacity of cold-acclimated and/or hyperphagic rats to respond calorigenically to catecholamines.« less

Accumulation of vitamin A in the hepatic stellate cell of arctic top predators.

PubMed

Senoo, Haruki; Imai, Katsuyuki; Mezaki, Yoshihiro; Miura, Mitsutaka; Morii, Mayako; Fujiwara, Mutsunori; Blomhoff, Rune

2012-10-01

We performed a systematic characterization of the hepatic vitamin A storage in mammals and birds of the Svalbard Archipelago and Greenland. The liver of top predators, including polar bear, Arctic fox, bearded seal, and glaucous gull, contained about 10-20 times more vitamin A than the liver of all other arctic animals studied, as well as their genetically related continental top predators. The values are also high compared to normal human and experimental animals like mouse and rat. This massive amount of hepatic vitamin A was located in large autofluorescent lipid droplets in hepatic stellate cells (HSCs; also called vitamin A-storing cells, lipocytes, interstitial cells, fat-storing cells, or Ito cells). The droplets made up most of the cells' cytoplasm. The development of such an efficient vitamin A-storing mechanism in HSCs may have contributed to the survival of top predators in the extreme environment of the arctic. These animals demonstrated no signs of hypervitaminosis A. We suggest that HSCs have capacity to take-up and store large amounts of vitamin A, which may play a pivotal role in maintenance of the food web, food chain, biodiversity, and eventually ecology of the arctic. Copyright © 2012 Wiley Periodicals, Inc.

Progenitors of Secondary Crest Myofibroblasts are Developmentally Committed in Early Lung Mesoderm

PubMed Central

Li, Changgong; Li, Min; Li, Sha; Xing, Yiming; Yang, Chang-Yo; Li, Aimin; Borok, Zea; De Langhe, Stijn; Minoo, Parviz

2015-01-01

Development of the mammalian lung is predicated on cross-communications between two highly interactive tissues, the endodermally-derived epithelium and the mesodermally-derived pulmonary mesenchyme. While much attention has been paid the lung epithelium, the pulmonary mesenchyme, partly due to lack of specific tractable markers remains under-investigated. The lung mesenchyme is derived from the lateral plate mesoderm and is the principal recipient of Hedgehog (Hh) signaling, a morphogenetic network that regulates multiple aspects of embryonic development. Using the Hh-responsive Gli1-creERT2 mouse line, we identified the mesodermal targets of Hh signaling at various time points during embryonic and postnatal lung development. Cell lineage analysis showed these cells serve as progenitors to contribute to multiple lineages of mesodermally-derived differentiated cell types that include parenchymal or interstitial myofibroblasts, parabronchial and perivascular smooth muscle as well as rare populations of cells within the mesothelium. Most importantly, Gli1-creERT2 identified the progenitors of secondary crest myofibroblasts, a hitherto intractable cell type that plays a key role in alveolar formation, a vital process about which little is currently known. Transcriptome analysis of Hh-targeted progenitor cells transitioning from the pseudoglandular to the saccular phase of lung development revealed important modulations of key signaling pathways. Amongst these, there was significant down-regulation of canonical WNT signaling. Ectopic stabilization of β-Catenin via inactivation of Apc by Gli1-creERT2 expanded the Hh-targeted progenitor pools, which caused the formation of fibroblastic masses within the lung parenchyma. The Gli1-creERT2 mouse line represents a novel tool in the analysis of mesenchymal cell biology and alveolar formation during lung development. PMID:25448080

Blood and interstitial flow in the hierarchical pore space architecture of bone tissue.

PubMed

Cowin, Stephen C; Cardoso, Luis

2015-03-18

There are two main types of fluid in bone tissue, blood and interstitial fluid. The chemical composition of these fluids varies with time and location in bone. Blood arrives through the arterial system containing oxygen and other nutrients and the blood components depart via the venous system containing less oxygen and reduced nutrition. Within the bone, as within other tissues, substances pass from the blood through the arterial walls into the interstitial fluid. The movement of the interstitial fluid carries these substances to the cells within the bone and, at the same time, carries off the waste materials from the cells. Bone tissue would not live without these fluid movements. The development of a model for poroelastic materials with hierarchical pore space architecture for the description of blood flow and interstitial fluid flow in living bone tissue is reviewed. The model is applied to the problem of determining the exchange of pore fluid between the vascular porosity and the lacunar-canalicular porosity in bone tissue due to cyclic mechanical loading and blood pressure. These results are basic to the understanding of interstitial flow in bone tissue that, in turn, is basic to understanding of nutrient transport from the vasculature to the bone cells buried in the bone tissue and to the process of mechanotransduction by these cells. Copyright © 2014 Elsevier Ltd. All rights reserved.

Blood and Interstitial flow in the hierarchical pore space architecture of bone tissue

PubMed Central

Cowin, Stephen C.; Cardoso, Luis

2015-01-01

There are two main types of fluid in bone tissue, blood and interstitial fluid. The chemical composition of these fluids varies with time and location in bone. Blood arrives through the arterial system containing oxygen and other nutrients and the blood components depart via the venous system containing less oxygen and reduced nutrition. Within the bone, as within other tissues, substances pass from the blood through the arterial walls into the interstitial fluid. The movement of the interstitial fluid carries these substances to the cells within the bone and, at the same time, carries off the waste materials from the cells. Bone tissue would not live without these fluid movements. The development of a model for poroelastic materials with hierarchical pore space architecture for the description of blood flow and interstitial fluid flow in living bone tissue is reviewed. The model is applied to the problem of determining the exchange of pore fluid between the vascular porosity and the lacunar-canalicular porosity in bone tissue due to cyclic mechanical loading and blood pressure. These results are basic to the understanding of interstitial flow in bone tissue that, in turn, is basic to understanding of nutrient transport from the vasculature to the bone cells buried in the bone tissue and to the process of mechanotransduction by these cells. PMID:25666410

Bilateral sertoli and interstitial cell tumours in abdominal testes of a goat with polled intersex syndrome (PIS).

PubMed

Canisso, I F; Coffee, L L; Ortved, K; Fubini, S L; Monteagudo, L V; Schlafer, D H; Gilbert, R O

2014-12-01

An 8-year-old, mixed breed, polled goat was presented for evaluation of male-like behaviour. Clinical findings included clitoromegaly, a heavily muscled neck, pronounced beard, and erect dorsal guard hairs, which are phenotypic characteristics commonly observed in intersex animals. Transrectal ultrasonography revealed the presence of two abdominal masses caudolateral to the uterine horns. Serum concentration of estradiol was elevated. Genetic evaluation was compatible with polled intersex syndrome defined by an XX karyotype without a Y chromosome or SRY gene. Based on gross and histologic evaluation, the abdominal masses were determined to be intra-abdominal testes, each of which was effaced by Sertoli cell and interstitial (Leydig) cell tumours. The Sertoli cell tumours (SCTs) represented two unique histologic patterns. Regardless of pattern, neoplastic Sertoli cells were consistently lipid laden and positive for vimentin. Interstitial cell tumours (ICTs) were negative for vimentin. Clinical and histopathologic findings suggest that prolonged exposure to steroids secreted by neoplastic Sertoli cells contributed to virilization. In addition, results from immunohistochemistry indicated that vimentin may be a valuable immunodiagnostic tool for differentiation between interstitial and Sertoli cell tumours in goats. © 2014 Blackwell Verlag GmbH.

Carbon, oxygen and their interaction with intrinsic point defects in solar silicon ribbon material: A speculative approach

NASA Technical Reports Server (NTRS)

Goesele, U.; Ast, D. G.

1983-01-01

Some background information on intrinsic point defects is provided and on carbon and oxygen in silicon in so far as it may be relevant for the efficiency of solar cells fabricated from EFG ribbon material. The co-precipitation of carbon and oxygen and especially of carbon and silicon self interstitials are discussed. A simple model for the electrical activity of carbon-self-interstitial agglomerates is presented. The self-interstitial content of these agglomerates is assumed to determine their electrical activity and that both compressive stresses (high self-interstitial content) and tensile stresses (low self-interstitial content) give rise to electrical activity of the agglomerates. The self-interstitial content of these carbon-related agglomerates may be reduced by an appropriate high temperature treatment and enhanced by a supersaturation of self-interstitials generated during formation of the p-n junction of solar cells. Oxygen present in supersaturation in carbon-rich silicon may be induced to form SiO, precipitates by self-interstitials generated during phosphorus diffusion. It is proposed that the SiO2-Si interface of the precipates gives rise to a continuum of donor stables and that these interface states are responsible for at least part of the light inhancement effects observed in oxygen containing EFG silicon after phosphorus diffusion.

IL-36α Regulates Tubulointerstitial Inflammation in the Mouse Kidney.

PubMed

Ichii, Osamu; Kimura, Junpei; Okamura, Tadashi; Horino, Taro; Nakamura, Teppei; Sasaki, Hayato; Elewa, Yaser Hosny Ali; Kon, Yasuhiro

2017-01-01

IL-36α, a member of the IL-1 family, is a crucial mediator of inflammatory responses. We previously found that IL-36α was overexpressed in injured distal tubules (DTs); however, its pathological function remains unclear. Herein, unilateral ureter obstruction (UUO) or folic acid (FA) injection was performed in mouse kidneys to assess the role of IL-36α in kidney injury. IL-36α mRNA and protein expression significantly increased in the kidneys within 24 h after UUO. IL-36α localized to dilated DTs. IL-36α expression significantly correlated with the progression of tubulointerstitial cell infiltration and tubular epithelium cell death in UUO kidneys and with renal dysfunction in FA-induced acute kidney injury mice. At 24 h after UUO, IL-36α + DT epithelial cells showed loose intercellular digitations. IL-1RL2, an IL-36α receptor protein, localized to podocytes, proximal tubules, and DTs in the healthy kidney. IL-1RL2 was expressed in interstitial cells and platelets or extended primary cilia of DT epithelial cells in UUO kidneys. IL-36α stimulation promoted the production of IL-6 and Prss35, an inflammatory cytokine and collagen remodeling-associated enzyme, respectively, in cultured NIH3T3 fibroblasts. UUO-treated IL-36α-knockout (KO) mice showed milder kidney injury features than wild-type (WT) mice did. In UUO kidneys from IL-36α-KO mice, the expression of genes associated with inflammatory response and sensory perception was significantly different from that in WT mice. Altogether, our data indicate an association between intrarenal IL-36α overexpression and the progression of tubulointerstitial inflammations and morpho-functional alterations of DT epithelial cells. IL-36α may be a novel kidney injury marker useful for evaluating DT damages.

Tranilast prevents renal interstitial fibrosis by blocking mast cell infiltration in a rat model of diabetic kidney disease.

PubMed

Yin, Dan-Dan; Luo, Jun-Hui; Zhao, Zhu-Ye; Liao, Ying-Jun; Li, Ying

2018-05-01

Renal interstitial fibrosis is a final pathway that is observed in various types of kidney diseases, including diabetic kidney disease (DKD). The present study investigated the effect of tranilast on renal interstitial fibrosis and the association between its role and mast cell infiltration in a rat model of DKD. A total of 30 healthy 6‑week‑old male Sprague‑Dawley rats were randomly divided into the following four groups: Normal control group; DKD model group; low‑dose tranilast group (200 mg/kg/day); and high‑dose tranilast group (400 mg/kg/day). The morphological alterations of tubulointerstitial fibrosis were evaluated by Masson's trichrome staining, while mast cell infiltration into the renal tubular interstitium was measured by toluidine blue staining and complement C3a receptor 1 (C3aR) immunohistochemical staining (IHC). The expression of fibronectin (FN), collagen I (Col‑I), stem cell factor (SCF) and proto‑oncogene c‑kit (c‑kit) was detected by IHC, western blotting and reverse transcription‑quantitative‑polymerase chain reaction. The results demonstrated that tubulointerstitial fibrosis and mast cell infiltration were observed in DKD model rats, and this was improved dose‑dependently in the tranilast treatment groups. The expression of FN, Col‑I, SCF and c‑kit mRNA and protein was upregulated in the tubulointerstitium of DKD model rats compared with the normal control rats, and tranilast inhibited the upregulated expression of these markers. Furthermore, the degree of SCF and c‑kit expression demonstrated a significant positive correlation with C3aR‑positive mast cells and the markers of renal interstitial fibrosis. The results of the present study indicate that mast cell infiltration may promote renal interstitial fibrosis via the SCF/c‑kit signaling pathway. Tranilast may prevent renal interstitial fibrosis through inhibition of mast cell infiltration mediated through the SCF/c-kit signaling pathway.

Cancer Cell Glycocalyx Mediates Mechanostransduction and Flow-Regulated Invasion

PubMed Central

Qazi, Henry; Palomino, Rocio; Shi, Zhong-Dong; Munn, Lance L.; Tarbell, John M.

2014-01-01

Mammalian cells are covered by a surface proteoglycan (glycocalyx) layer, and it is known that blood vessel-lining endothelial cells use the glycocalyx to sense and transduce the shearing forces of blood flow into intracellular signals. Tumor cells in vivo are exposed to forces from interstitial fluid flow that may affect metastatic potential but are not reproduced by most in vitro cell motility assays. We hypothesized that glycocalyx-mediated mechanotransduction of interstitial flow shear stress is an un-recognized factor that can significantly enhance metastatic cell motility and play a role in augmentation of invasion. Involvement of MMP levels, cell adhesion molecules (CD44, α3 integrin), and glycocalyx components (heparan sulfate and hyaluronan) were investigated in a cell/collagen gel suspension model designed to mimic the interstitial flow microenvironment. Physiologic levels of flow upregulated MMP levels and enhanced the motility of metastatic cells. Blocking the flow-enhanced expression of MMP actvity or adhesion molecules (CD44 and integrins) resulted in blocking the flow-enhanced migratory activity. The presence of a glycocalyx-like layer was verified around tumor cells, and the degradation of this layer by hyaluronidase and heparinase blocked the flow-regulated invasion. This study shows for the first time that interstitial flow enhancement of metastatic cell motility can be mediated by the cell surface glycocalyx – a potential target for therapeutics. PMID:24077103

Immunohistochemical evaluation of proliferation, apoptosis and steroidogenic enzymes in the ovary of rats with polycystic ovary.

PubMed

Lombardi, Leonardo Augusto; Simões, Ricardo Santos; Maganhin, Carla Cristina; Baracat, Maria Cândida Pinheiro; Silva-Sasso, Gisela Rodrigues; Florencio-Silva, Rinaldo; Soares, José Maria; Baracat, Edmund Chada

2014-07-01

to evaluate the immunohistochemical expression of proliferative, apoptotic and steroidogenic enzyme markers in the ovaries of rats with polycystic ovary syndrome (PCOS). twenty rats were divided into two groups: GCtrl - estrous phase, and PCOS - with polycystic ovaries. The GCtrl animals were subjected to a lighting period from 7 am to 7 pm, while the animals with PCOS group remained with continuous lighting for 60 days. Subsequently, the animals were anesthetized, the ovaries were removed and fixed in 10% formaldehyde, prior to paraffin embedding. Sections were stained using H.E. or subjected to immunohistochemical methods for the detection of Ki-67, cleaved caspase-3, CYP11A1, CYP17A1 and CYP19A1. The results were analyzed using Student's t-test (p < 0,05). morphological results showed evidence of interstitial cells originating from the inner theca cells of degenerating ovarian cysts in PCOS. Immunoexpression of Ki-67 was higher in the granulosa cells in GCtrl, and the theca interna cells in PCOS, while cleaved caspase-3 was higher in granulosa cells of ovarian cysts from PCOS and in the theca interna cells of GCtrl. Immunoreactivity of CYP11A1 in the theca interna, granulosa and interstitial cells was similar between the two groups, while CYP17A1 and CYP19A1 were higher in the granulosa and interstitial cells in the PCOS group. the results indicate that the interstitial cells are derived from the theca interna and that enzymatic changes occur in the theca interna and interstitial cells in ovaries of rats with PCOS, responsible for the high levels of androgens and estradiol.

Angiotensin II type 1 and 2 receptors and lymphatic vessels modulate lung remodeling and fibrosis in systemic sclerosis and idiopathic pulmonary fibrosis.

PubMed

Parra, Edwin Roger; Ruppert, Aline Domingos Pinto; Capelozzi, Vera Luiza

2014-01-01

To validate the importance of the angiotensin II receptor isotypes and the lymphatic vessels in systemic sclerosis and idiopathic pulmonary fibrosis. We examined angiotensin II type 1 and 2 receptors and lymphatic vessels in the pulmonary tissues obtained from open lung biopsies of 30 patients with systemic sclerosis and 28 patients with idiopathic pulmonary fibrosis. Their histologic patterns included cellular and fibrotic non-specific interstitial pneumonia for systemic sclerosis and usual interstitial pneumonia for idiopathic pulmonary fibrosis. We used immunohistochemistry and histomorphometry to evaluate the number of cells in the alveolar septae and the vessels stained by these markers. Survival curves were also used. We found a significantly increased percentage of septal and vessel cells immunostained for the angiotensin type 1 and 2 receptors in the systemic sclerosis and idiopathic pulmonary fibrosis patients compared with the controls. A similar percentage of angiotensin 2 receptor positive vessel cells was observed in fibrotic non-specific interstitial pneumonia and usual interstitial pneumonia. A significantly increased percentage of lymphatic vessels was present in the usual interstitial pneumonia group compared with the non-specific interstitial pneumonia and control groups. A Cox regression analysis showed a high risk of death for the patients with usual interstitial pneumonia and a high percentage of vessel cells immunostained for the angiotensin 2 receptor in the lymphatic vessels. We concluded that angiotensin II receptor expression in the lung parenchyma can potentially control organ remodeling and fibrosis, which suggests that strategies aimed at preventing high angiotensin 2 receptor expression may be used as potential therapeutic target in patients with pulmonary systemic sclerosis and idiopathic pulmonary fibrosis.

Augmented sphingosine 1 phosphate receptor-1 signaling in cardiac fibroblasts induces cardiac hypertrophy and fibrosis through angiotensin II and interleukin-6

PubMed Central

Ohkura, Sei-ichiro; Takashima, Shin-ichiro; Yoshioka, Kazuaki; Okamoto, Yasuo; Inagaki, Yutaka; Sugimoto, Naotoshi; Kitano, Teppei; Takamura, Masayuki; Wada, Takashi; Kaneko, Shuichi; Takuwa, Yoh

2017-01-01

Background: Cardiac fibroblasts, together with cardiomyocytes, occupy the majority of cells in the myocardium and are involved in myocardial remodeling. The lysophospholipid mediator sphigosine-1-phosphate (S1P) regulates functions of cardiovascular cells through multiple receptors including S1PR1–S1PR3. S1PR1 but not other S1P receptors was upregulated in angiotensin II-induced hypertrophic hearts. Therefore, we investigated a role of S1PR1 in fibroblasts for cardiac remodeling by employing transgenic mice that overexpressed S1PR1 under the control of α-smooth muscle actin promoter. In S1PR1-transgenic mouse heart, fibroblasts and/or myofibroblasts were hyperplastic, and those cells as well as vascular smooth muscle cells overexpressed S1PR1. Transgenic mice developed bi-ventricular hypertrophy by 12-week-old and diffuse interstitial fibrosis by 24-week-old without hemodynamic stress. Cardiac remodeling in transgenic mice was associated with greater ERK phosphorylation, upregulation of fetal genes, and systolic dysfunction. Transgenic mouse heart showed increased mRNA expression of angiotensin-converting enzyme and interleukin-6 (IL-6). Isolated fibroblasts from transgenic mice exhibited enhanced generation of angiotensin II, which in turn stimulated IL-6 release. Either an AT1 blocker or angiotensin-converting enzyme inhibitor prevented development of cardiac hypertrophy and fibrosis, systolic dysfunction and increased IL-6 expression in transgenic mice. Finally, administration of anti-IL-6 antibody abolished an increase in tyrosine phosphorylation of STAT3, a major signaling molecule downstream of IL-6, in the transgenic mouse heart and prevented development of cardiac hypertrophy in transgenic mice. These results demonstrate a promoting role of S1PR1 in cardiac fibroblasts for cardiac remodeling, in which angiotensin II—AT1 and IL-6 are involved. PMID:28771545

Inflammation and Fibrosis in Polycystic Kidney Disease.

PubMed

Song, Cheng Jack; Zimmerman, Kurt A; Henke, Scott J; Yoder, Bradley K

Polycystic kidney disease (PKD) is a commonly inherited disorder characterized by cyst formation and fibrosis (Wilson, N Engl J Med 350:151-164, 2004) and is caused by mutations in cilia or cilia-related proteins, such as polycystin 1 or 2 (Oh and Katsanis, Development 139:443-448, 2012; Kotsis et al., Nephrol Dial Transplant 28:518-526, 2013). A major pathological feature of PKD is the development of interstitial inflammation and fibrosis with an associated accumulation of inflammatory cells (Grantham, N Engl J Med 359:1477-1485, 2008; Zeier et al., Kidney Int 42:1259-1265, 1992; Ibrahim, Sci World J 7:1757-1767, 2007). It is unclear whether inflammation is a driving force for cyst formation or a consequence of the pathology (Ta et al., Nephrology 18:317-330, 2013) as in some murine models cysts are present prior to the increase in inflammatory cells (Phillips et al., Kidney Blood Press Res 30:129-144, 2007; Takahashi et al., J Am Soc Nephrol JASN 1:980-989, 1991), while in other models the increase in inflammatory cells is present prior to or coincident with cyst initiation (Cowley et al., Kidney Int 43:522-534, 1993, Kidney Int 60:2087-2096, 2001). Additional support for inflammation as an important contributor to cystic kidney disease is the increased expression of many pro-inflammatory cytokines in murine models and human patients with cystic kidney disease (Karihaloo et al., J Am Soc Nephrol JASN 22:1809-1814, 2011; Swenson-Fields et al., Kidney Int, 2013; Li et al., Nat Med 14:863-868, 2008a). Based on these data, an emerging model in the field is that disruption of primary cilia on tubule epithelial cells leads to abnormal cytokine cross talk between the epithelium and the inflammatory cells contributing to cyst growth and fibrosis (Ta et al., Nephrology 18:317-330, 2013). These cytokines are produced by interstitial fibroblasts, inflammatory cells, and tubule epithelial cells and activate multiple pathways including the JAK-STAT and NF-κB signaling (Qin et al., J Am Soc Nephrol JASN 23:1309-1318, 2012; Park et al., Am J Nephrol 32:169-178, 2010; Bhunia et al., Cell 109:157-168, 2002). Indeed, inflammatory cells are responsible for producing several of the pro-fibrotic growth factors observed in PKD patients with fibrosis (Nakamura et al., Am J Nephrol 20:32-36, 2000; Wilson et al., J Cell Physiol 150:360-369, 1992; Song et al., Hum Mol Genet 18:2328-2343, 2009; Schieren et al., Nephrol Dial Transplant 21:1816-1824, 2006). These growth factors trigger epithelial cell proliferation and myofibroblast activation that stimulate the production of extracellular matrix (ECM) genes including collagen types 1 and 3 and fibronectin, leading to reduced glomerular function with approximately 50% of ADPKD patients progressing to end-stage renal disease (ESRD). Therefore, treatments designed to reduce inflammation and slow the rate of fibrosis are becoming important targets that hold promise to improve patient life span and quality of life. In fact, recent studies in several PKD mouse models indicate that depletion of macrophages reduces cyst severity. In this chapter, we review the potential mechanisms of interstitial inflammation in PKD with a focus on ADPKD and discuss the role of interstitial inflammation in progression to fibrosis and ESRD.

Modeling and Measurement of Correlation between Blood and Interstitial Glucose Changes

PubMed Central

Shi, Ting; Li, Dachao; Li, Guoqing; Zhang, Yiming; Xu, Kexin; Lu, Luo

2016-01-01

One of the most effective methods for continuous blood glucose monitoring is to continuously measure glucose in the interstitial fluid (ISF). However, multiple physiological factors can modulate glucose concentrations and affect the lag phase between blood and ISF glucose changes. This study aims to develop a compensatory tool for measuring the delay in ISF glucose variations in reference to blood glucose changes. A theoretical model was developed based on biophysics and physiology of glucose transport in the microcirculation system. Blood and interstitial fluid glucose changes were measured in mice and rats by fluorescent and isotope methods, respectively. Computer simulation mimicked curves were fitted with data resulting from fluorescent measurements of mice and isotope measurements of rats, indicating that there were lag times for ISF glucose changes. It also showed that there was a required diffusion distance for glucose to travel from center of capillaries to interstitial space in both mouse and rat models. We conclude that it is feasible with the developed model to continuously monitor dynamic changes of blood glucose concentration through measuring glucose changes in ISF with high accuracy, which requires correct parameters for determining and compensating for the delay time of glucose changes in ISF. PMID:27239479

Single or group housing altered hormonal physiology and affected pituitary and interstitial cell kinetics

EPA Science Inventory

A significant negative correlation between testicular interstitial cell tumors and pituitary tumors in control male F344 rats has been reported associated with the number of animals per cage. Change in numbers of animals per cage may cause stress and increased serum corticosteroi...

Localization and expression of Orexin A and its receptor in mouse testis during different stages of postnatal development.

PubMed

Joshi, Deepanshu; Singh, Shio Kumar

2017-01-15

Orexin A (OXA), a hypothalamic neuropeptide, is involved in regulation of various biological functions and its actions are mediated through G-protein-coupled receptor, OX1R. This neuropeptide has emerged as a central neuroendocrine modulator of reproductive functions. Both OXA and OX1R have been shown to be expressed in peripheral organs such as gastrointestinal and genital tracts. In the present study, localization and expression of OXA and OX1R in mouse testis during different stages of postnatal development have been investigated. Immunohistochemical results demonstrated localization of OXA and OX1R in both the interstitial and the tubular compartments of the testis throughout the period of postnatal development. In testicular sections on 0day postpartum (dpp), gonocytes, Sertoli cells and foetal Leydig cells showed OXA and OX1R-immunopositive signals. At 10dpp, Sertoli cells, spermatogonia, early spermatocytes and Leydig cells showed immunopositive signals for both, the ligand and the receptor. On 30 and 90dpp, the spermatogonia, Sertoli cells, spermatocytes, spermatids and Leydig cells showed the OXA and OX1R-immunopositive signals. At 90dpp, strong OXA-positive signals were seen in Leydig cells, primary spermatocytes and spermatogonia, while OX1R-immunopositive intense signals were observed in Leydig cells and elongated spermatids. Further, semiquantitative RT-PCR and immunoblot analyses showed that OXA and OX1R were expressed in the testis both at transcript and protein levels during different stages of postnatal development. The expression of OXA and OX1R increased progressively from day of birth (0dpp) until adulthood (90dpp), with maximal expression at 90 dpp. The results suggest that OXA and OX1R are expressed in the testis and that they may help in proliferation and development of germ cells, Leydig cells and Sertoli cells, and in the spermatogenic process and steroidogenesis. Copyright © 2016 Elsevier Inc. All rights reserved.

Dynamical optical imaging monocytes/macrophages migration and activation in contact hypersensitivity (Conference Presentation)

NASA Astrophysics Data System (ADS)

Zhang, Zhihong

2017-02-01

Inflammatory monocytes/macrophages (Mon/Mφ) play an important role in cutaneous allergic inflammation. However, their migration and activation in dermatitis and how they accelerate the inflammatory reaction are largely unknown. Optical molecular imaging is the most promising tool for investigating the function and motility of immune cells in vivo. We have developed a multi-scale optical imaging approach to evaluate the spatio-temporal dynamic behavior and properties of immune cells from the whole field of organs to the cellular level at the inflammatory site in delayed type hypersensitivity reaction. Here, we developed some multi-color labeling mouse models based on the endogenous labeling with fluorescent proteins and the exogenous labeling with fluorescent dyes. We investigated the cell movement, cell interaction and function of immunocytes (e.g. Mon/Mφ, DC, T cells and neutrophils) in the skin allergy inflammation (e.g., contact hypersensitivity) by using intravital microscopy. The long-term imaging data showed that after inflammatory Mon/Mφ transendothelial migration in dermis, they migrating in interstitial space of dermis. Depletion of blood monocyte with clodronate liposome extremely reduced the inflammatory reaction. Our finding provided further insight into inflammatory Mon/Mφ mediating the inflammatory cascade through functional migration in allergic contact dermatitis.

An abundant tissue macrophage population in the adult murine heart with a distinct alternatively-activated macrophage profile.

PubMed

Pinto, Alexander R; Paolicelli, Rosa; Salimova, Ekaterina; Gospocic, Janko; Slonimsky, Esfir; Bilbao-Cortes, Daniel; Godwin, James W; Rosenthal, Nadia A

2012-01-01

Cardiac tissue macrophages (cTMs) are a previously uncharacterised cell type that we have identified and characterise here as an abundant GFP(+) population within the adult Cx(3)cr1(GFP/+) knock-in mouse heart. They comprise the predominant myeloid cell population in the myocardium, and are found throughout myocardial interstitial spaces interacting directly with capillary endothelial cells and cardiomyocytes. Flow cytometry-based immunophenotyping shows that cTMs exhibit canonical macrophage markers. Gene expression analysis shows that cTMs (CD45(+)CD11b(+)GFP(+)) are distinct from mononuclear CD45(+)CD11b(+)GFP(+) cells sorted from the spleen and brain of adult Cx(3)cr1(GFP/+) mice. Gene expression profiling reveals that cTMs closely resemble alternatively-activated anti-inflammatory M2 macrophages, expressing a number of M2 markers, including Mrc1, CD163, and Lyve-1. While cTMs perform normal tissue macrophage homeostatic functions, they also exhibit a distinct phenotype, involving secretion of salutary factors (including IGF-1) and immune modulation. In summary, the characterisation of cTMs at the cellular and molecular level defines a potentially important role for these cells in cardiac homeostasis.

Ava[L-Pro9,N-MeLeu10] substance P(7-11) (GR 73632) and Sar9, Met(O2)11 increase distention-induced peristalsis through activation of neurokinin-1 receptors on smooth muscle and interstitial cells of cajal.

PubMed

Nieuwmeyer, Florentine; Ye, Jing; Huizinga, Jan D

2006-04-01

Substance P is generally considered an excitatory neurotransmitter related to gut motor activity, although an inhibitory influence of neurokinin-1 (NK1) receptor activation on peristalsis has also been reported. With an optimized in vitro method to assess distention-induced peristalsis, our aim was to clarify the effect of NK1 receptor activation on peristaltic activity and to reveal the mechanisms by which NK1 activation alters peristalsis. Distention of the small intestine of the mouse and guinea pig induced periodic occurrence of rhythmic waves of propagating rings of circular muscle contraction, associated with slow waves and superimposed action potentials, that propelled intestinal contents aborally. Activation of NK1 receptors by Ava[l-Pro(9),N-MeLeu10] substance P(7-11) (GR 73632) and Sar(9), Met(O(2))(11) on smooth muscle cells resulted in prolongation of the activity periods and increased action potential generation occurring superimposed on the intestinal slow wave activity. Activation of NK1 receptors on interstitial cells of Cajal resulted in an increase in slow wave frequency. Slow wave amplitude increased, likely by increased cell-to-cell coupling. The NK1 antagonist (S)-1-(2-[3-(3,4-dichlorophenyl)-1-(3-isopropoxyphenylacetyl)piperidin-3-yl]ethyl)-4-phenyl-1-azoniabicyclo[2.2.2]octane chloride (SR 140333) induced a decrease in the slow wave frequency and duration of the activity periods evoked by distention, which makes it likely that NK1 receptor activation plays a role in the normal physiological distention-induced generation of peristaltic motor patterns. In summary, NK1 receptors play a role in normal development of peristalsis and NK1 receptor activation markedly increases propulsive peristaltic contractile activity.

Potassium and ANO1/TMEM16A chloride channel profiles distinguish atypical and typical smooth muscle cells from interstitial cells in the mouse renal pelvis

PubMed Central

Iqbal, Javed; Tonta, Mary A; Mitsui, Retsu; Li, Qun; Kett, Michelle; Li, Jinhua; Parkington, Helena C; Hashitani, Hikaru; Lang, Richard J

2012-01-01

BACKGROUND AND PURPOSE Although atypical smooth muscle cells (SMCs) in the proximal renal pelvis are thought to generate the pacemaker signals that drive pyeloureteric peristalsis, their location and electrical properties remain obscure. EXPERIMENTAL APPROACH Standard patch clamp, intracellular microelectrode and immunohistochemistry techniques were used. To unequivocally identify SMCs, transgenic mice with enhanced yellow fluorescent protein (eYFP) expressed in cells containing α-smooth muscle actin (α-SMA) were sometimes used. KEY RESULTS Atypical SMCs were distinguished from typical SMCs by the absence of both a transient 4-aminopyridine-sensitive K+ current (IKA) and spontaneous transient outward currents (STOCs) upon the opening of large-conductance Ca2+-activated K+ (BK) channels. Many typical SMCs displayed a slowly activating, slowly decaying Cl- current blocked by niflumic acid (NFA). Immunostaining for KV4.3 and ANO1/ TMEM16A Cl- channel subunits co-localized with α-SMA immunoreactive product predominately in the distal renal pelvis. Atypical SMCs fired spontaneous inward currents that were either selective for Cl- and blocked by NFA, or cation-selective and blocked by La3+. α-SMA- interstitial cells (ICs) were distinguished by the presence of a Xe991-sensitive KV7 current, BK channel STOCs and Cl- selective, NFA-sensitive spontaneous transient inward currents (STICs). Intense ANO1/ TMEM16A and KV7.5 immunostaining was present in Kit-α-SMA- ICs in the suburothelial and adventitial regions of the renal pelvis. CONCLUSIONS AND IMPLICATIONS We conclude that KV4.3+α-SMA+ SMCs are typical SMCs that facilitate muscle wall contraction, that ANO1/ TMEM16A and KV7.5 immunoreactivity may be selective markers of Kit- ICs and that atypical SMCs which discharge spontaneous inward currents are the pelviureteric pacemakers. PMID:22014103

Endothelial Dysfunction Exacerbates Renal Interstitial Fibrosis through Enhancing Fibroblast Smad3 Linker Phosphorylation in the Mouse Obstructed Kidney

PubMed Central

Sun, Yu Bo Yang; Qu, Xinli; Li, Xueling; Nikolic-Paterson, David J.; Li, Jinhua

2013-01-01

Endothelial dysfunction and enhanced transforming growth factor-β (TGF-β)/Smad3 signalling are common features of progressive renal fibrosis. This study investigated a potential link between these mechanisms. In unilateral ureteric obstruction (UUO) we observed an acute (6 hr) down-regulation of nitric oxide synthase 3 (NOS3/eNOS) levels and increased phosphorylation of the linker region of Smad3 at T179 and S208 in Smad3/JNK complexes. These events preceded Smad3 C-terminal domain phosphorylation and the induction of myofibroblast proliferation at 48 hrs. Mice deficient in NOS3 showed enhanced myofibroblast proliferation and collagen accumulation compared to wild type mice in a 7 day UUO model. This was associated with enhanced phosphorylation of Smad3 T179 and S208 by 92% and 88%, respectively, whereas Smad3-C-terminal phosphorylation was not affected. Resolvin D1 (RvD1) can suppress renal fibrosis in the UUO model, and further analysis herein showed that RvD1 protected against endothelial dysfunction and suppressed Smad3/JNK complex formation with a consequent reduction in phosphorylation of Smad3 T179 and S208 by 78% and 65%, respectively, while Smad3 C-terminal phosphorylation was unaltered. In vitro, conditioned media from mouse microvascular endothelial cells (MMEC) treated with a general inhibitor of nitric oxide synthase (L-NAME) augmented the proliferation and collagen production of renal fibroblasts (NRK49F cells) compared to control MMEC media and this was associated with increased phosphorylation of JNK and Smad3 T179 and S208, whereas Smad3-C-terminal domain phosphorylation was unaffected. The addition of RvD1 to L-NAME treated MMEC abrogated these effects of the conditioned media on renal fibroblasts. Finally, Smad3 T179/V and S208/A mutations significantly inhibit TGF-β1 induced up-regulation collagen I promoter. In conclusion, these data suggest that endothelial dysfunction can exacerbate renal interstitial fibrosis through increased fibroblast proliferation and collagen production via enhanced Smad3 linker phosphorylation. PMID:24391884

Endothelial dysfunction exacerbates renal interstitial fibrosis through enhancing fibroblast Smad3 linker phosphorylation in the mouse obstructed kidney.

PubMed

Sun, Yu Bo Yang; Qu, Xinli; Li, Xueling; Nikolic-Paterson, David J; Li, Jinhua

2013-01-01

Endothelial dysfunction and enhanced transforming growth factor-β (TGF-β)/Smad3 signalling are common features of progressive renal fibrosis. This study investigated a potential link between these mechanisms. In unilateral ureteric obstruction (UUO) we observed an acute (6 hr) down-regulation of nitric oxide synthase 3 (NOS3/eNOS) levels and increased phosphorylation of the linker region of Smad3 at T179 and S208 in Smad3/JNK complexes. These events preceded Smad3 C-terminal domain phosphorylation and the induction of myofibroblast proliferation at 48 hrs. Mice deficient in NOS3 showed enhanced myofibroblast proliferation and collagen accumulation compared to wild type mice in a 7 day UUO model. This was associated with enhanced phosphorylation of Smad3 T179 and S208 by 92% and 88%, respectively, whereas Smad3-C-terminal phosphorylation was not affected. Resolvin D1 (RvD1) can suppress renal fibrosis in the UUO model, and further analysis herein showed that RvD1 protected against endothelial dysfunction and suppressed Smad3/JNK complex formation with a consequent reduction in phosphorylation of Smad3 T179 and S208 by 78% and 65%, respectively, while Smad3 C-terminal phosphorylation was unaltered. In vitro, conditioned media from mouse microvascular endothelial cells (MMEC) treated with a general inhibitor of nitric oxide synthase (L-NAME) augmented the proliferation and collagen production of renal fibroblasts (NRK49F cells) compared to control MMEC media and this was associated with increased phosphorylation of JNK and Smad3 T179 and S208, whereas Smad3-C-terminal domain phosphorylation was unaffected. The addition of RvD1 to L-NAME treated MMEC abrogated these effects of the conditioned media on renal fibroblasts. Finally, Smad3 T179/V and S208/A mutations significantly inhibit TGF-β1 induced up-regulation collagen I promoter. In conclusion, these data suggest that endothelial dysfunction can exacerbate renal interstitial fibrosis through increased fibroblast proliferation and collagen production via enhanced Smad3 linker phosphorylation.

Characterization of Nestin-positive stem Leydig cells as a potential source for the treatment of testicular Leydig cell dysfunction

PubMed Central

Jiang, Mei Hua; Cai, Bing; Tuo, Ying; Wang, Jiancheng; Zang, Zhi Jun; Tu, Xiang'an; Gao, Yong; Su, Zhijian; Li, Weiqiang; Li, Guilan; Zhang, Min; Jiao, Jianwei; Wan, Zi; Deng, Chunhua; Lahn, Bruce T; Xiang, Andy Peng

2014-01-01

The ability to identify and isolate lineage-specific stem cells from adult tissues could facilitate cell replacement therapy. Leydig cells (LCs) are the primary source of androgen in the mammalian testis, and the prospective identification of stem Leydig cells (SLCs) may offer new opportunities for treating testosterone deficiency. Here, in a transgenic mouse model expressing GFP driven by the Nestin (Nes) promoter, we observed Nes-GFP+ cells located in the testicular interstitial compartment where SLCs normally reside. We showed that these Nes-GFP+ cells expressed LIFR and PDGFR-α, but not LC lineage markers. We further observed that these cells were capable of clonogenic self-renewal and extensive proliferation in vitro and could differentiate into neural or mesenchymal cell lineages, as well as LCs, with the ability to produce testosterone, under defined conditions. Moreover, when transplanted into the testes of LC-disrupted or aging models, the Nes-GFP+ cells colonized the interstitium and partially increased testosterone production, and then accelerated meiotic and post-meiotic germ cell recovery. In addition, we further demonstrated that CD51 might be a putative cell surface marker for SLCs, similar with Nestin. Taken together, these results suggest that Nes-GFP+ cells from the testis have the characteristics of SLCs, and our study would shed new light on developing stem cell replacement therapy for testosterone deficiency. PMID:25418539

Loss of neurogenesis in Hydra leads to compensatory regulation of neurogenic and neurotransmission genes in epithelial cells

PubMed Central

2016-01-01

Hydra continuously differentiates a sophisticated nervous system made of mechanosensory cells (nematocytes) and sensory–motor and ganglionic neurons from interstitial stem cells. However, this dynamic adult neurogenesis is dispensable for morphogenesis. Indeed animals depleted of their interstitial stem cells and interstitial progenitors lose their active behaviours but maintain their developmental fitness, and regenerate and bud when force-fed. To characterize the impact of the loss of neurogenesis in Hydra, we first performed transcriptomic profiling at five positions along the body axis. We found neurogenic genes predominantly expressed along the central body column, which contains stem cells and progenitors, and neurotransmission genes predominantly expressed at the extremities, where the nervous system is dense. Next, we performed transcriptomics on animals depleted of their interstitial cells by hydroxyurea, colchicine or heat-shock treatment. By crossing these results with cell-type-specific transcriptomics, we identified epithelial genes up-regulated upon loss of neurogenesis: transcription factors (Dlx, Dlx1, DMBX1/Manacle, Ets1, Gli3, KLF11, LMX1A, ZNF436, Shox1), epitheliopeptides (Arminins, PW peptide), neurosignalling components (CAMK1D, DDCl2, Inx1), ligand-ion channel receptors (CHRNA1, NaC7), G-Protein Coupled Receptors and FMRFRL. Hence epitheliomuscular cells seemingly enhance their sensing ability when neurogenesis is compromised. This unsuspected plasticity might reflect the extended multifunctionality of epithelial-like cells in early eumetazoan evolution. PMID:26598723

Loss of neurogenesis in Hydra leads to compensatory regulation of neurogenic and neurotransmission genes in epithelial cells.

PubMed

Wenger, Y; Buzgariu, W; Galliot, B

2016-01-05

Hydra continuously differentiates a sophisticated nervous system made of mechanosensory cells (nematocytes) and sensory-motor and ganglionic neurons from interstitial stem cells. However, this dynamic adult neurogenesis is dispensable for morphogenesis. Indeed animals depleted of their interstitial stem cells and interstitial progenitors lose their active behaviours but maintain their developmental fitness, and regenerate and bud when force-fed. To characterize the impact of the loss of neurogenesis in Hydra, we first performed transcriptomic profiling at five positions along the body axis. We found neurogenic genes predominantly expressed along the central body column, which contains stem cells and progenitors, and neurotransmission genes predominantly expressed at the extremities, where the nervous system is dense. Next, we performed transcriptomics on animals depleted of their interstitial cells by hydroxyurea, colchicine or heat-shock treatment. By crossing these results with cell-type-specific transcriptomics, we identified epithelial genes up-regulated upon loss of neurogenesis: transcription factors (Dlx, Dlx1, DMBX1/Manacle, Ets1, Gli3, KLF11, LMX1A, ZNF436, Shox1), epitheliopeptides (Arminins, PW peptide), neurosignalling components (CAMK1D, DDCl2, Inx1), ligand-ion channel receptors (CHRNA1, NaC7), G-Protein Coupled Receptors and FMRFRL. Hence epitheliomuscular cells seemingly enhance their sensing ability when neurogenesis is compromised. This unsuspected plasticity might reflect the extended multifunctionality of epithelial-like cells in early eumetazoan evolution. © 2015 The Authors.

Mesenchymal stem cell-derived inflammatory fibroblasts mediate interstitial fibrosis in the aging heart.

PubMed

Trial, JoAnn; Entman, Mark L; Cieslik, Katarzyna A

2016-02-01

Pathologic fibrosis in the aging mouse heart is associated with dysregulated resident mesenchymal stem cells (MSC) arising from reduced stemness and aberrant differentiation into dysfunctional inflammatory fibroblasts. Fibroblasts derived from aging MSC secrete higher levels of 1) collagen type 1 (Col1) that directly contributes to fibrosis, 2) monocyte chemoattractant protein-1 (MCP-1) that attracts leukocytes from the blood and 3) interleukin-6 (IL-6) that facilitates transition of monocytes into myeloid fibroblasts. The transcriptional activation of these proteins is controlled via the farnesyltransferase (FTase)-Ras-Erk pathway. The intrinsic change in the MSC phenotype acquired by advanced age is specific for the heart since MSC originating from bone wall (BW-MSC) or fibroblasts derived from them were free of these defects. The potential therapeutic interventions other than clinically approved strategies based on findings presented in this review are discussed as well. This article is a part of a Special Issue entitled "Fibrosis and Myocardial Remodeling". Copyright © 2015 Elsevier Ltd. All rights reserved.

Seed-competent HMW tau species accumulates in the cerebrospinal fluid of Alzheimer's disease mouse model and human patients

PubMed Central

Takeda, Shuko; Commins, Caitlin; DeVos, Sarah L.; Nobuhara, Chloe K.; Wegmann, Susanne; Roe, Allyson D.; Costantino, Isabel; Fan, Zhanyun; Nicholls, Samantha B.; Sherman, Alexis E.; Trisini Lipsanopoulos, Ana T.; Scherzer, Clemens R.; Carlson, George A.; Pitstick, Rose; Peskind, Elaine R.; Raskind, Murray A.; Li, Ge; Montine, Thomas J.; Frosch, Matthew P.; Hyman, Bradley T.

2016-01-01

Objective Cerebrospinal fluid (CSF) tau is an excellent surrogate marker for assessing neuropathological changes that occur in Alzheimer's disease (AD) patients. However, whether the elevated tau in AD CSF is just a marker of neurodegeneration or in fact a part of the disease process is uncertain. Moreover, it is unknown how CSF tau relates to the recently described soluble high-molecular-weight (HMW) species that is found in postmortem AD brain and can be taken up by neurons and seed aggregates. Methods We have examined seeding and uptake properties of brain extracellular tau from various sources including: interstitial fluid (ISF) and CSF from an AD transgenic mouse model, and postmortem ventricular and antemortem lumbar CSF from AD patients. Results We found that brain ISF and CSF tau from the AD mouse model can be taken up by cells and induce intracellular aggregates. Ventricular CSF from AD patients contained a rare HMW tau species that exerted a higher seeding activity. Notably, the HMW tau species was also detected in lumbar CSF from AD patients and its levels were significantly elevated compared with control subjects. HMW tau derived from CSF of AD patients was seed-competent in vitro. Interpretation These findings suggest that CSF from an AD brain contains potentially bioactive HMW tau species giving new insights into the role of CSF tau and biomarker development for AD. PMID:27351289

Intranasal Oxytocin for the Treatment of Pain Associated with Interstitial Cystitis

DTIC Science & Technology

2014-09-01

THIS PAGE U UU 8 19b. TELEPHONE NUMBER (include area code ) Table of Contents...electrical nerve stimulation, changes in diet, cessation in smoking, exercise, bladder training, physical therapy, and surgery . Unfortunately...Matzuk MM, Insel TR (2000) Infant vocalization , adult aggression, and fear behavior of an oxytocin null mutant mouse. Horm Behav 37:145–155.

Reduction in spontaneous firing of mouse excitatory layer 4 cortical neurons following visual classical conditioning

NASA Astrophysics Data System (ADS)

Bekisz, Marek; Shendye, Ninad; Raciborska, Ida; Wróbel, Andrzej; Waleszczyk, Wioletta J.

2017-08-01

The process of learning induces plastic changes in neuronal network of the brain. Our earlier studies on mice showed that classical conditioning in which monocular visual stimulation was paired with an electric shock to the tail enhanced GABA immunoreactivity within layer 4 of the monocular part of the primary visual cortex (V1), contralaterally to the stimulated eye. In the present experiment we investigated whether the same classical conditioning paradigm induces changes of neuronal excitability in this cortical area. Two experimental groups were used: mice that underwent 7-day visual classical conditioning and controls. Patch-clamp whole-cell recordings were performed from ex vivo slices of mouse V1. The slices were perfused with the modified artificial cerebrospinal fluid, the composition of which better mimics the brain interstitial fluid in situ and induces spontaneous activity. The neuronal excitability was characterized by measuring the frequency of spontaneous action potentials. We found that layer 4 star pyramidal cells located in the monocular representation of the "trained" eye in V1 had lower frequency of spontaneous activity in comparison with neurons from the same cortical region of control animals. Weaker spontaneous firing indicates decreased general excitability of star pyramidal neurons within layer 4 of the monocular representation of the "trained" eye in V1. Such effect could result from enhanced inhibitory processes accompanying learning in this cortical area.

Enhancement of CD147 on M1 macrophages induces differentiation of Th17 cells in the lung interstitial fibrosis.

PubMed

Geng, Jie-jie; Zhang, Kui; Chen, Li-na; Miao, Jin-lin; Yao, Meng; Ren, Ying; Fu, Zhi-guang; Chen, Zhi-nan; Zhu, Ping

2014-09-01

Lung interstitial fibrosis is a chronic lung disease, and few effective therapies are available to halt or reverse the progression of the disease. In murine and human lung fibrosis, the expression of CD147 is increased. However, the role of CD147 in lung fibrosis has not been identified, and it remains to be determined whether lung fibrosis would be improved by decreasing the expression of CD147. A murine bleomycin-induced lung interstitial fibrosis model was used in the experiments, and HAb18 mAbs and CsA were administered during the induction of lung fibrosis. In our study, we found that the HAb18 mAbs markedly reduced the collagen score and down-regulated M1 macrophages and Th17 cells. In vitro, flow cytometry analysis showed that M1 macrophages induced higher Th17 differentiation than M2 macrophages. After treatment with HAb18 mAbs or after reducing the expression of CD147 by lentivirus interference in M1 macrophages, the level of Th17 cells were significantly inhibited. In conclusion, HAb18 mAbs or CsA treatment ameliorates lung interstitial fibrosis. CD147 promoted M1 macrophage and induced the differentiation of Th17 cells in lung interstitial fibrosis, perhaps by regulating some cytokines such as IL-6, IL-1β, IL-12 and IL-23. These results indicated that CD147 may play an important role in the development of lung interstitial fibrosis. Copyright © 2014 Elsevier B.V. All rights reserved.

Is the appearance of macrophages in pulmonary tissue related to time of asphyxia?

PubMed

Vacchiano, G; D'Armiento, F; Torino, R

2001-01-01

In order to connect the appearance of macrophages and giant cells in pulmonary tissue with the time of asphyxia the authors analyzed 50 asphyxiated human lungs paying their attention on the number of alveolar and interstitial macrophages and giant cells. They compared histological specimens of 25 asphixiated humans lungs following a slow asphyxia (30 min or more) with 25 histological specimens of asphyxiated human lungs following a rapid asphyxia (10-15 min). Alveolar and interstitial macrophages and giant cells per section, were considered and numbered. Controls were done on histological examination of traumatized lungs. In the pulmonary alveoli following on acute asphyxia there were 27.7+/-4.4 macrophages per section. Subjects dead after a slow asphyxiation showed 68.2+/-7.1 alveolar macrophages per section (p<0.001). Interstitial macrophages were also frequently present. No differences are detectable in the number of polynuclear giant cells between rapidly and slowly asphyxiated human lungs. The number of alveolar and interstitial macrophages per section can be considered as a further histological evidence of a slow asphyxia and can differentiate a slow asphyxia from an acute one.

A distinct type of cell in myocardium: interstitial Cajal-like cells (ICLCs)

PubMed Central

Kostin, S; Popescu, L M

2009-01-01

Abstract The existence of a novel type of interstitial cells in the heart, interstitial Cajal-like cells (ICLCs), had been described for the first time in 2005. Their identification was mainly based on ultrastructural criteria: very long (tens up to hundreds of micrometres) and moniliform prolongations, which are extremely thin (less than 0.2 μm), below the resolving power of light microscopy. Myocardial ICLCs were also identified by methylene-blue vital staining, silver impregnation, and immunoreactivity for CD 34, vimentin, CD117/c-kit, etc. Although a series of studies provided evidence for the existence of ICLCs in human atria and rat ventricles, further investigations in other laboratories, using additional techniques, are required to substantiate the consistency of these findings. Here we provide further evidence for the existence of ICLCs in human and mammalian hearts (by transmission and scanning electron microscopy, as well as confocal laser scanning microscopy). Noteworthy, we confirm that ICLCs communicate with neighbouring cells via shedding (micro)vesicles. Although these so-called ICLCs represent a distinct type of cells, different from classical interstitial cells of Cajal, or fibroblasts, their role(s) in myocardium remain(s) to be established. Several hypotheses are proposed: (i) adult stromal (mesenchymal) stem cells, which might participate in cardiac repair/remodelling; (ii) intercellular signalling (e.g. via shedding microvesicles); (iii) chemo-mechanical transducers and (iv) players in pacemaking and/or arrhytmogenesis, and so on. PMID:19183408

Teaching a changing paradigm in physiology: a historical perspective on gut interstitial cells.

PubMed

Drumm, Bernard T; Baker, Salah A

2017-03-01

The study and teaching of gastrointestinal (GI) physiology necessitates an understanding of the cellular basis of contractile and electrical coupling behaviors in the muscle layers that comprise the gut wall. Our knowledge of the cellular origin of GI motility has drastically changed over the last 100 yr. While the pacing and coordination of GI contraction was once thought to be solely attributable to smooth muscle cells, it is now widely accepted that the motility patterns observed in the GI tract exist as a result of a multicellular system, consisting of not only smooth muscle cells but also enteric neurons and distinct populations of specialized interstitial cells that all work in concert to ensure proper GI functions. In this historical perspective, we focus on the emerging role of interstitial cells in GI motility and examine the key discoveries and experiments that led to a major shift in a paradigm of GI physiology regarding the role of interstitial cells in modulating GI contractile patterns. A review of these now classic experiments and papers will enable students and educators to fully appreciate the complex, multicellular nature of GI muscles as well as impart lessons on how shifting paradigms in physiology are fueled by new technologies that lead to new emerging discoveries. Copyright © 2017 the American Physiological Society.

Evidence for a Structural Motif in Toxins and Interleukin-2 That May Be Responsible for Binding to Endothelial Cells and Initiating Vascular Leak Syndrome

NASA Astrophysics Data System (ADS)

Baluna, Roxana; Rizo, Josep; Gordon, Brian E.; Ghetie, Victor; Vitetta, Ellen S.

1999-03-01

The dose-limiting toxicity of interleukin-2 (IL-2) and immunotoxin (IT) therapy in humans is vascular leak syndrome (VLS). VLS has a complex etiology involving damage to vascular endothelial cells (ECs), extravasation of fluids and proteins, interstitial edema, and organ failure. IL-2 and ITs prepared with the catalytic A chain of the plant toxin, ricin (RTA), and other toxins, damage human ECs in vitro and in vivo. Damage to ECs may initiate VLS; if this damage could be avoided without losing the efficacy of ITs or IL-2, larger doses could be administered. In this paper, we provide evidence that a three amino acid sequence motif, (x)D(y), in toxins and IL-2 damages ECs. Thus, when peptides from RTA or IL-2 containing this sequence motif are coupled to mouse IgG, they bind to and damage ECs both in vitro and, in the case of RTA, in vivo. In contrast, the same peptides with a deleted or mutated sequence do not. Furthermore, the peptide from RTA attached to mouse IgG can block the binding of intact RTA to ECs in vitro and vice versa. In addition, RTA, a fragment of Pseudomonas exotoxin A (PE38-lys), and fibronectin also block the binding of the mouse IgG-RTA peptide to ECs, suggesting that an (x)D(y) motif is exposed on all three molecules. Our results suggest that deletions or mutations in this sequence or the use of nondamaging blocking peptides may increase the therapeutic index of both IL-2, as well as ITs prepared with a variety of plant or bacterial toxins.

MRI-Based Computational Model of Heterogeneous Tracer Transport following Local Infusion into a Mouse Hind Limb Tumor

PubMed Central

Magdoom, Kulam Najmudeen; Pishko, Gregory L.; Rice, Lori; Pampo, Chris; Siemann, Dietmar W.; Sarntinoranont, Malisa

2014-01-01

Systemic drug delivery to solid tumors involving macromolecular therapeutic agents is challenging for many reasons. Amongst them is their chaotic microvasculature which often leads to inadequate and uneven uptake of the drug. Localized drug delivery can circumvent such obstacles and convection-enhanced delivery (CED) - controlled infusion of the drug directly into the tissue - has emerged as a promising delivery method for distributing macromolecules over larger tissue volumes. In this study, a three-dimensional MR image-based computational porous media transport model accounting for realistic anatomical geometry and tumor leakiness was developed for predicting the interstitial flow field and distribution of albumin tracer following CED into the hind-limb tumor (KHT sarcoma) in a mouse. Sensitivity of the model to changes in infusion flow rate, catheter placement and tissue hydraulic conductivity were investigated. The model predictions suggest that 1) tracer distribution is asymmetric due to heterogeneous porosity; 2) tracer distribution volume varies linearly with infusion volume within the whole leg, and exponentially within the tumor reaching a maximum steady-state value; 3) infusion at the center of the tumor with high flow rates leads to maximum tracer coverage in the tumor with minimal leakage outside; and 4) increasing the tissue hydraulic conductivity lowers the tumor interstitial fluid pressure and decreases the tracer distribution volume within the whole leg and tumor. The model thus predicts that the interstitial fluid flow and drug transport is sensitive to porosity and changes in extracellular space. This image-based model thus serves as a potential tool for exploring the effects of transport heterogeneity in tumors. PMID:24619021

Assessment of microcirculation dynamics during cutaneous wound healing phases in vivo using optical microangiography

PubMed Central

Yousefi, Siavash; Qin, Jia; Dziennis, Suzan; Wang, Ruikang K.

2014-01-01

Abstract. Cutaneous wound healing consists of multiple overlapping phases starting with blood coagulation following incision of blood vessels. We utilized label-free optical coherence tomography and optical microangiography (OMAG) to noninvasively monitor healing process and dynamics of microcirculation system in a mouse ear pinna wound model. Mouse ear pinna is composed of two layers of skin separated by a layer of cartilage and because its total thickness is around 500 μm, it can be utilized as an ideal model for optical imaging techniques. These skin layers are identical to human skin structure except for sweat ducts and glands. Microcirculatory system responds to the wound injury by recruiting collateral vessels to supply blood flow to hypoxic region. During the inflammatory phase, lymphatic vessels play an important role in the immune response of the tissue and clearing waste from interstitial fluid. In the final phase of wound healing, tissue maturation, and remodeling, the wound area is fully closed while blood vessels mature to support the tissue cells. We show that using OMAG technology allows noninvasive and label-free monitoring and imaging each phase of wound healing that can be used to replace invasive tissue sample histology and immunochemistry technologies. PMID:25036212

Mechanics of interstitial-lymphatic fluid transport: theoretical foundation and experimental validation.

PubMed

Swartz, M A; Kaipainen, A; Netti, P A; Brekken, C; Boucher, Y; Grodzinsky, A J; Jain, R K

1999-12-01

Interstitial fluid movement is intrinsically linked to lymphatic drainage. However, their relationship is poorly understood, and associated pathologies are mostly untreatable. In this work we test the hypothesis that bulk tissue fluid movement can be evaluated in situ and described by a linear biphasic theory which integrates the regulatory function of the lymphatics with the mechanical stresses of the tissue. To accomplish this, we develop a novel experimental and theoretical model using the skin of the mouse tail. We then use the model to demonstrate how interstitial-lymphatic fluid movement depends on a balance between the elasticity, hydraulic conductivity, and lymphatic conductance as well as to demonstrate how chronic swelling (edema) alters the equipoise between tissue fluid balance parameters. Specifically, tissue fluid equilibrium is perturbed with a continuous interstitial infusion of saline into the tip of the tail. The resulting gradients in tissue stress are measured in terms of interstitial fluid pressure using a servo-null system. These measurements are then fit to the theory to provide in vivo estimates of the tissue hydraulic conductivity, elastic modulus, and overall resistance to lymphatic drainage. Additional experiments are performed on edematous tails to show that although chronic swelling causes an increase in the hydraulic conductivity, its greatly increased distensibility (due to matrix remodeling) dampens the driving forces for fluid movement and leads to fluid stagnation. This model is useful for examining potential treatments for edema and lymphatic disorders as well as substances which may alter tissue fluid balance and/or lymphatic drainage.

Structure and composition of pulmonary arteries, capillaries and veins

PubMed Central

2013-01-01

The pulmonary vasculature is comprised of three anatomic compartments connected in series: the arterial tree, an extensive capillary bed, and the venular tree. Although in general this vasculature is thin-walled, structure is nonetheless complex. Contributions to structure (and thus potentially to function) from cells other than endothelial and smooth muscle cells as well as those from the extracellular matrix should be considered. This review is multifaceted, bringing together information regarding 1) classification of pulmonary vessels, 2) branching geometry in the pulmonary vascular tree, 3) a quantitative view of structure based on morphometry of the vascular wall, 4) the relationship of nerves, a variety of interstitial cells, matrix proteins, and striated myocytes to smooth muscle and endothelium in the vascular wall, 5) heterogeneity within cell populations and between vascular compartments, 6) homo- and heterotypic cell-cell junctional complexes, and 7) the relation of the pulmonary vasculature to that of airways. These issues for pulmonary vascular structure are compared, when data is available, across species from human to mouse and shrew. Data from studies utilizing vascular casting, light and electron microscopy, as well as models developed from those data, are discussed. Finally, the need for rigorous quantitative approaches to study of vascular structure in lung is highlighted. PMID:23606929

Cellular Plasticity in the Diabetic Myocardium

DTIC Science & Technology

2017-09-01

pathogenesis of cardiac dysfunction associated with metabolic disease . 8 Fig. 1: The db/db mouse recapitulates features of human Heart failure with ...patients, hypertensive heart disease is associated with development of interstitial and periarteriolar fibrosis even in the absence of significant cor...increased secretion of structural matrix proteins. The cardiac ECM in metabolic disease Diabetics exhibit a high incidence of heart failure with

A dural lymphatic vascular system that drains brain interstitial fluid and macromolecules

PubMed Central

Aspelund, Aleksanteri; Antila, Salli; Proulx, Steven T.; Karlsen, Tine Veronica; Karaman, Sinem; Detmar, Michael; Wiig, Helge

2015-01-01

The central nervous system (CNS) is considered an organ devoid of lymphatic vasculature. Yet, part of the cerebrospinal fluid (CSF) drains into the cervical lymph nodes (LNs). The mechanism of CSF entry into the LNs has been unclear. Here we report the surprising finding of a lymphatic vessel network in the dura mater of the mouse brain. We show that dural lymphatic vessels absorb CSF from the adjacent subarachnoid space and brain interstitial fluid (ISF) via the glymphatic system. Dural lymphatic vessels transport fluid into deep cervical LNs (dcLNs) via foramina at the base of the skull. In a transgenic mouse model expressing a VEGF-C/D trap and displaying complete aplasia of the dural lymphatic vessels, macromolecule clearance from the brain was attenuated and transport from the subarachnoid space into dcLNs was abrogated. Surprisingly, brain ISF pressure and water content were unaffected. Overall, these findings indicate that the mechanism of CSF flow into the dcLNs is directly via an adjacent dural lymphatic network, which may be important for the clearance of macromolecules from the brain. Importantly, these results call for a reexamination of the role of the lymphatic system in CNS physiology and disease. PMID:26077718

A dural lymphatic vascular system that drains brain interstitial fluid and macromolecules.

PubMed

Aspelund, Aleksanteri; Antila, Salli; Proulx, Steven T; Karlsen, Tine Veronica; Karaman, Sinem; Detmar, Michael; Wiig, Helge; Alitalo, Kari

2015-06-29

The central nervous system (CNS) is considered an organ devoid of lymphatic vasculature. Yet, part of the cerebrospinal fluid (CSF) drains into the cervical lymph nodes (LNs). The mechanism of CSF entry into the LNs has been unclear. Here we report the surprising finding of a lymphatic vessel network in the dura mater of the mouse brain. We show that dural lymphatic vessels absorb CSF from the adjacent subarachnoid space and brain interstitial fluid (ISF) via the glymphatic system. Dural lymphatic vessels transport fluid into deep cervical LNs (dcLNs) via foramina at the base of the skull. In a transgenic mouse model expressing a VEGF-C/D trap and displaying complete aplasia of the dural lymphatic vessels, macromolecule clearance from the brain was attenuated and transport from the subarachnoid space into dcLNs was abrogated. Surprisingly, brain ISF pressure and water content were unaffected. Overall, these findings indicate that the mechanism of CSF flow into the dcLNs is directly via an adjacent dural lymphatic network, which may be important for the clearance of macromolecules from the brain. Importantly, these results call for a reexamination of the role of the lymphatic system in CNS physiology and disease. © 2015 Aspelund et al.

Overexpression of SerpinE2/protease nexin-1 Contribute to Pathological Cardiac Fibrosis via increasing Collagen Deposition

PubMed Central

Li, Xuelian; Zhao, Dandan; Guo, Zhenfeng; Li, Tianshi; Qili, Muge; Xu, Bozhi; Qian, Ming; Liang, Haihai; E, Xiaoqiang; Chege Gitau, Samuel; Wang, Lu; Huangfu, Longtao; Wu, Qiuxia; Xu, Chaoqian; Shan, Hongli

2016-01-01

Although increases in cardiovascular load (pressure overload) are known to elicit ventricular remodeling including cardiomyocyte hypertrophy and interstitial fibrosis, the molecular mechanisms of pressure overload or AngII -induced cardiac interstitial fibrosis remain elusive. In this study, serpinE2/protease nexin-1 was over-expressed in a cardiac fibrosis model induced by pressure-overloaded via transverse aortic constriction (TAC) in mouse. Knockdown of serpinE2 attenuates cardiac fibrosis in a mouse model of TAC. At meantime, the results showed that serpinE2 significantly were increased with collagen accumulations induced by AngII or TGF-β stimulation in vitro. Intriguingly, extracellular collagen in myocardial fibroblast was reduced by knockdown of serpinE2 compared with the control in vitro. In stark contrast, the addition of exogenous PN-1 up-regulated the content of collagen in myocardial fibroblast. The MEK1/2- ERK1/2 signaling probably promoted the expression of serpinE2 via transcription factors Elk1 in myocardial fibroblast. In conclusion, stress-induced the ERK1/2 signaling pathway activation up-regulated serpinE2 expression, consequently led accumulation of collagen protein, and contributed to cardiac fibrosis. PMID:27876880

T Cell Interstitial Migration: Motility Cues from the Inflamed Tissue for Micro- and Macro-Positioning.

PubMed

Gaylo, Alison; Schrock, Dillon C; Fernandes, Ninoshka R J; Fowell, Deborah J

2016-01-01

Effector T cells exit the inflamed vasculature into an environment shaped by tissue-specific structural configurations and inflammation-imposed extrinsic modifications. Once within interstitial spaces of non-lymphoid tissues, T cells migrate in an apparent random, non-directional, fashion. Efficient T cell scanning of the tissue environment is essential for successful location of infected target cells or encounter with antigen-presenting cells that activate the T cell's antimicrobial effector functions. The mechanisms of interstitial T cell motility and the environmental cues that may promote or hinder efficient tissue scanning are poorly understood. The extracellular matrix (ECM) appears to play an important scaffolding role in guidance of T cell migration and likely provides a platform for the display of chemotactic factors that may help to direct the positioning of T cells. Here, we discuss how intravital imaging has provided insight into the motility patterns and cellular machinery that facilitates T cell interstitial migration and the critical environmental factors that may optimize the efficiency of effector T cell scanning of the inflamed tissue. Specifically, we highlight the local micro-positioning cues T cells encounter as they migrate within inflamed tissues, from surrounding ECM and signaling molecules, as well as a requirement for appropriate long-range macro-positioning within distinct tissue compartments or at discrete foci of infection or tissue damage. The central nervous system (CNS) responds to injury and infection by extensively remodeling the ECM and with the de novo generation of a fibroblastic reticular network that likely influences T cell motility. We examine how inflammation-induced changes to the CNS landscape may regulate T cell tissue exploration and modulate function.

Resveratrol Reduces Steroidogenesis in Rat Ovarian Theca-Interstitial Cells: The Role of Inhibition of Akt/PKB Signaling Pathway

PubMed Central

Ortega, Israel; Villanueva, Jesus A.; Wong, Donna H.; Cress, Amanda B.; Sokalska, Anna; Stanley, Scott D.

2012-01-01

Polycystic ovary syndrome is characterized by theca-interstitial hyperplasia and increased expression of steroidogenic genes, leading to excessive androgen production. Resveratrol, a natural polyphenol, promotes apoptosis and reduces rat theca-interstitial cell growth, in part by inhibiting the mevalonate pathway and decreasing the availability of substrates of isoprenylation [farnesyl-pyrophosphate (FPP) and geranylgeranyl-pyrophosphate (GGPP)]. This study evaluated the effect of resveratrol on rat theca-interstitial cell steroidogenesis. Because resveratrol may activate sirtuins, this study also investigated whether steroidogenesis was affected by sirtuin inhibitors (nicotinamide, sirtinol). Theca-interstitial cells were cultured with or without resveratrol (1–10 μm), GGPP (30 μm), FPP (30 μm), nicotinamide (1 mm), and/or sirtinol (10 μm). Resveratrol did not affect progesterone levels but reduced androgen production in a concentration-dependent fashion (androstenedione by up to 78% and androsterone by up to 76%). This inhibitory effect correlated with a decrease in mRNA expression of genes regulating androgen production, especially Cyp17a1 (by up to 73%). GGPP and FPP had no effect on androgen levels and Cyp17a1 mRNA levels and did not alter the effects induced by resveratrol. Similarly, sirtuin inhibitors did not reverse resveratrol-induced inhibition of steroidogenesis. However, resveratrol decreased activity of serine-threonine kinase/protein kinase B pathway, a cell-signaling pathway involved in ovarian steroidogenesis. The present findings indicate that resveratrol reduces androgen production primarily by inhibiting Cyp17a1 mRNA expression, and this inhibition may be mediated, in part, by blocking the activity of the serine-threonine kinase/protein kinase B pathway. These findings may be of clinical relevance to conditions associated with excessive production of androgens by theca cells, such as polycystic ovary syndrome. PMID:22719052

Spatial organization and coordination of slow waves in the mouse anorectum

PubMed Central

Hall, K A; Ward, S M; Cobine, C A; Keef, K D

2014-01-01

The internal anal sphincter (IAS) develops tone and is important for maintaining a high anal pressure while tone in the rectum is less. The mechanisms responsible for tone generation in the IAS are still uncertain. The present study addressed this question by comparing the electrical properties and morphology of the mouse IAS and distal rectum. The amplitude of tone and the frequency of phasic contractions was greater in the IAS than in rectum while membrane potential (Em) was less negative in the IAS than in rectum. Slow waves (SWs) were of greatest amplitude and frequency at the distal end of the IAS, declining in the oral direction. Dual microelectrode recordings revealed that SWs were coordinated over a much greater distance in the circumferential direction than in the oral direction. The circular muscle layer of the IAS was divided into five to eight ‘minibundles’ separated by connective tissue septa whereas few septa were present in the rectum. The limited coordination of SWs in the oral direction suggests that the activity in adjacent minibundles is not coordinated. Intramuscular interstitial cells of Cajal and platelet-derived growth factor receptor alpha-positive cells were present in each minibundle suggesting a role for one or both of these cells in SW generation. In summary, three important properties distinguish the IAS from the distal rectum: (1) a more depolarized Em; (2) larger and higher frequency SWs; and (3) the multiunit configuration of the muscle. All of these characteristics may contribute to greater tone generation in the IAS than in the distal rectum. PMID:24951622

TGF-β mediates early angiogenesis and latent fibrosis in an Emilin1-deficient mouse model of aortic valve disease

PubMed Central

Munjal, Charu; Opoka, Amy M.; Osinska, Hanna; James, Jeanne F.; Bressan, Giorgio M.; Hinton, Robert B.

2014-01-01

Aortic valve disease (AVD) is characterized by elastic fiber fragmentation (EFF), fibrosis and aberrant angiogenesis. Emilin1 is an elastin-binding glycoprotein that regulates elastogenesis and inhibits TGF-β signaling, but the role of Emilin1 in valve tissue is unknown. We tested the hypothesis that Emilin1 deficiency results in AVD, mediated by non-canonical (MAPK/phosphorylated Erk1 and Erk2) TGF-β dysregulation. Using histology, immunohistochemistry, electron microscopy, quantitative gene expression analysis, immunoblotting and echocardiography, we examined the effects of Emilin1 deficiency (Emilin1−/−) in mouse aortic valve tissue. Emilin1 deficiency results in early postnatal cell-matrix defects in aortic valve tissue, including EFF, that progress to latent AVD and premature death. The Emilin1−/− aortic valve displays early aberrant provisional angiogenesis and late neovascularization. In addition, Emilin1−/− aortic valves are characterized by early valve interstitial cell activation and proliferation and late myofibroblast-like cell activation and fibrosis. Interestingly, canonical TGF-β signaling (phosphorylated Smad2 and Smad3) is upregulated constitutively from birth to senescence, whereas non-canonical TGF-β signaling (phosphorylated Erk1 and Erk2) progressively increases over time. Emilin1 deficiency recapitulates human fibrotic AVD, and advanced disease is mediated by non-canonical (MAPK/phosphorylated Erk1 and Erk2) TGF-β activation. The early manifestation of EFF and aberrant angiogenesis suggests that these processes are crucial intermediate factors involved in disease progression and therefore might provide new therapeutic targets for human AVD. PMID:25056700

T Cell Interstitial Migration: Motility Cues from the Inflamed Tissue for Micro- and Macro-Positioning

PubMed Central

Gaylo, Alison; Schrock, Dillon C.; Fernandes, Ninoshka R. J.; Fowell, Deborah J.

2016-01-01

Effector T cells exit the inflamed vasculature into an environment shaped by tissue-specific structural configurations and inflammation-imposed extrinsic modifications. Once within interstitial spaces of non-lymphoid tissues, T cells migrate in an apparent random, non-directional, fashion. Efficient T cell scanning of the tissue environment is essential for successful location of infected target cells or encounter with antigen-presenting cells that activate the T cell’s antimicrobial effector functions. The mechanisms of interstitial T cell motility and the environmental cues that may promote or hinder efficient tissue scanning are poorly understood. The extracellular matrix (ECM) appears to play an important scaffolding role in guidance of T cell migration and likely provides a platform for the display of chemotactic factors that may help to direct the positioning of T cells. Here, we discuss how intravital imaging has provided insight into the motility patterns and cellular machinery that facilitates T cell interstitial migration and the critical environmental factors that may optimize the efficiency of effector T cell scanning of the inflamed tissue. Specifically, we highlight the local micro-positioning cues T cells encounter as they migrate within inflamed tissues, from surrounding ECM and signaling molecules, as well as a requirement for appropriate long-range macro-positioning within distinct tissue compartments or at discrete foci of infection or tissue damage. The central nervous system (CNS) responds to injury and infection by extensively remodeling the ECM and with the de novo generation of a fibroblastic reticular network that likely influences T cell motility. We examine how inflammation-induced changes to the CNS landscape may regulate T cell tissue exploration and modulate function. PMID:27790220

Ulcerative colitis: ultrastructure of interstitial cells in myenteric plexus.

PubMed

Rumessen, J J; Vanderwinden, J-M; Horn, T

2010-10-01

Interstitial cells of Cajal (ICC) are key regulatory cells in the gut. In the colon of patients with severe ulcerative colitis (UC), myenteric ICC had myoid ultrastructural features and were in close contact with nerve terminals. In all patients as opposed to controls, some ICC profiles showed degenerative changes, such as lipid droplets and irregular vacuoles. Nerve terminals often appeared swollen and empty. Glial cells, muscle cells, and fibroblast-like cells (FLC) showed no alterations. FLC enclosed macrophages (MLC), which were in close contact with naked axon terminals. The organization and cytological changes may be of pathophysiological significance in patients with UC.

Loss of CD11b Exacerbates Murine Complement-Mediated Tubulointerstitial Nephritis

PubMed Central

Wang, Ying; Chang, Anthony; Haas, Mark; Quigg, Richard John

2014-01-01

Acute complement activation occurs in the tubulointerstitium (TI) of kidneys transplanted from Crry−/−C3−/− mice into complement-sufficient wildtype mice, followed by marked inflammatory cell infiltration, tubular damage and interstitial fibrosis. We postulated iC3b-CD11b interactions were critical in this TI nephritis model. We transplanted Crry−/−C3−/− mouse kidneys into CD11b−/− and wildtype C57BL/6 mice. Surprisingly, there was greater inflammation in Crry−/−C3−/− kidneys in CD11b−/− recipients compared to those in wildtype hosts. Kidneys in CD11b−/− recipients had large numbers of CD11b−Ly6ChiCCR2hiF4/80+ cells consistent with inflammatory (M1) macrophages recruited from circulating monocytes of the host CD11b−/− animal. There was also an expanded population of CD11b+CD11c+Ly6C−F4/80hi cells. Since these cells were CD11b+, they must have originated from the transplanted kidney; their surface protein expression and appearance within the kidney were consistent with the intrinsic renal mononuclear cellular population. These cells were markedly expanded relative to all relevant controls, including the contralateral donor kidney and Crry−/−C3−/− mouse kidneys in CD11b+/+ wildtype recipients. Direct evidence for their in situ proliferation was the presence of nuclear Ki67 and PCNA in CD11b+F4/80+ cells. Thus, in this experimental model in which there is unrestricted C3 activation, CD11b+ monocytes limit their own infiltration into the kidney and prevent proliferation of endogenous mononuclear cells. This suggests a role for outside-in iC3b-CD11b signals in limiting intrinsic organ inflammation. PMID:24632830

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tabatowski, K.; Roggli, V.L.; Fulkerson, W.J.

A case of biopsy-proven giant cell interstitial pneumonia in a patient with occupational exposure to hard-metal dust is reported. Bronchial washings performed several days prior to open-lung biopsy yielded an almost exclusive population of nonpigmented alveolar macrophages and pleomorphic, phagocytic multinucleated giant cells. Microorganisms, viral inclusions in the giant cells, epithelioid histiocytes and well-formed granulomas were not seen. This cytologic picture strongly suggests the presence of giant cell interstitial pneumonia in a patient with restrictive lung disease, particularly when exposure to hard-metal dust is known or suspected. A specific diagnosis early in the course of the disease may facilitate removalmore » of the individual from the workplace and forestall the development of end-stage interstitial fibrosis. Additionally, the working environment may be modified to minimize inhalational exposure. Recognition of this entity by the cytopathologist may direct diagnostic efforts toward accurate histologic evaluation and the identification of particulates by microprobe analysis of either cellular or biopsy material.« less

Microfluidic enhancement of intramedullary pressure increases interstitial fluid flow and inhibits bone loss in hindlimb suspended mice.

PubMed

Kwon, Ronald Y; Meays, Diana R; Tang, W Joyce; Frangos, John A

2010-08-01

Interstitial fluid flow (IFF) has been widely hypothesized to mediate skeletal adaptation to mechanical loading. Although a large body of in vitro evidence has demonstrated that fluid flow stimulates osteogenic and antiresorptive responses in bone cells, there is much less in vivo evidence that IFF mediates loading-induced skeletal adaptation. This is due in large part to the challenges associated with decoupling IFF from matrix strain. In this study we describe a novel microfluidic system for generating dynamic intramedullary pressure (ImP) and IFF within the femurs of alert mice. By quantifying fluorescence recovery after photobleaching (FRAP) within individual lacunae, we show that microfluidic generation of dynamic ImP significantly increases IFF within the lacunocanalicular system. In addition, we demonstrate that dynamic pressure loading of the intramedullary compartment for 3 minutes per day significantly eliminates losses in trabecular and cortical bone mineral density in hindlimb suspended mice, enhances trabecular and cortical structural integrity, and increases endosteal bone formation rate. Unlike previously developed modalities for enhancing IFF in vivo, this is the first model that allows direct and dynamic modulation of ImP and skeletal IFF within mice. Given the large number of genetic tools for manipulating the mouse genome, this model is expected to serve as a powerful investigative tool in elucidating the role of IFF in skeletal adaptation to mechanical loading and molecular mechanisms mediating this process.

Electrode assembly for use in a solid polymer electrolyte fuel cell

DOEpatents

Raistrick, Ian D.

1989-01-01

A gas reaction fuel cell may be provided with a solid polymer electrolyte membrane. Porous gas diffusion electrodes are formed of carbon particles supporting a catalyst which is effective to enhance the gas reactions. The carbon particles define interstitial spaces exposing the catalyst on a large surface area of the carbon particles. A proton conducting material, such as a perfluorocarbon copolymer or ruthenium dioxide contacts the surface areas of the carbon particles adjacent the interstitial spaces. The proton conducting material enables protons produced by the gas reactions adjacent the supported catalyst to have a conductive path with the electrolyte membrane. The carbon particles provide a conductive path for electrons. A suitable electrode may be formed by dispersing a solution containing a proton conducting material over the surface of the electrode in a manner effective to coat carbon surfaces adjacent the interstitial spaces without impeding gas flow into the interstitial spaces.

Acute inhalation toxicity of 3-methylfuran in the mouse: pathology, cell kinetics, and respiratory rate effects

DOE Office of Scientific and Technical Information (OSTI.GOV)

Haschek, W.M.; Boyd, M.R.; Hakkinen, P.J.

1984-01-01

The acute inhalation toxicity of 3-methylfuran (3MF) was investigated in male BALB/c mice by morphologic examination of animals killed at varying timepoints following a 1-hr exposure to an initial chamber concentration of 14 to 37 mumol/liter (343 to 906 ppm). In addition, respiratory rate measurements and cell kinetics were used to assess quantitatively pulmonary damage and repair. Necrosis of nonciliated bronchiolar epithelial (Clara) cells was seen 1 day following exposure and was followed by regeneration, which was virtually complete, within 21 days. Cell kinetic studies showed peak bronchiolar cell proliferation at 3 days with a labeling index (LI) of 5.0%more » compared to 0.4% in controls. An increase in parenchymal cell proliferation was also noted coincident with a mild interstitial pneumonitis. This parenchymal proliferation, peaking at 10 days with an LI of 1.4% compared to 0.2% in controls, consisted primarily of type II epithelial and endothelial cell proliferation indicating possible delayed damage and repair of type I epithelial and endothelial cells. The respiratory rate showed an initial transient increase followed by a more prolonged decrease with eventual return to control levels. 3MF toxicity was also evidenced by a necrotizing suppurative rhinitis, centrilobular hepatic necrosis, lymphocyte necrosis in the thymus and spleen, sialoadenitis, and otitis media.« less

New insights into canted spiro carbon interstitial in graphite

NASA Astrophysics Data System (ADS)

EL-Barbary, A. A.

2017-12-01

The self-interstitial carbon is the key to radiation damage in graphite moderator nuclear reactor, so an understanding of its behavior is essential for plant safety and maximized reactor lifetime. The density functional theory is applied on four different graphite unit cells, starting from of 64 carbon atoms up to 256 carbon atoms, using AIMPRO code to obtain the energetic, athermal and mechanical properties of carbon interstitial in graphite. This study presents first principles calculations of the energy of formation that prove its high barrier to athermal diffusion (1.1 eV) and the consequent large critical shear stress (39 eV-50 eV) necessary to shear graphite planes in its presence. Also, for the first time, the gamma surface of graphite in two dimensions is calculated and found to yield the critical shear stress for perfect graphite. Finally, in contrast to the extensive literature describing the interstitial of carbon in graphite as spiro interstitial, in this work the ground state of interstitial carbon is found to be canted spiro interstitial.

Differential Mouse Pulmonary Dose and Time Course Responses to Titanium Dioxide Nanospheres and Nanobelts

PubMed Central

Porter, Dale W.

2013-01-01

Three anatase titanium dioxide (TiO2) nanoparticles (NPs) were prepared; nanospheres (NSs), short nanobelts (NB1), and long nanobelts (NB2). These NPs were used to investigate the effect of NP shape and length on lung toxicity. Mice were exposed (0–30 µg per mouse) by pharyngeal aspiration and pulmonary toxicity was assessed over a 112-day time course. Whole lung lavage data indicated that NB1- and NB2-exposed mice, but not NS-exposed mice, had significant dose- and time-dependent pulmonary inflammation and damage. Histopathological analyses at 112 days postexposure determined no interstitial fibrosis in any NS-exposed mice, an increased incidence in 30 µg NB1-exposed mice, and significant interstitial fibrosis in 30 µg NB2-exposed mice. At 112 days postexposure, lung burden of NS was decreased by 96.4% and NB2 by 80.5% from initial deposition levels. At 112 days postexposure, enhanced dark field microscopy determined that alveolar macro- phages were the dominant deposition site, but a fraction of NB1 and NB2 was observed in the alveolar interstitial spaces. For the 30 µg exposure groups at 112 days postexposure, confocal micro- scopy and immunofluorescent staining demonstrated that retained NB2 but not NS were present in the interstitium subjacent to the terminal bronchiole near the normal location of the smallest lymphatic capillaries in the lung. These lymphatic capillaries play a critical role in particle clearance, and the accumulation of NB2, but not NS, suggests possible impaired lymphatic clearance by the high aspect ratio particles. In summary, our data indicate that TiO2 NP shape alters pulmonary responses, with severity of responses being ranked as NS < NB1 < NB2. PMID:22956629

Smad2-dependent glycosaminoglycan elongation in aortic valve interstitial cells enhances binding of LDL to proteoglycans.

PubMed

Osman, Narin; Grande-Allen, K Jane; Ballinger, Mandy L; Getachew, Robel; Marasco, Silvana; O'Brien, Kevin D; Little, Peter J

2013-01-01

Calcific aortic valve disease is a progressive condition that shares some common pathogenic features with atherosclerosis. Transforming growth factor-β1 is a recognized mediator of atherosclerosis and is expressed in aortic valve lesions. Transforming growth factorβ1 stimulates glycosaminoglycan elongation of proteoglycans that is associated with increased lipid binding. We investigated the presence of transforming growth factor-β1 and downstream signaling intermediates in diseased human aortic valves and the effects of activated transforming growth factor-β1 receptor signaling on aortic valve interstitial cell proteoglycan synthesis and lipid binding as a possible mechanism for the initiation of the early lesion of calcific aortic valve disease. Diseased human aortic valve leaflets demonstrated strong immunohistochemical staining for transforming growth factor-β1 and phosphorylated Smad2/3. In primary porcine aortic valve interstitial cells, Western blots showed that transforming growth factor-β1 stimulated phosphorylation in both the carboxy and linker regions of Smad2/3, which was inhibited by the transforming growth factor-β1 receptor inhibitor SB431542. Gel electrophoresis and size exclusion chromatography demonstrated that SB431542 decreased transforming growth factor-β1-mediated [(35)S]-sulfate incorporation into proteoglycans in a dose-dependent manner. Further, in proteoglycans derived from transforming growth factor-β1-treated valve interstitial cells, gel mobility shift assays demonstrated that inhibition of transforming growth factor-β1 receptor signaling resulted in decreased lipid binding. Classic transforming growth factor-β1 signaling is present in human aortic valves in vivo and contributes to the modification of proteoglycans expressed by valve interstitial cells in vitro. These findings suggest that transforming growth factor-β1 may promote increased low-density lipoprotein binding in the early phases of calcific aortic valve disease. Copyright © 2013 Elsevier Inc. All rights reserved.

Activation of Protein Kinase C by Mycobacterial Cord Factor, Trehalose 6‐Monomycolate, Resulting in Tumor Necrosis Factor‐α Release in Mouse Lung Tissues

PubMed Central

Sueoka, Eisaburo; Nishiwaki, Shinji; Okabe, Sachiko; Iida, Naoyuki; Suganuma, Masami; Yano, Ikuya; Aoki, Kunio

1995-01-01

Cord factors are mycoloyl glycolipids in cell walls of bacteria belonging to Actinomycetales, such as Mycobacterium, Nocardia and Rhodococcus. They induce granuloma formation in the lung and interstitial pneumonitis, associated with production of macrophage‐derived cytokines. We studied how cord factors induce biological activities in the cells. Cord factors isolated from M. tuberculosis, trehalose 6‐monomycolate (mTMM) and trehalose 6,6′‐dimycolate (mTDM), enhanced protein kinase C (PKC) activation in the presence of phosphatidylserine (PtdSer), diacylglycerol and Ca2+, and mTMM activated PKCα more strongly than PKCβ or γ under the same assay conditions. Kinetic studies of mTMM in response to PKC activation revealed that mTMM increased the apparent affinity of PKC to Ca2+ in the presence of both PtdSer and diolein. Although this is similar to observations with unsaturated fatty acids, such as arachidonic acid, mTMM was synergistic with PtdSer for PKC activation, but arachidonic acid was not. mTMM was also different as regards PKC activation, as phorbol ester was. A single i.p. administration of mTMM to mouse induced tumor necrosis factor‐α (TNF‐α) in serum and in the lung, which is a unique target tissue of cord factors. Based on our recent finding that TNF‐α is an endogenous tumor promoter, the correlation between lung cancer and pulmonary tuberculosis is discussed. PMID:7559098

Activation of protein kinase C by mycobacterial cord factor, trehalose 6-monomycolate, resulting in tumor necrosis factor-alpha release in mouse lung tissues.

PubMed

Sueoka, E; Nishiwaki, S; Okabe, S; Iida, N; Suganuma, M; Yano, I; Aoki, K; Fujiki, H

1995-08-01

Cord factors are mycoloyl glycolipids in cell walls of bacteria belonging to Actinomycetales, such as Mycobacterium, Nocardia and Rhodococcus. They induce granuloma formation in the lung and interstitial pneumonitis, associated with production of macrophage-derived cytokines. We studied how cord factors induce biological activities in the cells. Cord factors isolated from M. tuberculosis, trehalose 6-monomycolate (mTMM) and trehalose 6,6'-dimycolate (mTDM), enhanced protein kinase C (PKC) activation in the presence of phosphatidylserine (PtdSer), diacylglycerol and Ca2+, and mTMM activated PKC alpha more strongly than PKC beta or gamma under the same assay conditions. Kinetic studies of mTMM in response to PKC activation revealed that mTMM increased the apparent affinity of PKC to Ca2+ in the presence of both PtdSer and diolein. Although this is similar to observations with unsaturated fatty acids, such as arachidonic acid, mTMM was synergistic with PtdSer for PKC activation, but arachidonic acid was not. mTMM was also different as regards PKC activation, as phorbol ester was. A single i.p. administration of mTMM to mouse induced tumor necrosis factor-alpha (TNF-alpha) in serum and in the lung, which is a unique target tissue of cord factors. Based on our recent finding that TNF-alpha is an endogenous tumor promoter, the correlation between lung cancer and pulmonary tuberculosis is discussed.

Progenitor-like cells derived from mouse kidney protect against renal fibrosis in a remnant kidney model via decreased endothelial mesenchymal transition.

PubMed

Chen, C L; Chou, K J; Fang, H C; Hsu, C Y; Huang, W C; Huang, C W; Huang, C K; Chen, H Y; Lee, P T

2015-12-02

Pathophysiological changes associated with chronic kidney disease impair angiogenic processes and increase renal fibrosis. Progenitor-like cells derived from adult kidney have been previously used to promote regeneration in acute kidney injury, even though it remained unclear whether the cells could be beneficial in chronic kidney disease (CKD). In this study, we established a CKD model by five-sixths nephrectomy and mouse kidney progenitor-like cells (MKPCs) were intravenously administered weekly for 5 weeks after establishing CKD. We examined the impact of MKPCs on the progression of renal fibrosis and the potential of MKPCs to preserve the angiogenic process and prevent endothelial mesenchymal transition in vivo and in vitro. Our results demonstrate that the MKPCs delayed interstitial fibrosis and the progression of glomerular sclerosis and ameliorated the decline of kidney function. At 17 weeks, the treated mice exhibited lower blood pressures, higher hematocrit levels, and larger kidney sizes than the control mice. In addition, the MKPC treatment prolonged the survival of the mice with chronic kidney injuries. We observed a decreased recruitment of macrophages and myofibroblasts in the interstitium and the increased tubular proliferation. Notably, MKPC both decreased the level of vascular rarefaction and prevented endothelial mesenchymal transition (EndoMT) in the remnant kidneys. Moreover, the conditioned medium from the MKPCs ameliorated endothelial cell death under hypoxic culture conditions and prevented TGF-β-induced EndoMT through downregulation of phosphorylated Smad 3 in vitro. MKPCs may be a beneficial treatment for kidney diseases characterized by progressive renal fibrosis. The enhanced preservation of angiogenic processes following MKPC injections may be associated with decreased fibrosis in the remnant kidney. These findings provide further understanding of the mechanisms involved in these processes and will help develop new cell-based therapeutic strategies for regenerative medicine in renal fibrosis.

Regional differences in the morphological and functional effects of aging on cerebral basement membranes and perivascular drainage of amyloid-β from the mouse brain.

PubMed

Hawkes, Cheryl A; Gatherer, Maureen; Sharp, Matthew M; Dorr, Adrienne; Yuen, Ho Ming; Kalaria, Rajesh; Weller, Roy O; Carare, Roxana O

2013-04-01

Development of cerebral amyloid angiopathy (CAA) and Alzheimer's disease (AD) is associated with failure of elimination of amyloid-β (Aβ) from the brain along perivascular basement membranes that form the pathways for drainage of interstitial fluid and solutes from the brain. In transgenic APP mouse models of AD, the severity of cerebral amyloid angiopathy is greater in the cerebral cortex and hippocampus, intermediate in the thalamus, and least in the striatum. In this study we test the hypothesis that age-related regional variation in (1) vascular basement membranes and (2) perivascular drainage of Aβ contribute to the different regional patterns of CAA in the mouse brain. Quantitative electron microscopy of the brains of 2-, 7-, and 23-month-old mice revealed significant age-related thickening of capillary basement membranes in cerebral cortex, hippocampus, and thalamus, but not in the striatum. Results from Western blotting and immunocytochemistry experiments showed a significant reduction in collagen IV in the cortex and hippocampus with age and a reduction in laminin and nidogen 2 in the cortex and striatum. Injection of soluble Aβ into the hippocampus or thalamus showed an age-related reduction in perivascular drainage from the hippocampus but not from the thalamus. The results of the study suggest that changes in vascular basement membranes and perivascular drainage with age differ between brain regions, in the mouse, in a manner that may help to explain the differential deposition of Aβ in the brain in AD and may facilitate development of improved therapeutic strategies to remove Aβ from the brain in AD. © 2013 The Authors Aging Cell © 2013 Blackwell Publishing Ltd/Anatomical Society of Great Britain and Ireland.

Effectiveness of Combined Therapy with Pirfenidone and Erythromycin for Unclassifiable Interstitial Pneumonia Induced by HTLV-1-associated Bronchioloalveolar Disorder (HABA)

PubMed Central

Yokohori, Naoko; Sato, Akitoshi; Hasegawa, Mizue; Katsura, Hideki; Hiroshima, Kenzo; Takemura, Tamiko

2017-01-01

Human T-cell lymphotropic virus type 1 (HTLV-1) is a retrovirus involved in the pathogenesis of adult T-cell leukemia (ATL) and HTVL-1-associated bronchioloalveolar disorder (HABA). The clinical and pathological findings of HABA have been characterized as either a diffuse panbronchiolitis (DPB) pattern or idiopathic interstitial pneumonia (IIP) pattern. Treatments for HABA include corticosteroids for the IIP pattern and erythromycin for the DPB pattern. We herein report a case of HABA-associated unclassifiable interstitial pneumonia that improved with combined therapy with pirfenidone and erythromycin. This is the first report on the effectiveness of combined therapy with pirfenidone and erythromycin for HABA. PMID:28050003

Giant-cell interstitial pneumonia in a gas station worker.

PubMed

Lee, S M; Moon, C H; Oh, Y B; Kim, H Y; Ahn, Y; Ko, E J; Joo, J E

1998-10-01

Giant-cell interstitial Pneumonia (GIP) is a very uncommon respiratory disease. The majority of cases of GIP are caused by exposure to cobalt, tungsten and other hard metals. In this report, we describe GIP in a patient who worked in gas station and dealt in propane gas vessels. He presented with clinical features of chronic interstitial lung disease and underwent an open lung biopsy that showed DIP-like reaction with large numbers of intra-alveolar macrophages and numerous large, multinucleated histiocytes which were admixed with the macrophages. Analysis of lung tissue for hard metals was done. Cobalt was the main component of detected hard metals. Corticosteroid therapy was started and he recovered fully.

Matrix remodeling maintains ESC self-renewal by activating Stat3

PubMed Central

Przybyla, Laralynne M.; Theunissen, Thorold W.; Jaenisch, Rudolf; Voldman, Joel

2013-01-01

While a variety of natural and synthetic matrices have been used to influence embryonic stem cell (ESC) self-renewal or differentiation, and ESCs also deposit a rich matrix of their own, the mechanisms behind how extracellular matrix affects cell fate are largely unexplored. The ESC matrix is continuously remodeled by matrix metalloproteinases (MMPs), a process that we find is enhanced by the presence of mouse embryonic fibroblast feeders in a paracrine manner. Matrix remodeling by MMPs aids in the self-renewal of ESCs, as inhibition of MMPs inhibits the ability of ESCs to self-renew. We also find that addition of the interstitial collagenase MMP1 is sufficient to maintain long-term LIF-independent mESC self-renewal in a dose-dependent manner. This remarkable ability is due to the presence of endogenously produced self-renewal-inducing signals, including the LIF-family ligand CNTF, that are normally trapped within the ECM and become exposed upon MMP-induced matrix remodeling to signal through JAK and Stat3. These results uncover a new role for feeder cells in maintaining self-renewal and show that mESCs normally produce sufficient levels of autocrine-acting pro-self-renewal ligands. PMID:23404867

Organizer formation in Hydra is disrupted by thalidomide treatment.

PubMed

Brooun, Maria; Manoukian, Armen; Shimizu, Hiroshi; Bode, Hans R; McNeill, Helen

2013-06-01

Thalidomide is a drug that is well known for its teratogenic properties in humans. Surprisingly, thalidomide does not have teratogenic effects on mouse development. We investigated the effect of thalidomide on patterning in hydra, an early metazoan with a very simple axial symmetry. Hydra develops asexually via Wnt-dependent organizer formation, leading to the budding of a new organism. We observe both induction and inhibition of organizer formation depending on cellular context. Interestingly, thalidomide treatment altered budding and the developing organizer, but had little effect on the adult. Expression of Hybra1, a marker of the organizer increased upon thalidomide treatment. However when the organizer is induced by ectopic activation of Wnt signaling via GSK3 inhibition, thalidomide suppresses induction. We show that inhibition of Wnt signaling is not mediated by induction of the BMP pathway. We show that thalidomide activity on organizer formation in hydra depends on the activity of casein kinase1 and the abundance of β-catenin. Finally, we find that interstitial cells, multipotent cells which give rise to nemoatocytes, neural, digestive and germline cells, are partially responsible for the inhibitory effect of thalidomide. Copyright © 2013 Elsevier Inc. All rights reserved.

Comparative study of two models of combined pulmonary fibrosis and emphysema in mice.

PubMed

Zhang, Wan-Guang; Wu, Si-Si; He, Li; Yang, Qun; Feng, Yi-Kuan; Chen, Yue-Tao; Zhen, Guo-Hua; Xu, Yong-Jian; Zhang, Zhen-Xiang; Zhao, Jian-Ping; Zhang, Hui-Lan

2017-04-01

Combined pulmonary fibrosis and emphysema (CPFE) is an "umbrella term" encompassing emphysema and pulmonary fibrosis, but its pathogenesis is not known. We established two models of CPFE in mice using tracheal instillation with bleomycin (BLM) or murine gammaherpesvirus 68 (MHV-68). Experimental mice were divided randomly into four groups: A (normal control, n=6), B (emphysema, n=6), C (emphysema+MHV-68, n=24), D (emphysema+BLM, n=6). Group C was subdivided into four groups: C1 (sacrificed on day 367, 7 days after tracheal instillation of MHV-68); C2 (day 374; 14days); C3 (day 381; 21days); C4 (day 388; 28days). Conspicuous emphysema and interstitial fibrosis were observed in BLM and MHV-68 CPFE mouse models. However, BLM induced diffuse pulmonary interstitial fibrosis with severely diffuse pulmonary inflammation; MHV-68 induced relatively modest inflammation and fibrosis, and the inflammation and fibrosis were not diffuse, but instead around bronchioles. Inflammation and fibrosis were detectable in the day-7 subgroup and reached a peak in the day-28 subgroup in the emphysema + MHV-68 group. Levels of macrophage chemoattractant protein-1, macrophage inflammatory protein-1α, interleukin-13, and transforming growth factor-β1 in bronchoalveolar lavage fluid were increased significantly in both models. Percentage of apoptotic type-2 lung epithelial cells was significantly higher; however, all four types of cytokine and number of macrophages were significantly lower in the emphysema+MHV-68 group compared with the emphysema +BLM group. The different changes in pathology between BLM and MHV-68 mice models demonstrated different pathology subtypes of CPFE: macrophage infiltration and apoptosis of type-II lung epithelial cells increased with increasing pathology score for pulmonary fibrosis. Copyright © 2017 Elsevier GmbH. All rights reserved.

FSP1-specific SMAD2 knockout in renal tubular, endothelial, and interstitial cells reduces fibrosis and epithelial-to-mesenchymal transition in murine STZ-induced diabetic nephropathy.

PubMed

Loeffler, Ivonne; Liebisch, Marita; Allert, Stefanie; Kunisch, Elke; Kinne, Raimund W; Wolf, Gunter

2018-04-01

Extracellular matrix deposition during tubulointerstitial fibrosis (TIF), a central pathological process in patients with diabetic nephropathy (DN), is driven by locally activated, disease-relevant myofibroblasts. Myofibroblasts can arise from various cellular sources, e.g., tubular epithelial cells via a process named epithelial-to-mesenchymal transition (EMT). Transforming growth factor beta 1 (TGF-β1) and its downstream Smad signaling play a critical role in both TIF and EMT. Whereas Smad3 is one central mediator, the role of the other prominently expressed variant, Smad2, is not completely understood. In this study, we sought to analyze the role of renal Smad2 in the development of TIF and EMT during streptozotocin-induced DN by using a fibroblast-specific protein 1 (FSP1)-promotor-driven SMAD2 knockout mouse model with decreased tubular, endothelial, and interstitial Smad2 expression. In contrast to wild-type diabetic mice, diabetic SMAD2 knockout mice showed the following features: (1) significantly reduced DN and TIF (shown by KIM1 expression; periodic acid Schiff staining; collagen I and III, fibronectin, and connective tissue growth factor deposition); (2) significantly reduced tubular EMT-like changes (e.g., altered Snail1, E-cadherin, matrix metalloproteinase 2, and vimentin deposition); and (3) significantly decreased expression of myofibroblast markers (α-smooth muscle actin, FSP1). As one mechanism for the protection against diabetes-induced TIF and EMT, decreased Smad3 protein levels and, as a possible consequence, reduced TGF-β1 levels were observed in diabetic SMAD2 knockout mice. Our findings thus support the important role of Smad2 for pro-fibrotic TGF-β/Smad3 signaling in experimental DN.

Acute renal failure: unusual complication of Epstein-Barr virus-induced infectious mononucleosis.

PubMed

Lei, P S; Lowichik, A; Allen, W; Mauch, T J

2000-12-01

A 17-year-old boy with juvenile rheumatoid arthritis presented with jaundice, confusion, hemolytic anemia, thrombocytopenia, and acute renal failure secondary to titer-confirmed acute Epstein-Barr virus (EBV). Renal biopsy specimen revealed interstitial nephritis with an inflammatory infiltrate composed of cytotoxic/suppressor T cells, and interstitial mononuclear cell nuclei expressed EBV encoded RNA-1 (EBER-1) mRNA. Methylprednisolone treatment resulted in rapid improvement.

The distribution of serum albumin in the rat testis, studied by electron microscope immunocytochemistry on ultrathin frozen sections.

PubMed

Christensen, A K; Komorowski, T E; Wilson, B; Ma, S F; Stevens, R W

1985-05-01

The distribution of serum albumin is of interest in the rat testis because this protein is the principal carrier for testosterone in the plasma and interstitial fluid of this species. We have localized extravascular serum albumin in the rat testis at the electron microscope level, using gold particle immunocytochemistry on ultrathin frozen sections of tissue fixed lightly by perfusion. The same localization was obtained with three different antisera. Preabsorption and normal rabbit serum controls were negative, and Western blots of testis extracts showed major activity only at the molecular weight of albumin. Serum albumin occurred in substantial concentration throughout extracellular space in the interstitial tissue, as well as in the space between the boundary layer and the base of the seminiferous epithelium. Immunoreactivity extended between Sertoli cells, as well as around spermatogonia and early primary spermatocytes (to stage 11), but did not traverse the Sertoli-Sertoli junctions that comprise the blood-testis barrier. Macrophages in the interstitial tissue showed some endocytic activity. If perfusion fixation was carried out in a manner that flushed most of the albumin from the interstitial space, then a layer of albumin remained on the surface of Leydig cells and many macrophages but was minimal or absent on the surface of other cell types that are normally in contact with albumin, such as Sertoli cells, spermatogonia, myoid cells, lymphatic endothelium, fibroblasts, or cells of blood vessels.

ATRX Plays a Key Role in Maintaining Silencing at Interstitial Heterochromatic Loci and Imprinted Genes

PubMed Central

Voon, Hsiao P.J.; Hughes, Jim R.; Rode, Christina; De La Rosa-Velázquez, Inti A.; Jenuwein, Thomas; Feil, Robert; Higgs, Douglas R.; Gibbons, Richard J.

2015-01-01

Summary Histone H3.3 is a replication-independent histone variant, which replaces histones that are turned over throughout the entire cell cycle. H3.3 deposition at euchromatin is dependent on HIRA, whereas ATRX/Daxx deposits H3.3 at pericentric heterochromatin and telomeres. The role of H3.3 at heterochromatic regions is unknown, but mutations in the ATRX/Daxx/H3.3 pathway are linked to aberrant telomere lengthening in certain cancers. In this study, we show that ATRX-dependent deposition of H3.3 is not limited to pericentric heterochromatin and telomeres but also occurs at heterochromatic sites throughout the genome. Notably, ATRX/H3.3 specifically localizes to silenced imprinted alleles in mouse ESCs. ATRX KO cells failed to deposit H3.3 at these sites, leading to loss of the H3K9me3 heterochromatin modification, loss of repression, and aberrant allelic expression. We propose a model whereby ATRX-dependent deposition of H3.3 into heterochromatin is normally required to maintain the memory of silencing at imprinted loci. PMID:25865896

Renal myofibroblasts contract collagen I matrix lattices in vitro.

PubMed

Kelynack, K J; Hewitson, T D; Pedagogos, E; Nicholls, K M; Becker, G J

1999-01-01

Myofibroblasts, cells with both fibroblastic and smooth muscle cell features, have been implicated in renal scarring. In addition to synthetic properties, contractile features and integrin expression may allow myofibroblasts to rearrange and contract interstitial collagenous proteins. Myofibroblasts from normal rat kidneys were grown in cell-populated collagen lattices to measure cell generated contraction. Following detachment of cell populated collagen lattices, myofibroblasts progressively contracted collagen lattices, reducing lattice diameter by 42% at 24 h. Alignment of myofibroblasts, rearrangement of fibrils and beta(1) integrin expression were observed within lattices. We postulate that interstitial myofibroblasts contribute to renal scarring through manipulation of collagenous proteins. Copyright 1999 S. Karger AG, Basel

Identification of myogenic-endothelial progenitor cells in the interstitial spaces of skeletal muscle.

PubMed

Tamaki, Tetsuro; Akatsuka, Akira; Ando, Kiyoshi; Nakamura, Yoshihiko; Matsuzawa, Hideyuki; Hotta, Tomomitsu; Roy, Roland R; Edgerton, V Reggie

2002-05-13

Putative myogenic and endothelial (myo-endothelial) cell progenitors were identified in the interstitial spaces of murine skeletal muscle by immunohistochemistry and immunoelectron microscopy using CD34 antigen. Enzymatically isolated cells were characterized by fluorescence-activated cell sorting on the basis of cell surface antigen expression, and were sorted as a CD34+ and CD45- fraction. Cells in this fraction were approximately 94% positive for Sca-1, and mostly negative (<3% positive) for CD14, 31, 49, 144, c-kit, and FLK-1. The CD34+/45- cells formed colonies in clonal cell cultures and colony-forming units displayed the potential to differentiate into adipocytes, endothelial, and myogenic cells. The CD34+/45- cells fully differentiated into vascular endothelial cells and skeletal muscle fibers in vivo after transplantation. Immediately after sorting, CD34+/45- cells expressed only c-met mRNA, and did not express any other myogenic cell-related markers such as MyoD, myf-5, myf-6, myogenin, M-cadherin, Pax-3, and Pax-7. However, after 3 d of culture, these cells expressed mRNA for all myogenic markers. CD34+/45- cells were distinct from satellite cells, as they expressed Bcrp1/ABCG2 gene mRNA (Zhou et al., 2001). These findings suggest that myo-endothelial progenitors reside in the interstitial spaces of mammalian skeletal muscles, and that they can potentially contribute to postnatal skeletal muscle growth.

MEAT SCIENCE AND MUSCLE BIOLOGY SYMPOSIUM

PubMed Central

Bi, P.; Kuang, S.

2012-01-01

Stem cell niche plays a critical role in regulating the behavior and function of adult stem cells that underlie tissue growth, maintenance, and regeneration. In the skeletal muscle, stem cells, called satellite cells, contribute to postnatal muscle growth and hypertrophy, and thus, meat production in agricultural animals. Satellite cells are located adjacent to mature muscle fibers underneath a sheath of basal lamina. Microenvironmental signals from extracellular matrix mediated by the basal lamina and from the host myofiber both impinge on satellite cells to regulate their activity. Furthermore, several types of muscle interstitial cells, including intramuscular preadipocytes and connective tissue fibroblasts, have recently been shown to interact with satellite cells and actively regulate the growth and regeneration of postnatal skeletal muscles. From this regard, interstitial adipogenic cells are not only important for marbling and meat quality, but also represent an additional cellular component of the satellite cell niche. At the molecular level, these interstitial cells may interact with satellite cells through cell surface ligands, such as delta-like 1 homolog (Dlk1) protein whose overexpression is thought to be responsible for muscle hypertrophy in callipyge sheep. In fact, extracellular Dlk1 protein has been shown to promote the myogenic differentiation of satellite cells. Understanding the cellular and molecular mechanisms within the stem cell niche that regulate satellite cell differentiation and maintain muscle homeostasis may lead to promising approaches to optimizing muscle growth and composition, thus improving meat production and quality. PMID:22100594

The regulation and regulatory role of collagenase in bone

NASA Technical Reports Server (NTRS)

Partridge, N. C.; Walling, H. W.; Bloch, S. R.; Omura, T. H.; Chan, P. T.; Pearman, A. T.; Chou, W. Y.

1996-01-01

Interstitial collagenase plays an important role in both the normal and pathological remodeling of collagenous extracellular matrices, including skeletal tissues. The enzyme is a member of the family of matrix metalloproteinases. Only one rodent interstitial collagenase has been found but there are two human enzymes, human collagenase-1 and -3, the latter being the homologue of the rat enzyme. In developing rat and mouse bone, collagenase is expressed by hypertrophic chondrocytes, osteoblasts, and osteocytes, a situation that is replicated in a fracture callus. Cultured osteoblasts derived from neonatal rat calvariae show greater amounts of collagenase transcripts late in differentiation. These levels can be regulated by parathyroid hormone (PTH), retinoic acid, and insulin-like growth factors, as well as the degree of matrix mineralization. Much of the work on collagenase in bone has been derived from studies on the rat osteosarcoma cell line, UMR 106-01. All bone-resorbing agents stimulate these cells to produce collagenase mRNA and protein, with PTH being the most potent stimulator. Determination of secreted levels of collagenase has been difficult because UMR cells, normal rat osteoblasts, and rat fibroblasts possess a scavenger receptor that removes the enzyme from the extracellular space, internalizes and degrades it, thus imposing another level of control. PTH can also regulate the abundance of the receptor as well as the expression and synthesis of the enzyme. Regulation of the collagenase gene by PTH appears to involve the cAMP pathway as well as a primary response gene, possibly Fos, which then contributes to induction of the collagenase gene. The rat collagenase gene contains an activator protein-1 sequence that is necessary for basal expression, but other promoter regions may also participate in PTH regulation. Thus, there are many levels of regulation of collagenase in bone perhaps constraining what would otherwise be a rampant enzyme.

Skeletal adaptation to intramedullary pressure-induced interstitial fluid flow is enhanced in mice subjected to targeted osteocyte ablation.

PubMed

Kwon, Ronald Y; Meays, Diana R; Meilan, Alexander S; Jones, Jeremiah; Miramontes, Rosa; Kardos, Natalie; Yeh, Jiunn-Chern; Frangos, John A

2012-01-01

Interstitial fluid flow (IFF) is a potent regulatory signal in bone. During mechanical loading, IFF is generated through two distinct mechanisms that result in spatially distinct flow profiles: poroelastic interactions within the lacunar-canalicular system, and intramedullary pressurization. While the former generates IFF primarily within the lacunar-canalicular network, the latter generates significant flow at the endosteal surface as well as within the tissue. This gives rise to the intriguing possibility that loading-induced IFF may differentially activate osteocytes or surface-residing cells depending on the generating mechanism, and that sensation of IFF generated via intramedullary pressurization may be mediated by a non-osteocytic bone cell population. To begin to explore this possibility, we used the Dmp1-HBEGF inducible osteocyte ablation mouse model and a microfluidic system for modulating intramedullary pressure (ImP) to assess whether structural adaptation to ImP-driven IFF is altered by partial osteocyte depletion. Canalicular convective velocities during pressurization were estimated through the use of fluorescence recovery after photobleaching and computational modeling. Following osteocyte ablation, transgenic mice exhibited severe losses in bone structure and altered responses to hindlimb suspension in a compartment-specific manner. In pressure-loaded limbs, transgenic mice displayed similar or significantly enhanced structural adaptation to Imp-driven IFF, particularly in the trabecular compartment, despite up to ∼50% of trabecular lacunae being uninhabited following ablation. Interestingly, regression analysis revealed relative gains in bone structure in pressure-loaded limbs were correlated with reductions in bone structure in unpressurized control limbs, suggesting that adaptation to ImP-driven IFF was potentiated by increases in osteoclastic activity and/or reductions in osteoblastic activity incurred independently of pressure loading. Collectively, these studies indicate that structural adaptation to ImP-driven IFF can proceed unimpeded following a significant depletion in osteocytes, consistent with the potential existence of a non-osteocytic bone cell population that senses ImP-driven IFF independently and potentially parallel to osteocytic sensation of poroelasticity-derived IFF.

Mutant prenyltransferase-like mitochondrial protein (PLMP) and mitochondrial abnormalities in kd/kd mice

PubMed Central

Peng, Min; Jarett, Leonard; Meade, Ray; Madaio, Michael P.; Hancock, Wayne W.; George, Alfred L.; Neilson, Eric G.; Gasser, David L.

2008-01-01

Background Mice that are homozygous for the kidney disease (kd) mutation are apparently healthy for the first 8 weeks of life, but spontaneously develop a severe form of interstitial nephritis that progresses to end-stage renal disease (ESRD) by 4 to 8 months of age. By testing for linkage to microsatellite markers, we previously localized the kd gene to a YAC/BAC contig. Methods The sequence of the entire critical region was examined, and candidate genes were identified. These candidate genes were sequenced in both mutant (kd/kd) mice and normal controls. The phenotype was further characterized by immunohistochemistry and electron microscopy. Transgenic mice were constructed that carried the wild-type allele of the prime candidate gene, and this transgene was transferred to a kd/kd background by breeding. Results We have obtained evidence that kd is a mutant allele of a novel gene for a prenyltransferase-like mitochondrial protein (PLMP). This gene is alternatively spliced, with the larger gene product having one domain that resembles transprenyltransferase and another that is similar to geranylgeranyl pyrophosphate synthase. The smaller gene product includes only the first domain. An antiserum to PLMP localizes to mitochondria, and ultrastructural defects are present in the mitochondria of renal tubular epithelial cells, and to a lesser extent, hepatocytes and heart cells from kd/kd mice. In a line of kd/kd mice that carried the wild-type PLMP allele as a transgene, only 1 out of 13 animals expressed the disease by 120 days of age. Conclusion The kd allele codes for a novel protein that localizes to the mitochondria, and the kd/kd mouse has dysmorphic mitochondria in the renal tubular epithelial cells. This mouse is therefore a unique animal model for studying mechanisms that lead to tubulointerstitial nephritis. PMID:15200409

Calcium responses in subserosal interstitial cells of the guinea-pig proximal colon.

PubMed

Tamada, H; Hashitani, H

2014-01-01

In the subserosal layer between the longitudinal muscle layer and mesothelium, heterogeneous populations of interstitial cells are distributed. As the distribution of nerve elements in this layer is sparse as compared with the nerve plexus layer or tunica muscularis, there may be unique communication among subserosal interstitial cells (SSICs). This study aimed to explore functional properties of SSICs. In subserosal preparations of the guinea-pig proximal colon, changes in intracellular Ca(2+) ([Ca(2+) ]i ) were visualized using Fluo-4 Ca(2+) imaging. Immunohistochemistry was also performed to identify the SSICs exhibiting Ca(2+) transients. A majority of SSICs responded to adenosine triphosphate (ATP, 10 μM) by increasing [Ca(2+) ]i , but remained quiescent during the application of acetylcholine (10 μM). ATP-induced Ca(2+) responses were mimicked by adenosine 5'-diphosphate (10 μM), MRS2365 (10 nM) but not α, β-methylene ATP (10 μM) or uridine triphosphate (10 μM), and could be reproduced in Ca(2+) -free solution, suggesting that ATP acts via P2Y receptors, most likely P2Y1 subtype, but not P2X receptors. Live staining of the same preparations after Ca(2+) imaging indicated the ATP-sensitive SSICs were not positive for c-Kit antibody, a specific marker for gastrointestinal interstitial cells of Cajal (ICC). Immunohistochemistry identified vimentin (mesenchymal cell marker)+/Kit- and SK3 (fibroblast-like cell (FLC) marker)+/Kit- cells that had a similar morphology to the ATP-sensitive SSICs in Ca(2+) imaging. A majority of the SSICs in the guinea-pig proximal colon, presumably FLC, are capable of responding to ATP and thus may contribute to smooth muscle relaxation upon stimulation with ATP released from non-neuronal cells. © 2013 John Wiley & Sons Ltd.

Immunophenotype of infiltrating cells in protocol renal allograft biopsies from tacrolimus-versus cyclosporine-treated patients.

PubMed

Serón, Daniel; O'Valle, Francisco; Moreso, Francesc; Gomà, Montse; Hueso, Miguel; Grinyó, Josep M; Garcia del Moral, Raimundo

2007-03-15

The prevalence of subclinical rejection is lower in patients receiving tacrolimus than in patients treated with cyclosporine. However, it is not known whether this difference is related to the modulation of a specific cell immunophenotype. We perform a two case-one control study in patients treated with tacrolimus (n=44) or cyclosporine (n=22) with a protocol biopsy performed at 4 to 6 months. Immunophenotype of infiltrating cells was evaluated with monoclonal antibodies directed against CD45 (all leukocytes), CD3 (T lymphocytes), CD68 (monocytes/macrophages), and CD20 (B lymphocytes) and expressed as interstitial positive cells/mm(2). The number of interstitial CD45 (290+/-209 vs. 696+/-560; P<0.01), CD3 (121+/-84 vs. 208+/-104; P<0.01), and CD68 (155+/-232 vs. 242+/-280; P<0.05) but not CD20 (137+/-119 vs. 197+/-154) positive cells was lower in tacrolimus-treated patients. T lymphocytes and macrophages interstitial infiltration was reduced in tacrolimus treated patients evaluated with protocol biopsies in comparison to cyclosporine-treated patients.

Cannabinoid-1 (CB1) receptors regulate colonic propulsion by acting at motor neurons within the ascending motor pathways in mouse colon.

PubMed

Sibaev, Andrei; Yüce, Birol; Kemmer, Markus; Van Nassauw, Luc; Broedl, Ulli; Allescher, Hans D; Göke, Burkhard; Timmermans, Jean-Pierre; Storr, Martin

2009-01-01

Cannabinoid-1 (CB(1)) receptors on myenteric neurons are involved in the regulation of intestinal motility. Our aim was to investigate CB(1) receptor involvement in ascending neurotransmission in mouse colon and to characterize the involved structures by functional and morphological means. Presence of the CB(1) receptor was investigated by RT-PCR, and immunohistochemistry was used for colabeling studies. Myenteric reflex responses were initiated by electrical stimulation (ES) at different distances, and junction potentials (JP) were recorded from circular smooth muscle cells by intracellular recording in an unpartitioned and a partitioned recording chamber. In vivo colonic propulsion was tested in wild-type and CB(1)(-/-) mice. Immunostaining with the cytoskeletal marker peripherin showed CB(1) immunoreactivity both on Dogiel type I and type II neurons. Further neurochemical characterization revealed CB(1) on choline acetyltransferase-, calretinin-, and 5-HT-immunopositive myenteric neurons, but nitrergic neurons appeared immunonegative for CB(1) immunostaining. Solitary spindle-shaped CB(1)-immunoreactive cells in between smooth muscle cells lacked specific markers for interstitial cells of Cajal or glial cells. ES elicited neuronally mediated excitatory JP (EJP) and inhibitory JP. Gradual increases in distance resulted in a wave-like EJP with EJP amplitudes being maximal at the location of stimulating electrode 6 and a maximal EJP projection distance of approximately 18 mm. The CB(1) receptor agonist WIN 55,212-2 reduced the amplitude of EJP and was responsible for shortening the oral spreading of the excitatory impulse. In a partitioned chamber, WIN 55,212-2 reduced EJP at the separated oral sites, proving that CB(1) activation inhibits interneuron-mediated neurotransmission. These effects were absent in the presence of the CB(1) antagonist SR141716A, which, when given alone, had no effect. WIN 55,212-2 inhibited colonic propulsion in wild-type mice but not in SR141716A-pretreated wild-type or CB(1)(-/-) mice. Activation of the CB(1) receptor modulates excitatory cholinergic neurotransmission in mouse colon by reducing amplitude and spatial spreading of the ascending electrophysiological impulses. This effect on electrophysiological spreading involves CB(1)-mediated effects on motor neurons and ascending interneurons and is likely to underlie the here reported in vivo reduction in colonic propulsion.

Contribution of intravascular versus interstitial purines and nitric oxide in the regulation of exercise hyperaemia in humans

PubMed Central

Hellsten, Y; Nyberg, M; Mortensen, S P

2012-01-01

The regulation of blood flow to skeletal muscle involves a complex interaction between several locally formed vasodilators that are produced both in the skeletal muscle interstitium and intravascularly. The gas nitric oxide (NO) and the purines ATP and adenosine, are potent vasodilators that are formed by multiple cell types and released into the skeletal muscle interstitium and in plasma in response to muscle contraction. Cellular sources of ATP and NO in plasma are erythrocytes and endothelial cells, whereas interstitial sources are skeletal muscle cells and endothelial cells. Adenosine originates primarily from extracellular degradation of ATP. During exercise the concentrations of ATP and adenosine increase markedly in the interstitium with smaller increases occurring in plasma, and thus the interstitial concentration during exercise is severalfold higher than in plasma. The concentration of NO metabolites (NOx) in interstitium and plasma does not change during exercise and is similar in the two compartments. Adenosine and NO have been shown to contribute to exercise hyperaemia whereas the role of ATP remains unclear due to lack of specific purinergic receptor blockers. The relative role of intravascular versus interstitial vasodilators is not known but evidence suggests that both compartments are important. In cardiovascular disease, a reduced capacity to form adenosine in the muscle interstitium may be a contributing factor in increased peripheral vascular resistance. PMID:22733661

Pericytes in kidney fibrosis.

PubMed

Ren, Shuyu; Duffield, Jeremy S

2013-07-01

Pericytes and perivascular fibroblasts have emerged as poorly appreciated yet extensive populations of mesenchymal cells in the kidney that play important roles in homeostasis and responses to injury. This review will update readers on the evolving understanding of the biology of these cells. Fate mapping has identified pericytes and perivascular fibroblasts as the major source of pathological fibrillar matrix-forming cells in interstitial kidney disease. In other organs similar cells have been described and independent fate mapping indicates that pericytes or perivascular cells are myofibroblast progenitors in multiple organs. Over the last year, new insights into the function of pericytes in kidney homeostasis has been uncovered and new molecular pathways that regulate detachment and their transdifferentiation into pathological myofibroblasts, including Wingless/Int, ephrin, transforming growth factor β, platelet derived growth factor, and Hedgehog signaling pathways, have been reported. In addition provocative studies indicate that microRNAs, which regulate posttranscriptional gene expression, may also play important roles in their transdifferentiation. Pericytes and perivascular fibroblasts are the major source of pathological collagen fiber-forming cells in interstitial kidney diseases. New avenues of research into their activation and differentiation has identified new drug candidates for the treatment of interstitial kidney disease.

Fibrinogen-like protein 2/fibroleukin prothrombinase contributes to tumor hypercoagulability via IL-2 and IFN-γ

PubMed Central

Su, Kai; Chen, Fang; Yan, Wei-Ming; Zeng, Qi-Li; Xu, Li; Xi, Dong; Pi, Bin; Luo, Xiao-Ping; Ning, Qin

2008-01-01

AIM: To examine the role of Fibrinogen-like protein 2 (fgl2)/fibroleukin in tumor development. Fgl2 has been reported to play a vital role in the pathogenesis in MHV-3 (mouse hepatitis virus) induced fulminant and severe hepatitis, spontaneous abortion, allo- and xeno- graft rejection by mediating “immune coagulation”. METHODS: Tumor tissues from 133 patients with six types of distinct cancers and the animal tumor tissues from human hepatocellular carcinoma (HCC) model on nude mice (established from high metastasis HCC cell line MHCC97LM6) were obtained. RESULTS: Hfgl2 was detected in tumor tissues from 127 out of 133 patients as well as tumor tissues collected from human HCC nude mice. Hfgl2 was highly expressed both in cancer cells and interstitial inflammatory cells including macrophages, NK cells, and CD8+ T lymphocytes and vascular endothelial cells. Hfgl2 mRNA was localized in cells that expressed hfgl2 protein. Fibrin (nogen) co-localization with hfgl2 expression was determined by dual immunohistochemical staining. In vitro, IL-2 and IFN-γ increased hfgl2 mRNA by 10-100 folds and protein expression in both THP-1 and HUVEC cell lines. One-stage clotting assays demonstrated that THP-1 and HUVEC cells expressing hfgl2 had increased procoagulant activity following cytokines stimulation. CONCLUSION: The hfg12 contributes to the hypercoagulability in cancer and may induce tumor angiogenesis and metastasis via cytokine induction. PMID:18932275

Ultrastructural study of relationships between c-kit immunoreactive interstitial cells and other cellular elements in the human colon.

PubMed

Mazzia, C; Porcher, C; Julé, Y; Christen, M O; Henry, M

2000-05-01

C-kit immunocytochemistry was performed on ultrathin sections of human distal colon. Our attention was focused on relationships between c-kit immunoreactive interstitial cells (c-kit ICs) and muscular cells and nervous elements located in the external muscular layers of the colonic wall. C-kit ICs established membrane apposition with both nerve fibers and smooth muscle cells of, respectively, the longitudinal and circular muscle layers, the myenteric area, and the extremus submucosus plexus. C-kit ICs also surrounded the external submucosus plexus and established membrane appositions with nerve elements located inside the myenteric ganglia. These membrane appositions were observed either at the level of the c-kit IC bodies or at that of their cytoplasmic processes. In some cases, membrane appositions were observed concomitantly between the c-kit ICs, nerve fibers, and smooth muscle cells. In all the regions studied, the c-kit ICs were also found to be located in the close vicinity of blood vessels and to have established close contacts with non-immunoreactive fibroblast-like cells. The results of the present study shed essential light on the relationships of c-kit ICs with the neighboring muscle cells and nerve elements, and confirm that the intercalated c-kit ICs well fit with the so-called "interstitial cells of Cajal".

Dicarbonyl stress in clinical obesity.

PubMed

Masania, Jinit; Malczewska-Malec, Malgorzata; Razny, Urszula; Goralska, Joanna; Zdzienicka, Anna; Kiec-Wilk, Beata; Gruca, Anna; Stancel-Mozwillo, Julita; Dembinska-Kiec, Aldona; Rabbani, Naila; Thornalley, Paul J

2016-08-01

The glyoxalase system in the cytoplasm of cells provides the primary defence against glycation by methylglyoxal catalysing its metabolism to D-lactate. Methylglyoxal is the precursor of the major quantitative advanced glycation endproducts in physiological systems - arginine-derived hydroimidazolones and deoxyguanosine-derived imidazopurinones. Glyoxalase 1 of the glyoxalase system was linked to anthropometric measurements of obesity in human subjects and to body weight in strains of mice. Recent conference reports described increased weight gain on high fat diet-fed mouse with lifelong deficiency of glyoxalase 1 deficiency, compared to wild-type controls, and decreased weight gain in glyoxalase 1-overexpressing transgenic mice, suggesting a functional role of glyoxalase 1 and dicarbonyl stress in obesity. Increased methylglyoxal, dicarbonyl stress, in white adipose tissue and liver may be a mediator of obesity and insulin resistance and thereby a risk factor for development of type 2 diabetes and non-alcoholic fatty liver disease. Increased methylglyoxal formation from glyceroneogenesis on adipose tissue and liver and decreased glyoxalase 1 activity in obesity likely drives dicarbonyl stress in white adipose tissue increasing the dicarbonyl proteome and related dysfunction. The clinical significance will likely emerge from on-going clinical evaluation of inducers of glyoxalase 1 expression in overweight and obese subjects. Increased transcapillary escape rate of albumin and increased total body interstitial fluid volume in obesity likely makes levels of glycation of plasma protein unreliable indicators of glycation status in obesity as there is a shift of albumin dwell time from plasma to interstitial fluid, which decreases overall glycation for a given glycemic exposure.

Microfluidic Enhancement of Intramedullary Pressure Increases Interstitial Fluid Flow and Inhibits Bone Loss in Hindlimb Suspended Mice

PubMed Central

Kwon, Ronald Y; Meays, Diana R; Tang, W Joyce; Frangos, John A

2010-01-01

Interstitial fluid flow (IFF) has been widely hypothesized to mediate skeletal adaptation to mechanical loading. Although a large body of in vitro evidence has demonstrated that fluid flow stimulates osteogenic and antiresorptive responses in bone cells, there is much less in vivo evidence that IFF mediates loading-induced skeletal adaptation. This is due in large part to the challenges associated with decoupling IFF from matrix strain. In this study we describe a novel microfluidic system for generating dynamic intramedullary pressure (ImP) and IFF within the femurs of alert mice. By quantifying fluorescence recovery after photobleaching (FRAP) within individual lacunae, we show that microfluidic generation of dynamic ImP significantly increases IFF within the lacunocanalicular system. In addition, we demonstrate that dynamic pressure loading of the intramedullary compartment for 3 minutes per day significantly eliminates losses in trabecular and cortical bone mineral density in hindlimb suspended mice, enhances trabecular and cortical structural integrity, and increases endosteal bone formation rate. Unlike previously developed modalities for enhancing IFF in vivo, this is the first model that allows direct and dynamic modulation of ImP and skeletal IFF within mice. Given the large number of genetic tools for manipulating the mouse genome, this model is expected to serve as a powerful investigative tool in elucidating the role of IFF in skeletal adaptation to mechanical loading and molecular mechanisms mediating this process. © 2010 American Society for Bone and Mineral Research. PMID:20200992

Refining human T-cell immunotherapy of cytomegalovirus disease: a mouse model with 'humanized' antigen presentation as a new preclinical study tool.

PubMed

Lemmermann, Niels A W; Reddehase, Matthias J

2016-12-01

With the cover headline 'T cells on the attack,' the journal Science celebrated individualized cancer immunotherapy by adoptive transfer of T cells as the 'Breakthrough of the Year' 2013 (J. Couzin-Frankel in Science 342:1432-1433, 2013). It is less well recognized and appreciated that individualized T cell immunotherapy of cytomegalovirus (CMV) infection is approaching clinical application for preventing CMV organ manifestations, interstitial CMV pneumonia in particular. This coincident medical development is particularly interesting as reactivated CMV infection is a major viral complication in the state of transient immunodeficiency after the therapy of hematopoietic malignancies by hematopoietic cell transplantation (HCT). It may thus be attractive to combine T cell immunotherapy of 'minimal residual disease/leukemia (MRD)' and CMV-specific T cell immunotherapy to combat both risks in HCT recipients simultaneously, and ideally with T cells derived from the respective HLA-matched HCT donor. Although clinical trials of human CMV-specific T cell immunotherapy were promising in that the incidence of virus reactivation and disease was found to be reduced with statistical significance, animal models are still instrumental for providing 'proof of concept' by directly documenting the prevention of viral multiple-organ histopathology and organ failure under controlled conditions of the absence versus presence of the therapy, which obviously is not feasible in an individual human patient. Further, animal models can make predictions regarding parameters that determine the efficacy of T cell immunotherapy for improved study design in clinical investigations, and they allow for manipulating host and virus genetics. The latter is of particular value as it opens the possibility for epitope specificity controls that are inherently missing in clinical trials. Here, we review a recently developed new mouse model that is more approximated to human CMV-specific T cell immunotherapy by 'humanizing' antigen presentation using antigenically chimeric CMV and HLA-transgenic mice to allow for an in vivo testing of the antiviral function of human CMV-specific T cells. As an important new message, this model predicts that T cell immunotherapy is most efficient if CD4 T cells are equipped with a transduced TCR directed against an epitope presented by MHC/HLA class-I for local delivery of 'cognate' help to CD8 effector T cells at infected MHC/HLA class-II-negative host tissue cells.

Smoking-related interstitial lung diseases.

PubMed

Caminati, A; Graziano, P; Sverzellati, N; Harari, S

2010-12-01

In pulmonary pathology, a wide spectrum of morphological changes is related to the consequences of smoking, and recognizing them on surgical specimens and on small transbronchial biopsies represents a challenge for the pathologist. Respiratory bronchiolitis, also referred to as smoker's bronchiolitis, is a common histologic feature found in the lung tissue of cigarette smokers. When identified as the sole histopathologic finding in the clinical setting of symptomatic interstitial lung disease, a diagnosis of respiratory bronchiolitis-interstitial lung disease is made. Since smoking is recognized to cause a variety of histologic patterns encompassing respiratory bronchiolitis, respiratory bronchiolitis-interstitial lung disease, desquamative interstitial pneumonia and pulmonary Langerhans cell hystiocytosis, smoking-related interstitial lung disease may be a useful concept to keep in mind for the pathologists. The relationship of smoking with each of these entities has been largely established on the basis of epidemiologic evidence. Although they have been retained as distinct and separate conditions in various classifications of interstitial lung diseases, these entities share a number of clinical, radiologic, and pathologic features suggesting that they represent a spectrum of patterns of interstitial lung disease occurring in predisposed individuals who smoke. Evaluation of histologic features, particularly in surgical lung biopsy samples, is important in making the distinction between these disorders. However, even after tissue biopsy, it may sometimes be difficult to clearly separate these entities. Recently, respiratory bronchiolitis-interstitial lung disease with fibrosis has been described and postulated that this is a smoking-related condition distinct from fibrotic non-specific interstitial pneumonia.

αvβ6 Integrin Regulates Renal Fibrosis and Inflammation in Alport Mouse

PubMed Central

Hahm, Kyungmin; Lukashev, Matvey E.; Luo, Yi; Yang, William J.; Dolinski, Brian M.; Weinreb, Paul H.; Simon, Kenneth J.; Chun Wang, Li; Leone, Diane R.; Lobb, Roy R.; McCrann, Donald J.; Allaire, Normand E.; Horan, Gerald S.; Fogo, Agnes; Kalluri, Raghu; Shield, Charles F.; Sheppard, Dean; Gardner, Humphrey A.; Violette, Shelia M.

2007-01-01

The transforming growth factor (TGF)-β-inducible integrin αvβ6 is preferentially expressed at sites of epithelial remodeling and has been shown to bind and activate latent precursor TGF-β. Herein, we show that αvβ6 is overexpressed in human kidney epithelium in membranous glomerulonephritis, diabetes mellitus, IgA nephropathy, Goodpasture’s syndrome, and Alport syndrome renal epithelium. To assess the potential regulatory role of αvβ6 in renal disease, we studied the effects of function-blocking αvβ6 monoclonal antibodies (mAbs) and genetic ablation of the β6 subunit on kidney fibrosis in Col4A3−/− mice, a mouse model of Alport syndrome. Expression of αvβ6 in Alport mouse kidneys was observed primarily in cortical tubular epithelial cells and in correlation with the progression of fibrosis. Treatment with αvβ6-blocking mAbs inhibited accumulation of activated fibroblasts and deposition of interstitial collagen matrix. Similar inhibition of renal fibrosis was observed in β6-deficient Alport mice. Transcript profiling of kidney tissues showed that αvβ6-blocking mAbs significantly inhibited disease-associated changes in expression of fibrotic and inflammatory mediators. Similar patterns of transcript modulation were produced with recombinant soluble TGF-β RII treatment, suggesting shared regulatory functions of αvβ6 and TGF-β. These findings demonstrate that αvβ6 can contribute to the regulation of renal fibrosis and suggest this integrin as a potential therapeutic target. PMID:17200187

A relationship between proteinuria and acute tubulointerstitial disease in rats with experimental nephrotic syndrome

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eddy, A.A.; McCulloch, L.; Liu, E.

1991-05-01

The relationship between tubulointerstitial nephritis and proteinuria was characterized in experimental nephrosis in rats. In one group, proteinuria induced by aminonucleoside of puromycin (PAN) was reduced by using an 8% protein diet and adding the angiotensin I-converting enzyme (ACE) inhibitor enalapril to the drinking water. Two control groups were injected with saline and PAN, respectively, and fed a 27% protein diet. The first group had significantly reduced albuminuria and a definite attenuation of tubular cell injury. There was a strong positive correlation between the number of interstitial macrophages and albuminuria. The beneficial effect was reproduced by dietary-protein restriction alone, whereasmore » ACE inhibition alone had an insignificant effect on the degree of proteinuria. Depletion of circulating T lymphocytes in one group of nephrotic rats eliminated interstitial lymphocytes but did not affect interstitial macrophage influx. Inhibition of the in situ proliferation of resident interstitial macrophages by unilateral kidney irradiation failed to change the intensity of the macrophage infiltration. Treatment of rats with sodium maleate produced proximal tubular cell toxicity but interstitial inflammation did not develop, suggesting that the latter is not a nonspecific response to tubular injury. These studies demonstrate a strong relationship between tubulointerstitial nephritis and the severity of proteinuria in experimental nephrosis.« less

Expression and function of 5-HT4 receptors in the mouse enteric nervous system.

PubMed

Liu, Mintsai; Geddis, Matthew S; Wen, Ying; Setlik, Wanda; Gershon, Michael D

2005-12-01

The aim of the current study was to identify enteric 5-HT(4) splice variants, locate enteric 5-HT(4) receptors, determine the relationship, if any, of the 5-HT(4) receptor to 5-HT(1P) activity, and to ascertain the function of 5-HT(4) receptors in enteric neurophysiology. 5-HT(4a), 5-HT(4b), 5-HT(4e), and 5-HT(4f) isoforms were found in mouse brain and gut. The ratio of 5-HT(4) expression to that of the neural marker, synaptophysin, was higher in gut than in brain but was similar in small and large intestines. Submucosal 5-HT(4) expression was higher than myenteric. Although transcripts encoding 5-HT(4a) and 5-HT(4b) isoforms were more abundant, those encoding 5-HT(4e) and 5-HT(4f) were myenteric plexus specific. In situ hybridization revealed the presence of transcripts encoding 5-HT(4) receptors in subsets of enteric neurons, interstitial cells of Cajal, and smooth muscle cells. IgY antibodies to mouse 5-HT(4) receptors were raised, affinity purified, and characterized. Nerve fibers in the circular muscle and the neuropil in ganglia of both plexuses were highly 5-HT(4) immunoreactive, although only a small subset of neurons contained 5-HT(4) immunoreactivity. No 5-HT(4)-immunoreactive nerves were detected in the mucosa. 5-HT and 5-HT(1P) agonists evoked a G protein-mediated long-lasting inward current that was neither mimicked by 5-HT(4) agonists nor blocked by 5-HT(4) antagonists. In contrast, the 5-HT(4) agonists renzapride and tegaserod increased the amplitudes of nicotinic evoked excitatory postsynaptic currents. Enteric neuronal 5-HT(4) receptors thus are presynaptic and probably exert their prokinetic effects by strengthening excitatory neurotransmission.

Altered vesicular glutamate transporter distributions in the mouse cochlear nucleus following cochlear insult

PubMed Central

Heeringa, Amarins N.; Stefanescu, Roxana A.; Raphael, Yehoash; Shore, Susan E.

2015-01-01

Vesicular glutamate transporters 1 and 2 (VGLUT1 and VGLUT2) have distinct distributions in the cochlear nucleus that correspond to the sources of the labeled terminals. VGLUT1 is mainly associated with terminals of auditory nerve fibers, whereas VGLUT2 is mainly associated with glutamatergic terminals deriving from other sources that project to the cochlear nucleus (CN), including somatosensory and vestibular terminals. Previous studies in guinea pig have shown that cochlear damage results in a decrease of VGLUT1-labeled puncta and an increase in VGLUT2-labeled puncta. This indicates cross-modal compensation that is of potential importance in somatic tinnitus. To examine whether this effect is consistent across species and to provide a background for future studies, using transgenesis, the current study examines VGLUT expression profiles upon cochlear insult by intracochlear kanamycin injections in the mouse. Intracochlear kanamycin injections abolished ipsilateral ABR responses in all animals and reduced ipsilateral spiral ganglion neuron densities in animals that were sacrificed after four weeks, but not in animals that were sacrificed after three weeks. In all unilaterally deafened animals, VGLUT1 density was decreased in CN regions that receive auditory nerve fiber terminals, i.e. in the deep layer of the dorsal cochlear nucleus (DCN), in the interstitial region where the auditory nerve enters the CN, and in the magnocellular region of the antero- and posteroventral CN. In contrast, density of VGLUT2 expression was upregulated in the fusiform cell layer of the DCN and in the granule cell lamina, which are known to receive somatosensory and vestibular terminals. These results show that a cochlear insult induces cross-modal compensation in the cochlear nucleus of the mouse, confirming previous findings in guinea pig, and that these changes are not dependent on the occurrence of spiral ganglion neuron degeneration. PMID:26705736

Altered vesicular glutamate transporter distributions in the mouse cochlear nucleus following cochlear insult.

PubMed

Heeringa, A N; Stefanescu, R A; Raphael, Y; Shore, S E

2016-02-19

Vesicular glutamate transporters 1 and 2 (VGLUT1 and VGLUT2) have distinct distributions in the cochlear nucleus that correspond to sources of the labeled terminals. VGLUT1 is mainly associated with terminals of auditory nerve fibers, whereas VGLUT2 is mainly associated with glutamatergic terminals deriving from other sources that project to the cochlear nucleus (CN), including somatosensory and vestibular terminals. Previous studies in guinea pig have shown that cochlear damage results in a decrease of VGLUT1-labeled puncta and an increase in VGLUT2-labeled puncta. This indicates cross-modal compensation that is of potential importance in somatic tinnitus. To examine whether this effect is consistent across species and to provide a background for future studies, using transgenesis, the current study examines VGLUT expression profiles upon cochlear insult by intracochlear kanamycin injections in the mouse. Intracochlear kanamycin injections abolished ipsilateral ABR responses in all animals and reduced ipsilateral spiral ganglion neuron densities in animals that were sacrificed after four weeks, but not in animals that were sacrificed after three weeks. In all unilaterally deafened animals, VGLUT1 density was decreased in CN regions that receive auditory nerve fiber terminals, i.e., in the deep layer of the dorsal cochlear nucleus (DCN), in the interstitial region where the auditory nerve enters the CN, and in the magnocellular region of the antero- and posteroventral CN. In contrast, density of VGLUT2 expression was upregulated in the fusiform cell layer of the DCN and in the granule cell lamina, which are known to receive somatosensory and vestibular terminals. These results show that a cochlear insult induces cross-modal compensation in the cochlear nucleus of the mouse, confirming previous findings in guinea pig, and that these changes are not dependent on the occurrence of spiral ganglion neuron degeneration. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Intrinsic A(1) adenosine receptor activation during ischemia or reperfusion improves recovery in mouse hearts.

PubMed

Peart, J; Headrick, J P

2000-11-01

We assessed the role of A(1) adenosine receptor (A(1)AR) activation by endogenous adenosine in the modulation of ischemic contracture and postischemic recovery in Langendorff-perfused mouse hearts subjected to 20 min of total ischemia and 30 min of reperfusion. In control hearts, the rate-pressure product (RPP) and first derivative of pressure development over time (+dP/dt) recovered to 57 +/- 3 and 58 +/- 3% of preischemia, respectively. Diastolic pressure remained elevated at 20 +/- 2 mmHg (compared with 3 +/- 1 mmHg preischemia). Interstitial adenosine, assessed by microdialysis, rose from approximately 0.3 to 1.9 microM during ischemia compared with approximately 15 microM in rat heart. Nonetheless, these levels will near maximally activate A(1)ARs on the basis of effects of exogenous adenosine and 2-chloroadenosine. Neither A(1)AR blockade with 200 nM 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) during the ischemic period alone nor A(1)AR activation with 50 nM N(6)-cyclopentyladenosine altered rapidity or extent of ischemic contracture. However, ischemic DPCPX treatment significantly depressed postischemic recovery of RPP and +dP/dt (44 +/- 3 and 40 +/- 4% of preischemia, respectively). DPCPX treatment during the reperfusion period alone also reduced recovery of RPP and +dP/dt (to 44 +/- 2 and 47 +/- 2% of preischemia, respectively). These data indicate that 1) interstitial adenosine is lower in mouse versus rat myocardium during ischemia, 2) A(1)AR activation by endogenous adenosine or exogenous agonists does not modify ischemic contracture in murine myocardium, 3) A(1)AR activation by endogenous adenosine during ischemia attenuates postischemic stunning, and 4) A(1)AR activation by endogenous adenosine during the reperfusion period also improves postischemic contractile recovery.

Structural effects in the interstitial solid solution system (La,Ce)(Fe,Si)13Cx-H: Correlation with hydrogenation kinetics

NASA Astrophysics Data System (ADS)

Hai, X.; Porcher, F.; Mayer, C.; Miraglia, S.

2018-02-01

Steady state and in-situ neutron powder diffraction on selected compositions of the magneto-caloric (La,Ce)(Fe,Si)13CxHy compounds has been used to locate the sites accommodated by the interstitial species and to reveal the structural modifications (breathing) that occur upon metal substitution and/or interstitial insertion. The latter type of measurement in which the sequential filling of interstitial sites is followed allows one to extract some useful hydrogenation kinetics data. This structural investigation has allowed to precise the deformations undergone by the complex metallic alloys La(Fe,Si)13 when subjected to light interstitial insertion or rare earth substitution at the cation site of the NaZn13-structure type. We attempt to correlate hydrogenation kinetics variations (depression or enhancement of the hydrogen absorption rate) with a particular inhomogeneous cell variation (breathing) and bonding of the NaZn13 structure-type.

Anode for a secondary, high-temperature electrochemical cell

DOEpatents

Vissers, Donald R.; Tani, Benjamin S.

1976-01-01

A high-temperature, secondary electrochemical cell includes an anode containing lithium, an electrolyte containing lithium ions and a cathode containing a chalcogen material such as sulfur or a metallic sulfide. The anode includes a porous substrate formed of, for instance, a compacted mass of entangled metallic fibers providing interstitial crevices for receiving molten lithium metal. The surfaces of the interstitial crevices are provided with a coating of cobalt metal to enhance the retention of the molten lithium metal within the substrate.

Hydrogen Passivation of Interstitial Zn Defects in Heteroepitaxial InP Cell Structures and Influence on Device Characteristics

NASA Technical Reports Server (NTRS)

Ringel, S. A.; Chatterjee, B.

2004-01-01

Hydrogen passivation of heteroepitaxial InP solar cells is of recent interest for deactivation of dislocations and other defects caused by the cell/substrate lattice mismatch that currently limit the photovoltaic performance of these devices. In this paper we present strong evidence that, in addition to direct hydrogen-dislocation interactions, hydrogen forms complexes with the high concentration of interstitial Zn defects present within the p(+) Zn-doped emitter of MOCVD-grown heteroepitaxial InP devices, resulting in a dramatic increase of the forward bias turn-on voltage by as much as 280 mV, from 680 mV to 960 mV. This shift is reproducible and thermally reversible and no such effect is observed for either n(+)p structures or homoepitaxial p(+)n structures grown under identical conditions. A combination of photoluminescence (PL), electrochemical C-V dopant profiling, SIMS and I-V measurements were performed on a set of samples having undergone a matrix of hydrogenation and post-hydrogenation annealing conditions to investigate the source of this voltage enhancement and confirm the expected role of interstitial Zn and hydrogen. A precise correlation between all measurements is demonstrated which indicates that Zn interstitials within the p(+) emitter and their interaction with hydrogen are indeed responsible for this device behavior.

3-compartment talaporfin sodium pharmacokinetic model by optimization using fluorescence measurement data from canine skin to estimate the concentration in interstitial space

NASA Astrophysics Data System (ADS)

Uno, Yuko; Ogawa, Emiyu; Aiyoshi, Eitaro; Arai, Tsunenori

2018-02-01

We constructed the 3-compartment talaporfin sodium pharmacokinetic model for canine by an optimization using the fluorescence measurement data from canine skin to estimate the concentration in the interstitial space. It is difficult to construct the 3-compartment model consisted of plasma, interstitial space, and cell because there is a lack of the dynamic information. Therefore, we proposed the methodology to construct the 3-compartment model using the measured talaporfin sodium skin fluorescence change considering originated tissue part by a histological observation. In a canine animal experiment, the talaporfin sodium concentration time history in plasma was measured by a spectrophotometer with a prepared calibration curve. The time history of talaporfin sodium Q-band fluorescence on left femoral skin of a beagle dog excited by talaporfin sodium Soret-band of 409 nm was measured in vivo by our previously constructed measurement system. The measured skin fluorescence was classified to its source, that is, specific ratio of plasma, interstitial space, and cell. We represented differential rate equations of the talaporfin sodium concentration in plasma, interstitial space, cell. The specific ratios and a converting constant to obtain absolute value of skin concentration were arranged. Minimizing the squared error of the difference between the measured fluorescence data and calculated concentration by the conjugate gradient method in MATLAB, the rate constants in the 3-compartment model were determined. The accuracy of the fitting operation was confirmed with determination coefficient of 0.98. We could construct the 3-compartment pharmacokinetic model for canine using the measured talaporfin sodium fluorescence change from canine skin.

Overview of the cellular and molecular basis of kidney fibrosis

PubMed Central

Eddy, Allison A

2014-01-01

The common pathogenetic pathway of progressive injury in patients with chronic kidney disease (CKD) is epitomized as normal kidney parenchymal destruction due to scarring (fibrosis). Understanding the fundamental pathways that lead to renal fibrosis is essential in order to develop better therapeutic options for human CKD. Although complex, four cellular responses are pivotal. (1) An interstitial inflammatory response that has multiple consequences—some harmful and others healing. (2) The appearance of a unique interstitial cell population of myofibroblasts, primarily derived from kidney stromal cells (fibroblasts and pericytes), that are the primary source of the various extracellular matrix proteins that form interstitial scars. (3) Tubular epithelial cells that have variable and time-dependent roles as early responders to injury and later as victims of fibrosis due to the loss of their regenerative abilities. (4) Loss of interstitial capillary integrity that compromises oxygen delivery and leads to a vicious cascade of hypoxia–oxidant stress that accentuates injury and fibrosis. In the absence of adequate angiogenic responses, a healthy interstitial capillary network is not maintained. The fibrotic ‘scar' that typifies CKD is an interesting consortium of multifunctional macromolecules that not only change in composition and structure over time, but can be degraded via extracellular and intracellular proteases. Although transforming growth factor beta appears to be the primary driver of kidney fibrosis, a vast array of additional molecules may have modulating roles. The importance of genetic and epigenetic factors is increasingly appreciated. An intriguing but incompletely understood cardiorenal syndrome underlies the high morbidity and mortality rates that develop in association with progressive kidney fibrosis. PMID:25401038

Computational Modeling of 3D Tumor Growth and Angiogenesis for Chemotherapy Evaluation

PubMed Central

Tang, Lei; van de Ven, Anne L.; Guo, Dongmin; Andasari, Vivi; Cristini, Vittorio; Li, King C.; Zhou, Xiaobo

2014-01-01

Solid tumors develop abnormally at spatial and temporal scales, giving rise to biophysical barriers that impact anti-tumor chemotherapy. This may increase the expenditure and time for conventional drug pharmacokinetic and pharmacodynamic studies. In order to facilitate drug discovery, we propose a mathematical model that couples three-dimensional tumor growth and angiogenesis to simulate tumor progression for chemotherapy evaluation. This application-oriented model incorporates complex dynamical processes including cell- and vascular-mediated interstitial pressure, mass transport, angiogenesis, cell proliferation, and vessel maturation to model tumor progression through multiple stages including tumor initiation, avascular growth, and transition from avascular to vascular growth. Compared to pure mechanistic models, the proposed empirical methods are not only easy to conduct but can provide realistic predictions and calculations. A series of computational simulations were conducted to demonstrate the advantages of the proposed comprehensive model. The computational simulation results suggest that solid tumor geometry is related to the interstitial pressure, such that tumors with high interstitial pressure are more likely to develop dendritic structures than those with low interstitial pressure. PMID:24404145

New transmitters and new targets in the autonomic nervous system.

PubMed

Barajas-López, C; Huizinga, J D

1993-12-01

Several recent findings have made research into the autonomic nervous system even more exciting, such as the revelation that nitric oxide is a major neurotransmitter, the delineation of the physiological roles for purines and vasoactive intestinal peptide, and the discovery that the interstitial cells of Cajal are major target cells for enteric innervation. Nitric oxide is probably the major neurotransmitter evoking inhibitory junction potentials in smooth muscle. ATP is a mediator of non-adrenergic non-cholinergic enteric innervation, as well as being a fast neurotransmitter in peripheral and autonomic neuro-neuronal synapses. The interactions between enteric nerves and both immune cells and interstitial cells of Cajal (as pacemaker cells of gut smooth muscle) are forcing a rethink of many aspects of gut physiology.

Immunohistochemical and ultrastructural characteristics of interstitial cells of Cajal in the rabbit duodenum. Presence of a single cilium

PubMed Central

Junquera, Concepción; Martínez-Ciriano, Carmen; Castiella, Tomás; Serrano, Pedro; Azanza, María Jesús; Ramón y Cajal Junquera, Santiago

2007-01-01

Abstract Santiago Ramón y Cajal discovered a new type of cell related to the myenteric plexus and also to the smooth muscle cells of the circular muscle layer of the intestine. Based on their morphology, relationships and staining characteristics, he considered these cells as primitive neurons. One century later, despite major improvements in cell biology, the interstitial cells of Cajal (ICCs) are still controversial for many researchers. The aim of study was to perform an immunohistochemical and ultrastructural characterization of the ICCs in the rabbit duo-denum. We have found interstitial cells that are positive for c-Kit, CD34 and nestin and are also positive for Ki67 protein, tightly associated with somatic cell proliferation. By means of electron microscopy, we describe ICCs around enteric ganglia. They present triangular or spindle forms and a very voluminous nucleus with scarce per-inuclear chromatin surrounded by a thin perinuclear cytoplasm that expands with long cytoplasmic processes. ICC processes penetrate among the smooth muscle cells and couple with the processes of other ICCs located in the connective tissue of the circular muscle layer and establish a three-dimensional network. Intercellular con-tacts by means of gap-like junctions are frequent. ICCs also establish gap-like junctions with smooth muscle cells. We also observe a population of interstitial cells of stellate morphology in the connective tissue that sur-rounds the muscle bundles in the circular muscle layer, usually close to nervous trunks. These cells establish different types of contacts with the muscle cells around them. In addition, the presence of a single cilium show-ing a structure 9 + 0 in an ICC is demonstrated for the first time. In conclusion, we report positive staining c-kit, CD34, nestin and Ki 67. ICCs fulfilled the usual transmission electron microscopy (TEM) criteria. A new ultrastructural characteristic of at least some ICCs is demonstrated: the presence of a single cilium. Some populations of ICCs in the rabbit duodenum present certain immunohistochemical and ultrastructural characteristics that often are present in progenitor cells. PMID:17760839

Tubule-Derived Wnts Are Required for Fibroblast Activation and Kidney Fibrosis.

PubMed

Zhou, Dong; Fu, Haiyan; Zhang, Lu; Zhang, Ke; Min, Yali; Xiao, Liangxiang; Lin, Lin; Bastacky, Sheldon I; Liu, Youhua

2017-08-01

Cell-cell communication via Wnt ligands is necessary in regulating embryonic development and has been implicated in CKD. Because Wnt ligands are ubiquitously expressed, the exact cellular source of the Wnts involved in CKD remains undefined. To address this issue, we generated two conditional knockout mouse lines in which Wntless (Wls), a dedicated cargo receptor that is obligatory for Wnt secretion, was selectively ablated in tubular epithelial cells or interstitial fibroblasts. Blockade of Wnt secretion by genetic deletion of Wls in renal tubules markedly inhibited myofibroblast activation and reduced renal fibrosis after unilateral ureteral obstruction. This effect associated with decreased activation of β -catenin and downstream gene expression and preserved tubular epithelial integrity. In contrast, fibroblast-specific deletion of Wls exhibited little effect on the severity of renal fibrosis after obstructive or ischemia-reperfusion injury. In vitro , incubation of normal rat kidney fibroblasts with tubule-derived Wnts promoted fibroblast proliferation and activation. Furthermore, compared with kidney specimens from patients without CKD, biopsy specimens from patients with CKD also displayed increased expression of multiple Wnt proteins, predominantly in renal tubular epithelium. These results illustrate that tubule-derived Wnts have an essential role in promoting fibroblast activation and kidney fibrosis via epithelial-mesenchymal communication. Copyright © 2017 by the American Society of Nephrology.

[Modern Views on Children's Interstitial Lung Disease].

PubMed

Boĭtsova, E V; Beliashova, M A; Ovsiannikov, D Iu

2015-01-01

Interstitial lung diseases (ILD, diffuse lung diseases) are a heterogeneous group of diseases in which a pathological process primarily involved alveoli and perialveolar interstitium, resulting in impaired gas exchange, restrictive changes of lung ventilation function and diffuse interstitial changes detectable by X-ray. Children's interstitial lung diseases is an topical problem ofpediatricpulmonoogy. The article presents current information about classification, epidemiology, clinical presentation, diagnostics, treatment and prognosis of these rare diseases. The article describes the differences in the structure, pathogenesis, detection of various histological changes in children's ILD compared with adult patients with ILD. Authors cite an instance of registers pediatric patients with ILD. The clinical semiotics of ILD, the possible results of objective research, the frequency of symptoms, the features of medical history, the changes detected on chest X-rays, CT semiotics described in detail. Particular attention was paid to interstitial lung diseases, occurring mainly in newborns and children during the first two years of life, such as congenital deficiencies of surfactant proteins, neuroendocrine cell hyperplasia of infancy, pulmonary interstitial glycogenosis. The diagnostic program for children's ILD, therapy options are presented in this article.

Characterization of the Tumor Secretome from Tumor Interstitial Fluid (TIF).

PubMed

Gromov, Pavel; Gromova, Irina

2016-01-01

Tumor interstitial fluid (TIF) surrounds and perfuses bodily tumorigenic tissues and cells, and can accumulate by-products of tumors and stromal cells in a relatively local space. Interstitial fluid offers several important advantages for biomarker and therapeutic target discovery, especially for cancer. Here, we describe the most currently accepted method for recovering TIF from tumor and nonmalignant tissues that was initially performed using breast cancer tissue. TIF recovery is achieved by passive extraction of fluid from small, surgically dissected tissue specimens in phosphate-buffered saline. We also present protocols for hematoxylin and eosin (H&E) staining of snap-frozen and formalin-fixed, paraffin-embedded (FFPE) tumor sections and for proteomic profiling of TIF and matched tumor samples by high-resolution two-dimensional gel electrophoresis (2D-PAGE) to enable comparative analysis of tumor secretome and paired tumor tissue.

Human bone perivascular niche-on-a-chip for studying metastatic colonization.

PubMed

Marturano-Kruik, Alessandro; Nava, Michele Maria; Yeager, Keith; Chramiec, Alan; Hao, Luke; Robinson, Samuel; Guo, Edward; Raimondi, Manuela Teresa; Vunjak-Novakovic, Gordana

2018-02-06

Eight out of 10 breast cancer patients die within 5 years after the primary tumor has spread to the bones. Tumor cells disseminated from the breast roam the vasculature, colonizing perivascular niches around blood capillaries. Slow flows support the niche maintenance by driving the oxygen, nutrients, and signaling factors from the blood into the interstitial tissue, while extracellular matrix, endothelial cells, and mesenchymal stem cells regulate metastatic homing. Here, we show the feasibility of developing a perfused bone perivascular niche-on-a-chip to investigate the progression and drug resistance of breast cancer cells colonizing the bone. The model is a functional human triculture with stable vascular networks within a 3D native bone matrix cultured on a microfluidic chip. Providing the niche-on-a-chip with controlled flow velocities, shear stresses, and oxygen gradients, we established a long-lasting, self-assembled vascular network without supplementation of angiogenic factors. We further show that human bone marrow-derived mesenchymal stem cells, which have undergone phenotypical transition toward perivascular cell lineages, support the formation of capillary-like structures lining the vascular lumen. Finally, breast cancer cells exposed to interstitial flow within the bone perivascular niche-on-a-chip persist in a slow-proliferative state associated with increased drug resistance. We propose that the bone perivascular niche-on-a-chip with interstitial flow promotes the formation of stable vasculature and mediates cancer cell colonization.

Tubular Epithelial NF-κB Activity Regulates Ischemic AKI

PubMed Central

Vigolo, Emilia; Hinze, Christian; Park, Joon-Keun; Roël, Giulietta; Balogh, András; Choi, Mira; Wübken, Anne; Cording, Jimmi; Blasig, Ingolf E.; Luft, Friedrich C.; Scheidereit, Claus; Schmidt-Ott, Kai M.; Schmidt-Ullrich, Ruth; Müller, Dominik N.

2016-01-01

NF-κB is a key regulator of innate and adaptive immunity and is implicated in the pathogenesis of AKI. The cell type–specific functions of NF-κB in the kidney are unknown; however, the pathway serves distinct functions in immune and tissue parenchymal cells. We analyzed tubular epithelial-specific NF-κB signaling in a mouse model of ischemia-reperfusion injury (IRI)–induced AKI. NF-κB reporter activity and nuclear localization of phosphorylated NF-κB subunit p65 analyses in mice revealed that IRI induced widespread NF-κB activation in renal tubular epithelia and in interstitial cells that peaked 2–3 days after injury. To genetically antagonize tubular epithelial NF-κB activity, we generated mice expressing the human NF-κB super-repressor IκBαΔN in renal proximal, distal, and collecting duct epithelial cells. Compared with control mice, these mice exhibited improved renal function, reduced tubular apoptosis, and attenuated neutrophil and macrophage infiltration after IRI-induced AKI. Furthermore, tubular NF-κB–dependent gene expression profiles revealed temporally distinct functional gene clusters for apoptosis, chemotaxis, and morphogenesis. Primary proximal tubular cells isolated from IκBαΔN-expressing mice and exposed to hypoxia-mimetic agent cobalt chloride exhibited less apoptosis and expressed lower levels of chemokines than cells from control mice did. Our results indicate that postischemic NF-κB activation in renal tubular epithelia aggravates tubular injury and exacerbates a maladaptive inflammatory response. PMID:26823548

Telocytes and putative stem cells in the lungs: electron microscopy, electron tomography and laser scanning microscopy.

PubMed

Popescu, Laurentiu M; Gherghiceanu, Mihaela; Suciu, Laura C; Manole, Catalin G; Hinescu, Mihail E

2011-09-01

This study describes a novel type of interstitial (stromal) cell - telocytes (TCs) - in the human and mouse respiratory tree (terminal and respiratory bronchioles, as well as alveolar ducts). TCs have recently been described in pleura, epicardium, myocardium, endocardium, intestine, uterus, pancreas, mammary gland, etc. (see www.telocytes.com ). TCs are cells with specific prolongations called telopodes (Tp), frequently two to three per cell. Tp are very long prolongations (tens up to hundreds of μm) built of alternating thin segments known as podomers (≤ 200 nm, below the resolving power of light microscope) and dilated segments called podoms, which accommodate mitochondria, rough endoplasmic reticulum and caveolae. Tp ramify dichotomously, making a 3-dimensional network with complex homo- and heterocellular junctions. Confocal microscopy reveals that TCs are c-kit- and CD34-positive. Tp release shed vesicles or exosomes, sending macromolecular signals to neighboring cells and eventually modifying their transcriptional activity. At bronchoalveolar junctions, TCs have been observed in close association with putative stem cells (SCs) in the subepithelial stroma. SCs are recognized by their ultrastructure and Sca-1 positivity. Tp surround SCs, forming complex TC-SC niches (TC-SCNs). Electron tomography allows the identification of bridging nanostructures, which connect Tp with SCs. In conclusion, this study shows the presence of TCs in lungs and identifies a TC-SC tandem in subepithelial niches of the bronchiolar tree. In TC-SCNs, the synergy of TCs and SCs may be based on nanocontacts and shed vesicles.

Hyperandrogenism from an ovarian interstitial-cell tumor in an alpaca.

PubMed

Gilbert, Rosanne; Kutzler, Michelle; Valentine, Beth A; Semevolos, Stacy

2006-11-01

An 8-year-old intact female Huacaya alpaca (Lama pacos) was presented for recent development of male behavior. Serum testosterone concentration was determined to be 969.1 pg/ml by using radioimmunoassay, while the range in 33 healthy female adult intact alpacas was 11.7-62.1 pg/ml. An ovarian mass was suspected, and an exploratory laparotomy was performed. A tan mass was present on the left ovary. Histologically, the mass was composed of closely packed, plump, polygonal cells with central round nuclei with granular chromatin and abundant eosinophilic finely granular to vesiculate cytoplasm. An ovarian benign interstitial (Leydig) cell tumor was diagnosed.

Zika Virus Trafficking and Interactions in the Human Male Reproductive Tract.

PubMed

da Silva, Lucia Regina Cangussu

2018-05-11

Sexual transmission of Zika virus (ZIKV) is a matter of great concern. Infectious viral particles can be shed in semen for as long as six months after infection and can be transferred to male and female sexual partners during unprotected sexual intercourse. The virus can be found inside spermatozoa and could be directly transferred to the oocyte during fertilization. Sexual transmission of ZIKV can contribute to the rise in number of infected individuals in endemic areas as well as in countries where the mosquito vector does not thrive. There is also the possibility, as has been demonstrated in mouse models, that the vaginal deposition of ZIKV particles present in semen could lead to congenital syndrome. In this paper, we review the current literature to understand ZIKV trafficking from the bloodstream to the human male reproductive tract and viral interactions with host cells in interstitial spaces, tubule walls, annexed glands and semen. We hope to highlight gaps to be filled by future research and potential routes for vaccine and antiviral development.

The effects of Robertsonian fusions on chiasma frequency and distribution in the house mouse (Mus musculus domesticus) from a hybrid zone in northern Scotland.

PubMed

Bidau, C J; Giménez, M D; Palmer, C L; Searle, J B

2001-09-01

Chiasma frequency and distribution were studied in male Mus musculus domesticus from the John O'Groats-standard chromosomal hybrid zone in northern Scotland. Individuals of the John O'Groats race (2n=32; homozygous for the Robertsonian fusions 4.10, 6.13, 9.12 and 11.14) and the standard race (2n=40, all telocentric), and hybrids with various karyotypes, were examined. Chiasma frequency was significantly negatively correlated with the number of Robertsonian configurations in the meiotic cell. The decrease of chiasma frequency can be attributed to intrachromosomal effects that reduce the number of chiasmata in Robertsonian bivalents (formed in homozygotes for Robertsonian fusions) and trivalents (formed in heterozygotes). However, the reduction is more pronounced in Robertsonian bivalents and is related to a shift of chiasmata to the distal ends of the chromosome arms. A different type of repatterning occurs in trivalents where there is a significant increase in proximal and interstitial chiasmata.

Targeted inactivation of the murine Abca3 gene leads to respiratory failure in newborns with defective lamellar bodies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hammel, Markus; Michel, Geert; Hoefer, Christina

2007-08-10

Mutations in the human ABCA3 gene, encoding an ABC-transporter, are associated with respiratory failure in newborns and pediatric interstitial lung disease. In order to study disease mechanisms, a transgenic mouse model with a disrupted Abca3 gene was generated by targeting embryonic stem cells. While heterozygous animals developed normally and were fertile, individuals homozygous for the altered allele (Abca3-/-) died within one hour after birth from respiratory failure, ABCA3 protein being undetectable. Abca3-/- newborns showed atelectasis of the lung in comparison to a normal gas content in unaffected or heterozygous littermates. Electron microscopy demonstrated the absence of normal lamellar bodies inmore » type II pneumocytes. Instead, condensed structures with apparent absence of lipid content were found. We conclude that ABCA3 is required for the formation of lamellar bodies and lung surfactant function. The phenotype of respiratory failure immediately after birth corresponds to the clinical course of severe ABCA3 mutations in human newborns.« less

Fgf10-positive cells represent a progenitor cell population during lung development and postnatally

PubMed Central

El Agha, Elie; Herold, Susanne; Alam, Denise Al; Quantius, Jennifer; MacKenzie, BreAnne; Carraro, Gianni; Moiseenko, Alena; Chao, Cho-Ming; Minoo, Parviz; Seeger, Werner; Bellusci, Saverio

2014-01-01

The lung mesenchyme consists of a widely heterogeneous population of cells that play crucial roles during development and homeostasis after birth. These cells belong to myogenic, adipogenic, chondrogenic, neuronal and other lineages. Yet, no clear hierarchy for these lineages has been established. We have previously generated a novel Fgf10iCre knock-in mouse line that allows lineage tracing of Fgf10-positive cells during development and postnatally. Using these mice, we hereby demonstrate the presence of two waves of Fgf10 expression during embryonic lung development: the first wave, comprising Fgf10-positive cells residing in the submesothelial mesenchyme at early pseudoglandular stage (as well as their descendants); and the second wave, comprising Fgf10-positive cells from late pseudoglandular stage (as well as their descendants). Our lineage-tracing data reveal that the first wave contributes to the formation of parabronchial and vascular smooth muscle cells as well as lipofibroblasts at later developmental stages, whereas the second wave does not give rise to smooth muscle cells but to lipofibroblasts as well as an Nkx2.1- E-Cad- Epcam+ Pro-Spc+ lineage that requires further in-depth analysis. During alveologenesis, Fgf10-positive cells give rise to lipofibroblasts rather than alveolar myofibroblasts, and during adult life, a subpopulation of Fgf10-expressing cells represents a pool of resident mesenchymal stromal (stem) cells (MSCs) (Cd45- Cd31- Sca-1+). Taken together, we show for the first time that Fgf10-expressing cells represent a pool of mesenchymal progenitors in the embryonic and postnatal lung. Our findings suggest that Fgf10-positive cells could be useful for developing stem cell-based therapies for treating interstitial lung diseases. PMID:24353064

Macrophage and epithelial cell H-ferritin expression regulates renal inflammation

PubMed Central

Bolisetty, Subhashini; Zarjou, Abolfazl; Hull, Travis D.; Traylor, Amie; Perianayagam, Anjana; Joseph, Reny; Kamal, Ahmed I; Arosio, Paolo; Soares, Miguel P; Jeney, Viktoria; Balla, Jozsef; George, James F.; Agarwal, Anupam

2015-01-01

Inflammation culminating in fibrosis contributes to progressive kidney disease. Crosstalk between the tubular epithelium and interstitial cells regulates inflammation by a coordinated release of cytokines and chemokines. Here we studied the role of heme oxygenase-1 (HO-1) and the heavy subunit of ferritin (FtH) in macrophage polarization and renal inflammation. Deficiency in HO-1 was associated with increased FtH expression, accumulation of macrophages with a dysregulated polarization profile, and increased fibrosis following unilateral ureteral obstruction in mice; a model of renal inflammation and fibrosis. Macrophage polarization in vitro was predominantly dependent on FtH expression in isolated bone marrow-derived mouse monocytes. Utilizing transgenic mice with conditional deletion of FtH in the proximal tubules (FtHPT−/−) or myeloid cells (FtHLysM−/−), we found that myeloid FtH deficiency did not affect polarization or accumulation of macrophages in the injured kidney compared to wild-type (FtH+/+) controls. However, tubular FtH deletion led to a marked increase in pro-inflammatory macrophages. Furthermore, injured kidneys from FtHPT−/− mice expressed significantly higher levels of inflammatory chemokines and fibrosis compared to kidneys from FtH+/+ and FtHLysM−/− mice. Thus, there are differential effects of FtH in macrophages and epithelial cells, which underscores the critical role of FtH in tubular-macrophage crosstalk during kidney injury. PMID:25874599

Vinpocetine inhibits NF-kappaB-dependent inflammation via an IKK-dependent but PDE-independent mechanism.

PubMed

Jeon, Kye-Im; Xu, Xiangbin; Aizawa, Toru; Lim, Jae Hyang; Jono, Hirofumi; Kwon, Dong-Seok; Abe, Jun-Ichi; Berk, Bradford C; Li, Jian-Dong; Yan, Chen

2010-05-25

Inflammation is a hallmark of many diseases, such as atherosclerosis, chronic obstructive pulmonary disease, arthritis, infectious diseases, and cancer. Although steroids and cyclooxygenase inhibitors are effective antiinflammatory therapeutical agents, they may cause serious side effects. Therefore, developing unique antiinflammatory agents without significant adverse effects is urgently needed. Vinpocetine, a derivative of the alkaloid vincamine, has long been used for cerebrovascular disorders and cognitive impairment. Its role in inhibiting inflammation, however, remains unexplored. Here, we show that vinpocetine acts as an antiinflammatory agent in vitro and in vivo. In particular, vinpocetine inhibits TNF-alpha-induced NF-kappaB activation and the subsequent induction of proinflammatory mediators in multiple cell types, including vascular smooth muscle cells, endothelial cells, macrophages, and epithelial cells. We also show that vinpocetine inhibits monocyte adhesion and chemotaxis, which are critical processes during inflammation. Moreover, vinpocetine potently inhibits TNF-alpha- or LPS-induced up-regulation of proinflammatory mediators, including TNF-alpha, IL-1beta, and macrophage inflammatory protein-2, and decreases interstitial infiltration of polymorphonuclear leukocytes in a mouse model of TNF-alpha- or LPS-induced lung inflammation. Interestingly, vinpocetine inhibits NF-kappaB-dependent inflammatory responses by directly targeting IKK, independent of its well-known inhibitory effects on phosphodiesterase and Ca(2+) regulation. These studies thus identify vinpocetine as a unique antiinflammatory agent that may be repositioned for the treatment of many inflammatory diseases.

Vinpocetine inhibits NF-κB–dependent inflammation via an IKK-dependent but PDE-independent mechanism

PubMed Central

Jeon, Kye-Im; Xu, Xiangbin; Aizawa, Toru; Lim, Jae Hyang; Jono, Hirofumi; Kwon, Dong-Seok; Berk, Bradford C.; Li, Jian-Dong; Yan, Chen

2010-01-01

Inflammation is a hallmark of many diseases, such as atherosclerosis, chronic obstructive pulmonary disease, arthritis, infectious diseases, and cancer. Although steroids and cyclooxygenase inhibitors are effective antiinflammatory therapeutical agents, they may cause serious side effects. Therefore, developing unique antiinflammatory agents without significant adverse effects is urgently needed. Vinpocetine, a derivative of the alkaloid vincamine, has long been used for cerebrovascular disorders and cognitive impairment. Its role in inhibiting inflammation, however, remains unexplored. Here, we show that vinpocetine acts as an antiinflammatory agent in vitro and in vivo. In particular, vinpocetine inhibits TNF-α–induced NF-κB activation and the subsequent induction of proinflammatory mediators in multiple cell types, including vascular smooth muscle cells, endothelial cells, macrophages, and epithelial cells. We also show that vinpocetine inhibits monocyte adhesion and chemotaxis, which are critical processes during inflammation. Moreover, vinpocetine potently inhibits TNF-α- or LPS-induced up-regulation of proinflammatory mediators, including TNF-α, IL-1β, and macrophage inflammatory protein-2, and decreases interstitial infiltration of polymorphonuclear leukocytes in a mouse model of TNF-α- or LPS-induced lung inflammation. Interestingly, vinpocetine inhibits NF-κB–dependent inflammatory responses by directly targeting IKK, independent of its well-known inhibitory effects on phosphodiesterase and Ca2+ regulation. These studies thus identify vinpocetine as a unique antiinflammatory agent that may be repositioned for the treatment of many inflammatory diseases. PMID:20448200

Acute and chronic rejection: compartmentalization and kinetics of counterbalancing signals in cardiac transplants.

PubMed

Kaul, A M K; Goparaju, S; Dvorina, N; Iida, S; Keslar, K S; de la Motte, C A; Valujskikh, A; Fairchild, R L; Baldwin, W M

2015-02-01

Acute and chronic rejection impact distinct compartments of cardiac allografts. Intramyocardial mononuclear cell infiltrates define acute rejection, whereas chronic rejection affects large arteries. Hearts transplanted from male to female C57BL/6 mice undergo acute rejection with interstitial infiltrates at 2 weeks that resolve by 6 weeks when large arteries develop arteriopathy. These processes are dependent on T cells because no infiltrates developed in T cell-deficient mice and transfer of CD4 T cells restored T cell as well as macrophage infiltrates and ultimately neointima formation. Markers of inflammatory macrophages were up-regulated in the interstitium acutely and decreased as markers of wound healing macrophages increased chronically. Programmed cell death protein, a negative costimulator, and its ligand PDL1 were up-regulated in the interstitium during resolution of acute rejection. Blocking PDL1:PD1 interactions in the acute phase increased interstitial T cell infiltrates. Toll-like receptor (TLR) 4 and its endogenous ligand hyaluronan were increased in arteries with neointimal expansion. Injection of hyaluronan fragments increased intragraft production of chemokines. Our data indicate that negative costimulatory pathways are critical for the resolution of acute interstitial infiltrates. In the arterial compartment recognition of endogenous ligands including hyaluronan by the innate TLRs may support the progression of arteriopathy. © Copyright 2015 The American Society of Transplantation and the American Society of Transplant Surgeons.

A review of recent advances in the assessment of bone porosity, permeability, and interstitial fluid flow

PubMed Central

Cardoso, Luis; Fritton, Susannah P.; Gailani, Gaffar; Benalla, Mohammed; Cowin, Stephen C.

2012-01-01

This contribution reviews recent research performed to assess the porosity and permeability of bone tissue with the objective of understanding interstitial fluid movement. Bone tissue mechanotransduction is considered to occur due to the passage of interstitial pore fluid adjacent to dendritic cell structures in the lacunar-canalicular porosity. The movement of interstitial fluid is also necessary for the nutrition of osteocytes. This review will focus on four topics related to improved assessment of bone interstitial fluid flow. First, the advantages and limitations of imaging technologies to visualize bone porosities and architecture at several length scales are summarized. Second, recent efforts to measure the vascular porosity and lacunar-canalicular microarchitecture are discussed. Third, studies associated with the measurement and estimation of the fluid pressure and permeability in the vascular and lacunar-canalicular domains are summarized. Fourth, the development of recent models to represent the interchange of fluids between the bone porosities is described. PMID:23174418

Generation and Long-term Maintenance of Nerve-free Hydra.

PubMed

Tran, Cassidy M; Fu, Sharon; Rowe, Trevor; Collins, Eva-Maria S

2017-07-07

The interstitial cell lineage of Hydra includes multipotent stem cells, and their derivatives: gland cells, nematocytes, germ cells, and nerve cells. The interstitial cells can be eliminated through two consecutive treatments with colchicine, a plant-derived toxin that kills dividing cells, thus erasing the potential for renewal of the differentiated cells that are derived from the interstitial stem cells. This allows for the generation of Hydra that lack nerve cells. A nerve-free polyp cannot open its mouth to feed, egest, or regulate osmotic pressure. Such animals, however, can survive and be cultured indefinitely in the laboratory if regularly force-fed and burped. The lack of nerve cells allows for studies of the role of the nervous system in regulating animal behavior and regeneration. Previously published protocols for nerve-free Hydra maintenance involve outdated techniques such as mouth-pipetting with hand-pulled micropipette tips to feed and clean the Hydra. Here, an improved protocol for maintenance of nerve-free Hydra is introduced. Fine-tipped forceps are used to force open the mouth and insert freshly killed Artemia. Following force-feeding, the body cavity of the animal is flushed with fresh medium using a syringe and hypodermic needle to remove undigested material, referred to here as "burping". This new method of force-feeding and burping nerve-free Hydra through the use of forceps and syringes eliminates the need for mouth-pipetting using hand-pulled micropipette tips. It thus makes the process safer and significantly more time efficient. To ensure that the nerve cells in the hypostome have been eliminated, immunohistochemistry using anti-tyrosine-tubulin is conducted.

Comparative physiology of mice and rats: radiometric measurement of vascular parameters in rodent tissues.

PubMed

Boswell, C Andrew; Mundo, Eduardo E; Ulufatu, Sheila; Bumbaca, Daniela; Cahaya, Hendry S; Majidy, Nicholas; Van Hoy, Marjie; Schweiger, Michelle G; Fielder, Paul J; Prabhu, Saileta; Khawli, Leslie A

2014-05-05

A solid understanding of physiology is beneficial in optimizing drug delivery and in the development of clinically predictive models of drug disposition kinetics. Although an abundance of data exists in the literature, it is often confounded by the use of various experimental methods and a lack of consensus in values from different sources. To help address this deficiency, we sought to directly compare three important vascular parameters at the tissue level using the same experimental approach in both mice and rats. Interstitial volume, vascular volume, and blood flow were radiometrically measured in selected harvested tissues of both species by extracellular marker infusion, red blood cell labeling, and rubidium chloride bolus distribution, respectively. The latter two parameters were further compared by whole-body autoradiographic imaging. An overall good interspecies agreement was observed for interstitial volume and blood flow on a weight-normalized basis in most tissues. In contrast, the measured vascular volumes of most rat tissues were higher than for mouse. Mice and rats, the two most commonly utilized rodent species in translational drug development, should not be considered as interchangeable in terms of vascular volume per gram of tissue. This will be particularly critical in biodistribution studies of drugs, as the amount of drug in the residual blood of tissues is often not negligible, especially for biologic drugs (e.g., antibodies) having long circulation half-lives. Physiologically based models of drug pharmacokinetics and/or pharmacodynamics also rely on accurate knowledge of biological parameters in tissues. For tissue parameters with poor interspecies agreement, the significance and possible drivers are discussed.

Photoacoustic physio-chemical analysis for prostate cancer diagnosis (Conference Presentation)

NASA Astrophysics Data System (ADS)

Xu, Guan; Cheng, Qian; Huang, Shengsong; Qin, Ming; Hopkins, Thomas; Lee, Chang H.; Kopelman, Raoul; Chao, Wan-yu; Keller, Evan T.; Wu, Denglong; Wang, Xueding

2017-03-01

Photoacoustic physio-chemical analysis (PAPCA) is a recently developed technology capable of simultaneously quantifying the content of molecular components and the corresponding microarchitectures in biological tissue. We have successfully quantified the diagnostic information in livers with PAPCA. In this study, we implemented PAPCA to the diagnosis of prostate cancers. 4 human prostates were scanned ex vivo. The PA signals from normal and cancerous regions in the prostates were acquired by an interstitial needle PA probe. A total of 14 interstitial measurements, including 6 within the normal regions and 8 in the cancerous regions, were acquired. The observed changes in molecular components, including lipid, collagen and hemoglobin were consistent with the findings by other research groups. The changes were quantified by PA spectral analysis (PASA) at wavelengths where strong optical absorption of the relevant molecular components was found. Statistically significant differences among the PASA parameters were observed (p=0.025 at significance of 0.05). A support vector machine model for differentiating the normal and cancerous tissue was established. With the limited number of samples, an 85% diagnostic accuracy was found. The diagnostic information in the PCPCA can be further enriched by targeted optical contrast agents visualizing the microarchitecture in PCa tissues. F3 PAA-PEG nanoparticles was employed to stain the PCa cells in a transgenic mouse model, in which the microarchitectures of normal and cancerous prostate tissues are comparable to that in human. Statistically significant differences were observed between the contrast-enhanced normal and cancerous regions (p=0.038 at a significance of 0.05).

CERA Attenuates Kidney Fibrogenesis in the db/db Mouse by Influencing the Renal Myofibroblast Generation

PubMed Central

Fischer, Christin; Deininger, Natalie; Wolf, Gunter; Loeffler, Ivonne

2018-01-01

Tubulointerstitial fibrosis (TIF) is a pivotal pathophysiological process in patients with diabetic nephropathy (DN). Multiple profibrotic factors and cell types, including transforming growth factor beta 1 (TGF-β1) and interstitial myofibroblasts, respectively, are responsible for the accumulation of extracellular matrix in the kidney. Matrix-producing myofibroblasts can originate from different sources and different mechanisms are involved in the activation process of the myofibroblasts in the fibrotic kidney. In this study, 16-week-old db/db mice, a model for type 2 DN, were treated for two weeks with continuous erythropoietin receptor activator (CERA), a synthetic erythropoietin variant with possible non-hematopoietic, tissue-protective effects. Non-diabetic and diabetic mice treated with placebo were used as controls. The effects of CERA on tubulointerstitial fibrosis (TIF) as well as on the generation of the matrix-producing myofibroblasts were evaluated by morphological, immunohistochemical, and molecular biological methods. The placebo-treated diabetic mice showed significant signs of beginning renal TIF (shown by picrosirius red staining; increased connective tissue growth factor (CTGF), fibronectin and collagen I deposition; upregulated KIM1 expression) together with an increased number of interstitial myofibroblasts (shown by different mesenchymal markers), while kidneys from diabetic mice treated with CERA revealed less TIF and fewer myofibroblasts. The mechanisms, in which CERA acts as an anti-fibrotic agent/drug, seem to be multifaceted: first, CERA inhibits the generation of matrix-producing myofibroblasts and second, CERA increases the ability for tissue repair. Many of these CERA effects can be explained by the finding that CERA inhibits the renal expression of the cytokine TGF-β1. PMID:29385703

CERA Attenuates Kidney Fibrogenesis in the db/db Mouse by Influencing the Renal Myofibroblast Generation.

PubMed

Fischer, Christin; Deininger, Natalie; Wolf, Gunter; Loeffler, Ivonne

2018-01-30

Tubulointerstitial fibrosis (TIF) is a pivotal pathophysiological process in patients with diabetic nephropathy (DN). Multiple profibrotic factors and cell types, including transforming growth factor beta 1 (TGF-β1) and interstitial myofibroblasts, respectively, are responsible for the accumulation of extracellular matrix in the kidney. Matrix-producing myofibroblasts can originate from different sources and different mechanisms are involved in the activation process of the myofibroblasts in the fibrotic kidney. In this study, 16-week-old db / db mice, a model for type 2 DN, were treated for two weeks with continuous erythropoietin receptor activator (CERA), a synthetic erythropoietin variant with possible non-hematopoietic, tissue-protective effects. Non-diabetic and diabetic mice treated with placebo were used as controls. The effects of CERA on tubulointerstitial fibrosis (TIF) as well as on the generation of the matrix-producing myofibroblasts were evaluated by morphological, immunohistochemical, and molecular biological methods. The placebo-treated diabetic mice showed significant signs of beginning renal TIF (shown by picrosirius red staining; increased connective tissue growth factor (CTGF), fibronectin and collagen I deposition; upregulated KIM1 expression) together with an increased number of interstitial myofibroblasts (shown by different mesenchymal markers), while kidneys from diabetic mice treated with CERA revealed less TIF and fewer myofibroblasts. The mechanisms, in which CERA acts as an anti-fibrotic agent/drug, seem to be multifaceted: first, CERA inhibits the generation of matrix-producing myofibroblasts and second, CERA increases the ability for tissue repair. Many of these CERA effects can be explained by the finding that CERA inhibits the renal expression of the cytokine TGF-β1.

Primary carcinoma of the distal male urethra: a case treated with lymphadenectomy and interstitial radiation therapy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ticho, B.H.; Perez-Tamayo, C.; Konnak, J.W.

1988-06-01

We report a case of primary squamous cell carcinoma of the distal male urethra with a single inguinal node metastasis. Treatment consisted of unilateral pelvic and inguinal lymphadenectomy, and a combined course of external beam and interstitial radiation therapy to the distal urethra and penis by the Henschke modification of the Paris technique.

Characterization of the Chicken Ovarian Cancer Model

DTIC Science & Technology

2005-08-01

Typical staining of tumor in an area of stromal hyperplasia . Figure 5. Adenocarcinoma, Ovary, Chicken, Her2/neu. Intense staining of tumor cells in...variable amounts of interstitial fibrovascular tissue and/or smooth muscle bundles, which accounted for their scirrhous nature grossly. 22...Figure 25). Figure 25. Peritoneum, Adenocarcinoma. Interstitial fibrovascular tissue and/or smooth muscle bundles account for the scirrhous

Severe acute interstitial lung disease in a patient with anaplastic lymphoma kinase rearrangement-positive non-small cell lung cancer treated with alectinib.

PubMed

Yamamoto, Yuzo; Okamoto, Isamu; Otsubo, Kohei; Iwama, Eiji; Hamada, Naoki; Harada, Taishi; Takayama, Koichi; Nakanishi, Yoichi

2015-10-01

Alectinib, the second generation anaplastic lymphoma kinase (ALK) inhibitor, has significant potency in patients with ALK rearrangement positive non-small cell lung cancer (NSCLC), and its toxicity is generally well tolerable. We report a patient who developed severe acute interstitial lung disease after alectinib treatment. An 86-year-old woman with stage IV lung adenocarcinoma positive for rearrangement of ALK gene was treated with alectinib. On the 215th day after initiation of alectinib administration, she was admitted to our hospital with the symptom of progressive dyspnea. Computed tomography (CT) revealed diffuse ground glass opacities and consolidations in both lungs, and analysis of bronchoalveolar lavage fluid revealed pronounced lymphocytosis. There was no evidence of infection or other specific causes of her condition, and she was therefore diagnosed with interstitial lung disease induced by alectinib. Her CT findings and respiratory condition improved after steroid pulse therapy. As far as we are aware, this is the first reported case of alectinib-induced severe interstitial lung disease (ILD). We should be aware of the possibility of such a severe adverse event and should therefore carefully monitor patients treated with this drug.

Increased Dietary Sodium Induces COX2 Expression by activating NFκB in Renal Medullary Interstitial Cells

PubMed Central

Zhao, Min; Davis, Linda S.; Blackwell, Timothy S.; Yull, Fiona; Breyer, Matthew D.; Hao, Chuan-Ming

2013-01-01

High salt diet induces renal medullary COX2 expression. Selective blockade of renal medullary COX2 activity in rats causes salt sensitive hypertension, suggesting a role for renal medullary COX2 in maintaining systemic sodium balance. The present study characterized the cellular location of COX2 induction in the kidney of mice following high salt diet and examined the role of NFκB in mediating this COX2 induction in response to increased dietary salt. High salt diet (8% NaCl) for 3 days markedly increased renal medullary COX2 expression in C57Bl/6J mice. Co-immunofluorescence using a COX2 antibody and antibodies against AQP2, ClC-K, AQP1 and CD31 showed that high salt diet-induced COX2 was selectively expressed in renal medullary interstitial cells. By using NFκB reporter transgenic mice, we observed a 7 fold increase of luciferase activity in the renal medulla of the NFκB-luciferase reporter mice following high salt diet, and a robust induction of EGFP expression mainly in renal medullary interstitial cells of the NFκB-EGFP reporter mice following high salt diet. Treating high salt diet fed C57Bl/6J mice with selective IκB kinase inhibitor IMD-0354 (8mg/kg bw) substantially suppressed COX2 induction in renal medulla, and also significantly reduced urinary PGE2. These data therefore suggest that renal medullary interstitial cell NFκB plays an important role in mediating renal medullary COX2 expression and promoting renal PGE2 synthesis in response to increased dietary sodium. PMID:23900806

Distribution of mast cell subtypes in interstitial cystitis: implications for novel diagnostic and therapeutic strategies?

PubMed

Malik, Shabana T; Birch, Brian R; Voegeli, David; Fader, Mandy; Foria, Vipul; Cooper, Alan J; Walls, Andrew F; Lwaleed, Bashir A

2018-05-15

To identify the presence and geographical distribution of mast cell (MC) subtypes: MC T (tryptase positive-chymase negative) and MC TC (tryptase positive-chymase positive) in bladder tissue. Bladder tissue was obtained from patients with painful bladder syndrome/interstitial cystitis (n=14) and normal histology from University Hospital Southampton tissue bank. Sequential tissue slices were immunohistochemically stained for MC subtypes using anti-MC tryptase (for MC T and MC TC ) and anti-MC chymase (for MC TC ). Stained sections were photographed, and positively stained MCs were quantified using ImageJ. Data were analysed using descriptive statistics and individual paired t-tests. There was a significant difference in the density of MCs between each layer of the disease bladder, with the greatest accumulation within the detrusor (p<0.001). There was a significant increase in MC TC subtype in the lamina (p=0.009) in painful bladder syndrome/interstitial cystitis. Our results suggest that mastocytosis is present within all layers of disease bladder, especially the muscle layer. The varying increase in MC subtypes in the lamina and mucosa may explain the variability in painful bladder syndrome/interstitial cystitis symptoms. A high influx of MC TC in the mucosa of individuals who also had ulceration noted within their diagnostic notes may be of the Hunner's ulcer subclassification. These findings suggest a relationship between the pathogenesis of MC subtypes and the clinical presentation of painful bladder syndrome/interstitial cystitis. A cohort study would further elucidate the diagnostic and/or therapeutic potential of MCs in patients with painful bladder syndrome/interstitial cystitis. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Resident alveolar macrophages are master regulators of arrested alveolarization in experimental bronchopulmonary dysplasia.

PubMed

Kalymbetova, Tatiana V; Selvakumar, Balachandar; Rodríguez-Castillo, José Alberto; Gunjak, Miša; Malainou, Christina; Heindl, Miriam Ruth; Moiseenko, Alena; Chao, Cho-Ming; Vadász, István; Mayer, Konstantin; Lohmeyer, Jürgen; Bellusci, Saverio; Böttcher-Friebertshäuser, Eva; Seeger, Werner; Herold, Susanne; Morty, Rory E

2018-06-01

Trophic functions for macrophages are emerging as key mediators of developmental processes, including bone, vessel, and mammary gland development. Yolk sac-derived macrophages mature in the distal lung shortly after birth. Myeloid-lineage macrophages are recruited to the lung and are activated under pathological conditions. These pathological conditions include bronchopulmonary dysplasia (BPD), a common complication of preterm birth characterized by stunted lung development, where the formation of alveoli is blocked. No study has addressed causal roles for immune cells in lung alveolarization. We employed antibody-based and transgenic death receptor-based depletion approaches to deplete or prevent lung recruitment of immune cell populations in a hyperoxia-based mouse model of BPD. Neither neutrophils nor exudate macrophages (which might include lung interstitial macrophages) contributed to structural perturbations to the lung that were provoked by hyperoxia; however, cells of the Csf1r-expressing monocyte/macrophage lineage were implicated as causal mediators of stunted lung development. We propose that resident alveolar macrophages differentiate into a population of CD45 + CD11c + SiglecF + CD11b + CD68 + MHCII + cells, which are activated by hyperoxia, and contribute to disturbances to the structural development of the immature lung. This is the first report that causally implicates immune cells in pathological disturbances to postnatal lung organogenesis. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Low-level overexpression of p53 promotes warfarin-induced calcification of porcine aortic valve interstitial cells by activating Slug gene transcription.

PubMed

Gao, Li; Ji, Yue; Lu, Yan; Qiu, Ming; Shen, Yejiao; Wang, Yaqing; Kong, Xiangqing; Shao, Yongfeng; Sheng, Yanhui; Sun, Wei

2018-03-09

The most frequently used oral anti-coagulant warfarin has been implicated in inducing calcification of aortic valve interstitial cells (AVICs), whereas the mechanism is not fully understood. The low-level activation of p53 is found to be involved in osteogenic transdifferentiation and calcification of AVICs. Whether p53 participates in warfarin-induced AVIC calcification remains unknown. In this study, we investigated the role of low-level p53 overexpression in warfarin-induced porcine AVIC (pAVIC) calcification. Immunostaining, quantitative PCR, and Western blotting revealed that p53 was expressed in human and pAVICs and that p53 expression was slightly increased in calcific human aortic valves compared with non-calcific valves. Terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling staining indicated that apoptosis slightly increased in calcific aortic valves than in non-calcific valves. Warfarin treatment led to a low-level increase of p53 mRNA and protein in both pAVICs and mouse aortic valves. Low-level overexpression of p53 in pAVICs via an adenovirus vector did not affect pAVIC apoptosis but promoted warfarin-induced calcium deposition and expression of osteogenic markers. shRNA-mediated p53 knockdown attenuated the pAVIC calcium deposition and osteogenic marker expression. Moreover, ChIP and luciferase assays showed that p53 was recruited to the slug promoter and activated slug expression in calcific pAVICs. Of note, overexpression of Slug increased osteogenic marker Runx2 expression, but not pAVIC calcium deposition, and Slug knockdown attenuated pAVIC calcification and p53-mediated pAVIC calcium deposition and expression of osteogenic markers. In conclusion, we found that p53 plays an important role in warfarin induced pAVIC calcification, and increased slug transcription by p53 is required for p53-mediated pAVIC calcification. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Intravital imaging of osteocytes in mouse calvaria using third harmonic generation microscopy

PubMed Central

Cisek, Richard; Wein, Marc N.; Turcotte, Raphaël; Haase, Christa; Yeh, Shu-Chi A.; Bharadwaj, Srinidhi; Raphael, Anthony P.; Paudel, Hari; Alt, Clemens; Liu, Tzu-Ming; Kronenberg, Henry M.; Lin, Charles P.

2017-01-01

Osteocytes are the most abundant cell in the bone, and have multiple functions including mechanosensing and regulation of bone remodeling activities. Since osteocytes are embedded in the bone matrix, their inaccessibility makes in vivo studies problematic. Therefore, a non-invasive technique with high spatial resolution is desired. The purpose of this study is to investigate the use of third harmonic generation (THG) microscopy as a noninvasive technique for high-resolution imaging of the lacunar-canalicular network (LCN) in live mice. By performing THG imaging in combination with two- and three-photon fluorescence microscopy, we show that THG signal is produced from the bone-interstitial fluid boundary of the lacuna, while the interstitial fluid-osteocyte cell boundary shows a weaker THG signal. Canaliculi are also readily visualized by THG imaging, with canaliculi oriented at small angles relative to the optical axis exhibiting stronger signal intensity compared to those oriented perpendicular to the optical axis (parallel to the image plane). By measuring forward- versus epi-detected THG signals in thinned versus thick bone samples ex vivo, we found that the epi-collected THG from the LCN of intact bone contains a superposition of backward-directed and backscattered forward-THG. As an example of a biological application, THG was used as a label-free imaging technique to study structural variations in the LCN of live mice deficient in both histone deacetylase 4 and 5 (HDAC4, HDAC5). Three-dimensional analyses were performed and revealed statistically significant differences between the HDAC4/5 double knockout and wild type mice in the number of osteocytes per volume and the number of canaliculi per lacunar surface area. These changes in osteocyte density and dendritic projections occurred without differences in lacunar size. This study demonstrates that THG microscopy imaging of the LCN in live mice enables quantitative analysis of osteocytes in animal models without the use of dyes or physical sectioning. PMID:29065178

Csf1r-mApple Transgene Expression and Ligand Binding In Vivo Reveal Dynamics of CSF1R Expression within the Mononuclear Phagocyte System.

PubMed

Hawley, Catherine A; Rojo, Rocio; Raper, Anna; Sauter, Kristin A; Lisowski, Zofia M; Grabert, Kathleen; Bain, Calum C; Davis, Gemma M; Louwe, Pieter A; Ostrowski, Michael C; Hume, David A; Pridans, Clare; Jenkins, Stephen J

2018-03-15

CSF1 is the primary growth factor controlling macrophage numbers, but whether expression of the CSF1 receptor differs between discrete populations of mononuclear phagocytes remains unclear. We have generated a Csf1r -mApple transgenic fluorescent reporter mouse that, in combination with lineage tracing, Alexa Fluor 647-labeled CSF1-Fc and CSF1, and a modified Δ Csf1- enhanced cyan fluorescent protein (ECFP) transgene that lacks a 150 bp segment of the distal promoter, we have used to dissect the differentiation and CSF1 responsiveness of mononuclear phagocyte populations in situ. Consistent with previous Csf1r- driven reporter lines, Csf1r -mApple was expressed in blood monocytes and at higher levels in tissue macrophages, and was readily detectable in whole mounts or with multiphoton microscopy. In the liver and peritoneal cavity, uptake of labeled CSF1 largely reflected transgene expression, with greater receptor activity in mature macrophages than monocytes and tissue-specific expression in conventional dendritic cells. However, CSF1 uptake also differed between subsets of monocytes and discrete populations of tissue macrophages, which in macrophages correlated with their level of dependence on CSF1 receptor signaling for survival rather than degree of transgene expression. A double Δ Csf1r -ECFP- Csf1r -mApple transgenic mouse distinguished subpopulations of microglia in the brain, and permitted imaging of interstitial macrophages distinct from alveolar macrophages, and pulmonary monocytes and conventional dendritic cells. The Csf1r- mApple mice and fluorescently labeled CSF1 will be valuable resources for the study of macrophage and CSF1 biology, which are compatible with existing EGFP-based reporter lines. Copyright © 2018 The Authors.

NK receptors, Substance P, Ano1 expression and ultrastructural features of the muscle coat in Cav-1−/− mouse ileum

PubMed Central

Cipriani, G; Serboiu, Crenguta S; Gherghiceanu, Mihaela; Simonetta Faussone-Pellegrini, Maria; Vannucchi, Maria Giuliana

2011-01-01

Abstract Caveolin (Cav)-1 is an integral membrane protein of caveolae playing a crucial role in various signal transduction pathways. Caveolae represent the sites for calcium entry and storage especially in smooth muscle cells (SMC) and interstitial cells of Cajal (ICC). Cav-1−/− mice lack caveolae and show abnormalities in pacing and contractile activity of the small intestine. Presently, we investigated, by transmission electron microscopy (TEM) and immunohistochemistry, whether the absence of Cav-1 in Cav-1−/− mouse small intestine affects ICC, SMC and neuronal morphology, the expression of NK1 and NK2 receptors, and of Ano1 (also called Dog1 or TMEM16A), an essential molecule for slow wave activity in gastrointestinal muscles. ICC were also labelled with c-Kit and tachykinergic neurons with Substance P (SP). In Cav-1−/− mice: (i) ICC were Ano1-negative but maintained c-Kit expression, (ii) NK1 and NK2 receptor immunoreactivity was more intense and, in the SMC, mainly intracytoplasmatic, (iii) SP-immunoreactivity was significantly reduced. Under TEM: (i) ICC, SMC and telocytes lacked typical caveolae but had few and large flask-shaped vesicles we called large-sized caveolae; (ii) SMC and ICC contained an extraordinary high number of mitochondria, (iii) neurons were unchanged. To maintain intestinal motility, loss of caveolae and reduced calcium availability in Cav-1–knockout mice seem to be balanced by a highly increased number of mitochondria in ICC and SMC. Loss of Ano-1 expression, decrease of SP content and consequently overexpression of NK receptors suggest that all these molecules are Cav-1–associated proteins. PMID:21535398

NK receptors, Substance P, Ano1 expression and ultrastructural features of the muscle coat in Cav-1(-/-) mouse ileum.

PubMed

Cipriani, G; Serboiu, Crenguta S; Gherghiceanu, Mihaela; Faussone-Pellegrini, Maria Simonetta; Vannucchi, Maria Giuliana

2011-11-01

Caveolin (Cav)-1 is an integral membrane protein of caveolae playing a crucial role in various signal transduction pathways. Caveolae represent the sites for calcium entry and storage especially in smooth muscle cells (SMC) and interstitial cells of Cajal (ICC). Cav-1(-/-) mice lack caveolae and show abnormalities in pacing and contractile activity of the small intestine. Presently, we investigated, by transmission electron microscopy (TEM) and immunohistochemistry, whether the absence of Cav-1 in Cav-1(-/-) mouse small intestine affects ICC, SMC and neuronal morphology, the expression of NK1 and NK2 receptors, and of Ano1 (also called Dog1 or TMEM16A), an essential molecule for slow wave activity in gastrointestinal muscles. ICC were also labelled with c-Kit and tachykinergic neurons with Substance P (SP). In Cav-1(-/-) mice: (i) ICC were Ano1-negative but maintained c-Kit expression, (ii) NK1 and NK2 receptor immunoreactivity was more intense and, in the SMC, mainly intracytoplasmatic, (iii) SP-immunoreactivity was significantly reduced. Under TEM: (i) ICC, SMC and telocytes lacked typical caveolae but had few and large flask-shaped vesicles we called large-sized caveolae; (ii) SMC and ICC contained an extraordinary high number of mitochondria, (iii) neurons were unchanged. To maintain intestinal motility, loss of caveolae and reduced calcium availability in Cav-1-knockout mice seem to be balanced by a highly increased number of mitochondria in ICC and SMC. Loss of Ano-1 expression, decrease of SP content and consequently overexpression of NK receptors suggest that all these molecules are Cav-1-associated proteins. © 2011 The Authors Journal of Cellular and Molecular Medicine © 2011 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.

Identification of Novel Rosavirus Species That Infects Diverse Rodent Species and Causes Multisystemic Dissemination in Mouse Model

PubMed Central

Fan, Rachel Y. Y.; Zhang, Anna J. X.; Chan, Brandon C. C.; Lam, Carol S. F.; Yip, Cyril C. Y.; Chan, Kwok-Hung; Chen, Zhi-Wei; Yuen, Kwok-Yung

2016-01-01

While novel picornaviruses are being discovered in rodents, their host range and pathogenicity are largely unknown. We identified two novel picornaviruses, rosavirus B from the street rat, Norway rat, and rosavirus C from five different wild rat species (chestnut spiny rat, greater bandicoot rat, Indochinese forest rat, roof rat and Coxing's white-bellied rat) in China. Analysis of 13 complete genome sequences showed that “Rosavirus B” and “Rosavirus C” represent two potentially novel picornavirus species infecting different rodents. Though being most closely related to rosavirus A, rosavirus B and C possessed distinct protease cleavage sites and variations in Yn-Xm-AUG sequence in 5’UTR and myristylation site in VP4. Anti-rosavirus B VP1 antibodies were detected in Norway rats, whereas anti-rosavirus C VP1 and neutralizing antibodies were detected in Indochinese forest rats and Coxing's white-bellied rats. While the highest prevalence was observed in Coxing's white-bellied rats by RT-PCR, the detection of rosavirus C from different rat species suggests potential interspecies transmission. Rosavirus C isolated from 3T3 cells causes multisystemic diseases in a mouse model, with high viral loads and positive viral antigen expression in organs of infected mice after oral or intracerebral inoculation. Histological examination revealed alveolar fluid exudation, interstitial infiltration, alveolar fluid exudate and wall thickening in lungs, and hepatocyte degeneration and lymphocytic/monocytic inflammatory infiltrates with giant cell formation in liver sections of sacrificed mice. Since rosavirus A2 has been detected in fecal samples of children, further studies should elucidate the pathogenicity and emergence potential of different rosaviruses. PMID:27737017

Myogenic Progenitor Cells Control Extracellular Matrix Production by Fibroblasts during Skeletal Muscle Hypertrophy.

PubMed

Fry, Christopher S; Kirby, Tyler J; Kosmac, Kate; McCarthy, John J; Peterson, Charlotte A

2017-01-05

Satellite cells, the predominant stem cell population in adult skeletal muscle, are activated in response to hypertrophic stimuli and give rise to myogenic progenitor cells (MPCs) within the extracellular matrix (ECM) that surrounds myofibers. This ECM is composed largely of collagens secreted by interstitial fibrogenic cells, which influence satellite cell activity and muscle repair during hypertrophy and aging. Here we show that MPCs interact with interstitial fibrogenic cells to ensure proper ECM deposition and optimal muscle remodeling in response to hypertrophic stimuli. MPC-dependent ECM remodeling during the first week of a growth stimulus is sufficient to ensure long-term myofiber hypertrophy. MPCs secrete exosomes containing miR-206, which represses Rrbp1, a master regulator of collagen biosynthesis, in fibrogenic cells to prevent excessive ECM deposition. These findings provide insights into how skeletal stem and progenitor cells interact with other cell types to actively regulate their extracellular environments for tissue maintenance and adaptation. Copyright © 2017 Elsevier Inc. All rights reserved.

Skeletal Adaptation to Intramedullary Pressure-Induced Interstitial Fluid Flow Is Enhanced in Mice Subjected to Targeted Osteocyte Ablation

PubMed Central

Kwon, Ronald Y.; Meays, Diana R.; Meilan, Alexander S.; Jones, Jeremiah; Miramontes, Rosa; Kardos, Natalie; Yeh, Jiunn-Chern; Frangos, John A.

2012-01-01

Interstitial fluid flow (IFF) is a potent regulatory signal in bone. During mechanical loading, IFF is generated through two distinct mechanisms that result in spatially distinct flow profiles: poroelastic interactions within the lacunar-canalicular system, and intramedullary pressurization. While the former generates IFF primarily within the lacunar-canalicular network, the latter generates significant flow at the endosteal surface as well as within the tissue. This gives rise to the intriguing possibility that loading-induced IFF may differentially activate osteocytes or surface-residing cells depending on the generating mechanism, and that sensation of IFF generated via intramedullary pressurization may be mediated by a non-osteocytic bone cell population. To begin to explore this possibility, we used the Dmp1-HBEGF inducible osteocyte ablation mouse model and a microfluidic system for modulating intramedullary pressure (ImP) to assess whether structural adaptation to ImP-driven IFF is altered by partial osteocyte depletion. Canalicular convective velocities during pressurization were estimated through the use of fluorescence recovery after photobleaching and computational modeling. Following osteocyte ablation, transgenic mice exhibited severe losses in bone structure and altered responses to hindlimb suspension in a compartment-specific manner. In pressure-loaded limbs, transgenic mice displayed similar or significantly enhanced structural adaptation to Imp-driven IFF, particularly in the trabecular compartment, despite up to ∼50% of trabecular lacunae being uninhabited following ablation. Interestingly, regression analysis revealed relative gains in bone structure in pressure-loaded limbs were correlated with reductions in bone structure in unpressurized control limbs, suggesting that adaptation to ImP-driven IFF was potentiated by increases in osteoclastic activity and/or reductions in osteoblastic activity incurred independently of pressure loading. Collectively, these studies indicate that structural adaptation to ImP-driven IFF can proceed unimpeded following a significant depletion in osteocytes, consistent with the potential existence of a non-osteocytic bone cell population that senses ImP-driven IFF independently and potentially parallel to osteocytic sensation of poroelasticity-derived IFF. PMID:22413015

Kidney pericytes: roles in regeneration and fibrosis.

PubMed

Kramann, Rafael; Humphreys, Benjamin D

2014-07-01

Renal pericytes have been neglected for many years, but recently they have become an intensively studied cell population in renal biology and pathophysiology. Pericytes are stromal cells that support vasculature, and a subset of pericytes are mesenchymal stem cells. In kidney, pericytes have been reported to play critical roles in angiogenesis, regulation of renal medullary and cortical blood flow, and serve as progenitors of interstitial myofibroblasts in renal fibrogenesis. They interact with endothelial cells through distinct signaling pathways and their activation and detachment from capillaries after acute or chronic kidney injury may be critical for driving chronic kidney disease progression. By contrast, during kidney homeostasis it is likely that pericytes serve as a local stem cell population that replenishes differentiated interstitial and vascular cells lost during aging. This review describes both the regenerative properties of pericytes as well as involvement in pathophysiologic conditions such as fibrogenesis. Copyright © 2014 Elsevier Inc. All rights reserved.

Mapping oxygen concentration in the awake mouse brain

PubMed Central

Lyons, Declan G; Parpaleix, Alexandre; Roche, Morgane; Charpak, Serge

2016-01-01

Although critical for brain function, the physiological values of cerebral oxygen concentration have remained elusive because high-resolution measurements have only been performed during anesthesia, which affects two major parameters modulating tissue oxygenation: neuronal activity and blood flow. Using measurements of capillary erythrocyte-associated transients, fluctuations of oxygen partial pressure (Po2) associated with individual erythrocytes, to infer Po2 in the nearby neuropil, we report the first non-invasive micron-scale mapping of cerebral Po2 in awake, resting mice. Interstitial Po2 has similar values in the olfactory bulb glomerular layer and the somatosensory cortex, whereas there are large capillary hematocrit and erythrocyte flux differences. Awake tissue Po2 is about half that under isoflurane anesthesia, and within the cortex, vascular and interstitial Po2 values display layer-specific differences which dramatically contrast with those recorded under anesthesia. Our findings emphasize the importance of measuring energy parameters non-invasively in physiological conditions to precisely quantify and model brain metabolism. DOI: http://dx.doi.org/10.7554/eLife.12024.001 PMID:26836304

Mapping oxygen concentration in the awake mouse brain.

PubMed

Lyons, Declan G; Parpaleix, Alexandre; Roche, Morgane; Charpak, Serge

2016-02-02

Although critical for brain function, the physiological values of cerebral oxygen concentration have remained elusive because high-resolution measurements have only been performed during anesthesia, which affects two major parameters modulating tissue oxygenation: neuronal activity and blood flow. Using measurements of capillary erythrocyte-associated transients, fluctuations of oxygen partial pressure (Po2) associated with individual erythrocytes, to infer Po2 in the nearby neuropil, we report the first non-invasive micron-scale mapping of cerebral Po2 in awake, resting mice. Interstitial Po2 has similar values in the olfactory bulb glomerular layer and the somatosensory cortex, whereas there are large capillary hematocrit and erythrocyte flux differences. Awake tissue Po2 is about half that under isoflurane anesthesia, and within the cortex, vascular and interstitial Po2 values display layer-specific differences which dramatically contrast with those recorded under anesthesia. Our findings emphasize the importance of measuring energy parameters non-invasively in physiological conditions to precisely quantify and model brain metabolism.

Characterization of lymphocyte populations in nonspecific interstitial pneumonia*

PubMed Central

Keogh, Karina A; Limper, Andrew H

2005-01-01

Study objectives Nonspecific interstitial pneumonia (NSIP) has been identified as a distinct entity with a more favorable prognosis and better response to immunosuppressive therapies than usual interstitial pneumonia (UIP). However the inflammatory profile of NSIP has not been characterized. Design Using immunohistochemistry techniques on open lung biopsy specimens, the infiltrate in NSIP was characterized in terms of T and B cells, and macrophages, and the T cell population further identified as either CD4 (helper) or CD8 (suppressor-cytotoxic) T cells. The extent of Th1 and Th2 cytokine producing cells was determined and compared to specimens from patients with UIP. Results In ten NSIP tissue samples 41.4 ± 4% of mononuclear cells expressed CD3, 24.7 ± 1.8% CD4, 19.1 ± 2% CD8, 27.4 ± 3.9% CD20, and 14.3 ± 1.6% had CD68 expression. Mononuclear cells expressed INFγ 21.9 ± 1.9% of the time and IL-4 in 3.0 ± 1%. In contrast, biopsies from eight patients with UIP demonstrated substantially less cellular staining for either cytokine (INFγ; 4.6 ± 1.7% and IL-4; 0.6 ± 0.3%). Significant populations of CD20 positive B-cells were also identified. Conclusion The lymphocytic infiltrate in NSIP is characterized by an elevated CD4/CD8 T-cell ratio, and is predominantly of Th1 type, with additional populations rich in B-cells. Such features are consistent with the favorable clinical course observed in patients with NSIP compared to UIP. PMID:16287509

Pulsed and oscillating gradient MRI for assessment of cell size and Extracellular space (POMACE) in mouse gliomas

PubMed Central

Reynaud, Olivier; Winters, Kerryanne Veronica; Hoang, Dung Minh; Wadghiri, Youssef Zaim; Novikov, Dmitry S.; Kim, Sungheon Gene

2016-01-01

Solid tumor microstructure is related to aggressiveness of tumor, interstitial pressure and drug delivery pathways that are closely associated with treatment response, metastatic spread and prognosis. In this study, we introduce a novel diffusion MRI data analysis framework, Pulsed and Oscillating gradient MRI for Assessment of Cell size and Extracellular space (POMACE), and demonstrate its feasibility in a mouse tumor model. In vivo and ex vivo POMACE experiments were performed on mice bearing the GL261 murine glioma model (n=8). Since the complete diffusion time-dependence is in general non-analytical, the tumor microstructure was modeled in an appropriate time/frequency regime by impermeable spheres (radius Rcell, intracellular diffusivity Dics) surrounded by extracellular space (approximated by constant apparent diffusivity Decs in volume fraction ECS). POMACE parametric maps (ECS, Rcell, Dics, Decs) were compared with conventional diffusion weighted imaging metrics, electron microscopy (EM), alternative ECS determination based on effective medium theory (EMT), and optical microscopy performed on the same samples. It was shown that Decs can be approximated by its long-time tortuosity limit in the range [1/(88 Hz) - 31 ms]. ECS estimations (44±7% in vivo and 54±11% ex vivo) were in agreement with EMT-based ECS and literature on brain gliomas. Ex vivo, ECS maps correlated well with optical microscopy. Cell sizes (Rcell=4.8±1.3 in vivo and 4.3±1.4 μm ex vivo) were consistent with EM measurements (4.7±1.8 μm). In conclusion, Rcell and ECS can be quantified and mapped in vivo and ex vivo in brain tumors using the proposed POMACE method. Our experimental results support that POMACE provides a way to interpret the frequency- or time-dependence of the diffusion coefficient in tumors in terms of objective biophysical parameters of neuronal tissue, which can be used for non-invasive monitoring of preclinical cancer studies and treatment efficacy. PMID:27448059

Interstitial Photodynamic Therapy in Treating Patients With Recurrent Head and Neck Cancer

ClinicalTrials.gov

2017-09-11

Recurrent Laryngeal Squamous Cell Carcinoma; Recurrent Laryngeal Verrucous Carcinoma; Recurrent Lip and Oral Cavity Squamous Cell Carcinoma; Recurrent Metastatic Squamous Cell Carcinoma in the Neck With Occult Primary; Recurrent Oral Cavity Verrucous Carcinoma; Recurrent Oropharyngeal Squamous Cell Carcinoma; Tongue Carcinoma

GDF11 induces kidney fibrosis, renal cell epithelial-to-mesenchymal transition, and kidney dysfunction and failure.

PubMed

Pons, Marianne; Koniaris, Leonidas G; Moe, Sharon M; Gutierrez, Juan C; Esquela-Kerscher, Aurora; Zimmers, Teresa A

2018-05-03

GDF11 modulates embryonic patterning and kidney organogenesis. Herein, we sought to define GDF11 function in the adult kidney and in renal diseases. In vitro renal cell lines, genetic, and murine in vivo renal injury models were examined. Among tissues tested, Gdf11 was highest in normal adult mouse kidney. Expression was increased acutely after 5/6 nephrectomy, ischemia-reperfusion injury, kanamycin toxicity, or unilateral ureteric obstruction. Systemic, high-dose GDF11 administration in adult mice led to renal failure, with accompanying kidney atrophy, interstitial fibrosis, epithelial-to-mesenchymal transition of renal tubular cells, and eventually death. These effects were associated with phosphorylation of SMAD2 and could be blocked by follistatin. In contrast, Gdf11 heterozygous mice showed reduced renal Gdf11 expression, renal fibrosis, and expression of fibrosis-associated genes both at baseline and after unilateral ureteric obstruction compared with wild-type littermates. The kidney-specific consequences of GDF11 dose modulation are direct effects on kidney cells. GDF11 induced proliferation and activation of NRK49f renal fibroblasts and also promoted epithelial-to-mesenchymal transition of IMCD-3 tubular epithelial cells in a SMAD3-dependent manner. Taken together, these data suggest that GDF11 and its downstream signals are critical in vivo mediators of renal injury. These effects are through direct actions of GDF11 on renal tubular cells and fibroblasts. Thus, regulation of GDF11 presents a therapeutic target for diseases involving renal fibrosis and impaired tubular function. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Increased dietary sodium induces COX2 expression by activating NFκB in renal medullary interstitial cells.

PubMed

He, Wenjuan; Zhang, Min; Zhao, Min; Davis, Linda S; Blackwell, Timothy S; Yull, Fiona; Breyer, Matthew D; Hao, Chuan-Ming

2014-02-01

High salt diet induces renal medullary cyclooxygenase 2 (COX2) expression. Selective blockade of renal medullary COX2 activity in rats causes salt-sensitive hypertension, suggesting a role for renal medullary COX2 in maintaining systemic sodium balance. The present study characterized the cellular location of COX2 induction in the kidney of mice following high salt diet and examined the role of NFκB in mediating this COX2 induction in response to increased dietary salt. High salt diet (8 % NaCl) for 3 days markedly increased renal medullary COX2 expression in C57Bl/6 J mice. Co-immunofluorescence using a COX2 antibody and antibodies against aquaporin-2, ClC-K, aquaporin-1, and CD31 showed that high salt diet-induced COX2 was selectively expressed in renal medullary interstitial cells. By using NFκB reporter transgenic mice, we observed a sevenfold increase of luciferase activity in the renal medulla of the NFκB-luciferase reporter mice following high salt diet, and a robust induction of enhanced green fluorescent protein (EGFP) expression mainly in renal medullary interstitial cells of the NFκB-EGFP reporter mice following high salt diet. Treating high salt diet-fed C57Bl/6 J mice with selective IκB kinase inhibitor IMD-0354 (8 mg/kg bw) substantially suppressed COX2 induction in renal medulla, and also significantly reduced urinary prostaglandin E2 (PGE2). These data therefore suggest that renal medullary interstitial cell NFκB plays an important role in mediating renal medullary COX2 expression and promoting renal PGE2 synthesis in response to increased dietary sodium.

Biological and mechanical evaluation of a Bio-Hybrid scaffold for autologous valve tissue engineering.

PubMed

Jahnavi, S; Saravanan, U; Arthi, N; Bhuvaneshwar, G S; Kumary, T V; Rajan, S; Verma, R S

2017-04-01

Major challenge in heart valve tissue engineering for paediatric patients is the development of an autologous valve with regenerative capacity. Hybrid tissue engineering approach is recently gaining popularity to design scaffolds with desired biological and mechanical properties that can remodel post implantation. In this study, we fabricated aligned nanofibrous Bio-Hybrid scaffold made of decellularized bovine pericardium: polycaprolactone-chitosan with optimized polymer thickness to yield the desired biological and mechanical properties. CD44 + , αSMA + , Vimentin + and CD105 - human valve interstitial cells were isolated and seeded on these Bio-Hybrid scaffolds. Subsequent biological evaluation revealed interstitial cell proliferation with dense extra cellular matrix deposition that indicated the viability for growth and proliferation of seeded cells on the scaffolds. Uniaxial mechanical tests along axial direction showed that the Bio-Hybrid scaffolds has at least 20 times the strength of the native valves and its stiffness is nearly 3 times more than that of native valves. Biaxial and uniaxial mechanical studies on valve interstitial cells cultured Bio-Hybrid scaffolds revealed that the response along the axial and circumferential direction was different, similar to native valves. Overall, our findings suggest that Bio-Hybrid scaffold is a promising material for future development of regenerative heart valve constructs in children. Copyright © 2016 Elsevier B.V. All rights reserved.

IFPA Award in Placentology lecture: molecular regulation of human trophoblast invasion.

PubMed

Knöfler, M; Pollheimer, J

2012-02-01

Invasion of extravillous trophoblast cell types into maternal uterine tissues is essential for successful human placental development and progression of pregnancy. Whereas endovascular trophoblasts migrate into the maternal spiral arteries, interstitial trophoblasts invade the decidual stroma, colonize the vessels from outside and communicate with diverse uterine cell types such as decidual stromal cells, macrophages and uterine NK cells. For example, interstitial trophoblasts expressing polymorphic human leukocyte antigen-C interact with uterine NK cells through binding to their killer immunoglobulin-like receptors which likely plays a role in trophoblast invasion and reproductive success of pregnancy. Both extravillous trophoblast subtypes are critically involved in the vascular transformation of the spiral arteries into dilated conduits ensuring appropriate blood flow into the intervillous space. Failures in this remodeling process are thought to be associated with severe forms of fetal growth restriction, preeclampsia and other pregnancy complications warranting studies on the molecular regulation of extravillous trophoblast differentiation. Moreover, interstitial trophoblast-derived hormones may regulate diverse biological functions in the decidua. In particular, human chorionic gonadotrophin has been shown to promote angiogenesis and to suppress apoptosis of endometrial stromal cells. In return, decidual cells produce a plethora of soluble factors controlling trophoblast invasion in a time- and distance-dependent manner. However, the underlying mechanisms have not been fully elucidated. Here, we will summarize autocrine as well as paracrine factors regulating invasion of extravillous trophoblasts and discuss critical signaling cascades involved. In addition, we will focus on key regulatory transcription factors controlling cell column proliferation and differentiation of the human extravillous trophoblast. Copyright © 2012 Elsevier Ltd. All rights reserved.

Targeting of tumor endothelium by RGD-grafted PLGA-nanoparticles.

PubMed

Danhier, Fabienne; Pourcelle, Vincent; Marchand-Brynaert, Jacqueline; Jérôme, Christine; Feron, Olivier; Préat, Véronique

2012-01-01

The destruction of the neovessels in solid tumors can cause the death of tumor cells resulting from the lack of oxygen and nutrients. Peculiarities of the tumor vasculature, however, also position angiogenic endothelial cells as obvious targets to address cytotoxic drugs into the tumor. In particular, the identification of a three-amino acids sequence, arginine-glycine-aspartate (RGD), as a fundamental recognition site for proliferating endothelial attachment to the extracellular matrix leads to the development of tumor-targeting ligands for nanoparticles. The RGD peptide can target the α(v)β(3) integrin overexpressed by the tumor endothelium, and thereby increases the accumulation of drug-loaded RGD-grafted nanoparticles. RGD-nanoparticles may thus extravasate more efficiently and enter the tumor via the enhanced permeability and retention (EPR) effect. This combination of active and passive processes leads to the penetration of nanoparticles into the tumor tissue, followed by cellular uptake and intracellular delivery of the cytotoxic payload. Since cancer cells may also express α(v)β(3) integrin, the entrapping of RGD-nanoparticles into the tumor interstitial fluid may yet be facilitated through direct binding to cancer cells. Here, we describe methods used for the preparation of RGD-nanoparticles and for the validation of their potential of tumor endothelium targeting both in vitro and in vivo. We also illustrate how RGD-nanoparticles may be more suited than nontargeted modalities for the tumor delivery of poorly soluble and/or highly cytotoxic drugs, using different mouse tumor xenograft models. Copyright © 2012 Elsevier Inc. All rights reserved.

The effect of interstitial pressure on tumor growth: coupling with the blood and lymphatic vascular systems

PubMed Central

Wu, Min; Frieboes, Hermann B.; McDougall, Steven R.; Chaplain, Mark A.J.; Cristini, Vittorio; Lowengrub, John

2013-01-01

The flow of interstitial fluid and the associated interstitial fluid pressure (IFP) in solid tumors and surrounding host tissues have been identified as critical elements in cancer growth and vascularization. Both experimental and theoretical studies have shown that tumors may present elevated IFP, which can be a formidable physical barrier for delivery of cell nutrients and small molecules into the tumor. Elevated IFP may also exacerbate gradients of biochemical signals such as angiogenic factors released by tumors into the surrounding tissues. These studies have helped to understand both biochemical signaling and treatment prognosis. Building upon previous work, here we develop a vascular tumor growth model by coupling a continuous growth model with a discrete angiogenesis model. We include fluid/oxygen extravasation as well as a continuous lymphatic field, and study the micro-environmental fluid dynamics and their effect on tumor growth by accounting for blood flow, transcapillary fluid flux, interstitial fluid flow, and lymphatic drainage. We thus elucidate further the non-trivial relationship between the key elements contributing to the effects of interstitial pressure in solid tumors. In particular, we study the effect of IFP on oxygen extravasation and show that small blood/lymphatic vessel resistance and collapse may contribute to lower transcapillary fluid/oxygen flux, thus decreasing the rate of tumor growth. We also investigate the effect of tumor vascular pathologies, including elevated vascular and interstitial hydraulic conductivities inside the tumor as well as diminished osmotic pressure differences, on the fluid flow across the tumor capillary bed, the lymphatic drainage, and the IFP. Our results reveal that elevated interstitial hydraulic conductivity together with poor lymphatic function is the root cause of the development of plateau profiles of the IFP in the tumor, which have been observed in experiments, and contributes to a more uniform distribution of oxygen, solid tumor pressure and a broad-based collapse of the tumor lymphatics. We also find that the rate that IFF is fluxed into the lymphatics and host tissue is largely controlled by an elevated vascular hydraulic conductivity in the tumor. We discuss the implications of these results on microenvironmental transport barriers, and the tumor invasive and metastatic potential. Our results suggest the possibility of developing strategies of targeting tumor cells based on the cues in the interstitial fluid. PMID:23220211

Ultrastructure of the genital organs in interstitial polychaetes. III. Penes and ejaculatory ducts in Hesionides arenaria (Hesionidae)

NASA Astrophysics Data System (ADS)

Westheide, W.

1982-12-01

The ejaculatory ducts of the two paired copulatory organs in the interstitial polychaete Hesionides arenaria are ciliated tubes, which open into simple, partly groove-like, non-stiffened penis papillae. The larger part of the ducts within the dorsal body wall is surrounded by circular muscle cells. Voluminous gland cell bodies lie between the pharynx-gut system and the body wall in the anterior part of the body; they extend anteriorly like long, thin necks, of which severl are always united in prominent strands. Their distal ends are expanded and penetrate the ducts. Six different types of glands can be distinguished according to the ultrastructure of their secretory granules. They produce the sheath of the double spermatophore or probably contain lytic enzymes that provide for the penetration of sperm into the body of the female. Differences in ultrastructure of the male organs in the interstitial genera Hesionides and Microphthalmus do not support the recent erection of the subfamily “Microphthalminae”.

Roles of interstitial fluid pH in diabetes mellitus: Glycolysis and mitochondrial function

PubMed Central

Marunaka, Yoshinori

2015-01-01

The pH of body fluids is one the most important key factors regulating various cell function such as enzyme activity and protein-protein interaction via modification of its binding affinity. Therefore, to keep cell function normal, the pH of body fluids is maintained constant by various systems. Insulin resistance is one of the most important, serious factors making the body condition worse in diabetes mellitus. I have recently found that the pH of body (interstitial) fluids is lower in diabetes mellitus than that in non-diabetic control, and that the lowered pH is one of the causes producing insulin resistance. In this review article, I introduce importance of body (interstitial) fluid pH in regulation of body function, evidence on abnormal regulation of body fluid pH in diabetes mellitus, and relationship between the body fluid pH and insulin resistance. Further, this review proposes perspective therapies on the basis of regulation of body fluid pH including propolis (honeybee product) diet. PMID:25685283

Decoy receptor 3 inhibits renal mononuclear leukocyte infiltration and apoptosis and prevents progression of IgA nephropathy in mice.

PubMed

Ka, Shuk-Man; Hsieh, Tai-Tzu; Lin, Shih-Hua; Yang, Sung-Sen; Wu, Chin-Chen; Sytwu, Huey-Kang; Chen, Ann

2011-12-01

The progression of IgA nephropathy (IgAN), the most frequent type of primary glomerulonephritis, is associated with high levels of mononuclear leukocyte infiltration into the kidney. These cells consist mainly of T cells and macrophages. Our previous study showed that a decoy receptor 3 (DCR3) gene therapy can prevent the development of a mouse autoimmune glomerulonephritis model by its potent immune modulating effects (Ka SM, Sytwu HK, Chang DM, Hsieh SL, Tsai PY, Chen A. J Am Soc Nephrol 18: 2473-2485, 2007). Here, we tested the hypothesis that DCR3 might prevent the progression of IgAN, an immune complex-mediated primary glomerulonephritis, by inhibiting T cell activation, renal T cell/macrophage infiltration, and protecting the kidney from apoptosis. We used a progressive IgAN (Prg-IgAN) model in B cell-deficient mice, because the mice are characterized by a dramatic proliferation of activated T cells systemically and progressive NF-κB activation in the kidney. We treated the animals with short-term gene therapy with DCR3 plasmids by hydrodynamics-based gene delivery. When the mice were euthanized on day 21, we found that, compared with empty vector-treated (disease control) Prg-IgAN mice, DCR3 gene therapy resulted in 1) systemic inhibition of T cell activation and proliferation; 2) lower serum levels of proinflammatory cytokines; 3) improved proteinuria, renal function, and renal pathology (inhibiting the development of marked glomerular proliferation, crescent formation, glomerulosclerosis, and interstitial inflammation); 5) suppression of T cell and macrophage infiltration into the periglomerular interstitium of the kidney; and 5) a reduction in apoptotic figures in the kidney. On the basis of these findings, DCR3 might be useful therapeutically in preventing the progression of IgAN.

The interstitial stem cells in Hydractinia and their role in regeneration.

PubMed

Gahan, James M; Bradshaw, Brian; Flici, Hakima; Frank, Uri

2016-10-01

Hydractinia species have been animal models in developmental biology and comparative immunology for over a century, but are having a renaissance due to the establishment of modern genetic and genomic tools by the growing community of researchers utilizing them. Hydractinia has a predictable and accessible life cycle and its stem cell system, known as interstitial- or i-cells has been a paradigm for animal stem cells since the late 1800s. In adult Hydractinia, i-cells continuously provide progenitors to sustain clonal growth, tissue homeostasis, sexual reproduction and regeneration. We review recent developments in stem cell and regeneration research centered on this animal. Hydractinia joins an established team of cnidarian genetic models in times of rapid progress in these disciplines. While each animal is particularly suited to specific experimental settings, jointly they can provide an integrative insight into the diversity of animal stem cell systems, how they drive regeneration, and how they evolved. Copyright © 2016 Elsevier Ltd. All rights reserved.

Viruses and interstitial cystitis: adenovirus genomes cannot be demonstrated in urinary bladder biopsies.

PubMed

Hukkanen, V; Haarala, M; Nurmi, M; Klemi, P; Kiilholma, P

1996-01-01

Microbes may be involved in the pathogenesis of interstitial cystitis (IC). Adenoviruses and BK virus (BKV) can infect epithelial cells in urinary bladder and they are causative agents for hemorrhagic cystitis. We therefore studied the presence of adenovirus and BKV genomes in urinary bladder tissue specimens of patients with IC using polymerase chain reaction (PCR) and in situ hybridization (ISH). Controls were specimens from cases with transitional cell carcinoma of the bladder. Nucleic acids were extracted from paraffin sections of the bladder tissue for PCR. Primers detecting all adenovirus types were used. In situ hybridization was carried out for the paraffin sections using digoxigenin-labeled DNA probes for adenovirus and BKV. The adenovirus DNA PCR was able to detect one to two infected cells/specimen. All the seven IC cases studied and six controls were negative for adenovirus DNA by PCR and ISH. The ISH test for BKV genomes was also considered negative in IC cases and controls. The specimens which were negative in PCR tests yielded a signal with beta-globin primers, thus being amplifiable. We conclude that adenovirus and BKV do not play a major pathogenetic role in interstitial cystitis.

Changes in membrane cholesterol affect caveolin-1 localization and ICC-pacing in mouse jejunum.

PubMed

Daniel, E E; Bodie, Gregory; Mannarino, Marco; Boddy, Geoffrey; Cho, Woo-Jung

2004-07-01

Pacing of mouse is dependent on the spontaneous activity of interstitial cells of Cajal in the myenteric plexus (ICC-MP). These ICC, as well as intestinal smooth muscle, contain small membrane invaginations called caveolae. Caveolae are signaling centers formed by insertions of caveolin proteins in the inner aspect of the plasma membrane. Caveolins bind signaling proteins and thereby negatively modulate their signaling. We disrupted caveolae by treating intestinal segments with methyl beta-clodextrin (CD) to remove cholesterol or with water-soluble cholesterol (WSC) to load cholesterol. Both of these treatments reduced pacing frequencies, and these effects were reversed by the other agent. These treatments also inhibited paced contractions, but complete reversal was not observed. To evaluate the specificity of the effects of CD and WSC, additional studies were made of their effects on responses to carbamoyl choline and to stimulation of cholinergic nerves. Neither of these treatments affected these sets of responses compared with their respective time controls. Immunochemical and ultrastructural studies showed that caveolin 1 was present in smooth muscle membranes and ICC-MP. CD depleted both caveolin 1 and caveolae, whereas WSC increased the amount of caveolin 1 immunoreactivity and altered its distribution but failed to increase the number of caveolae. The effects of each agent were reversed in major part by the other. We conclude that signaling through caveolae may play a role in pacing by ICC but does not affect responses to acetylcholine from nerves or when added exogenously.

Expression of hepatitis B virus 1.3-fold genome plasmid in an SV40 T-antigen-immortalized mouse hepatic cell line

PubMed Central

Song, Xiu-Guang; Bian, Peng-Fei; Yu, Shu-Li; Zhao, Xiu-Hua; Xu, Wei; Bu, Xue-Hui; Li, Xia; Ma, Li-Xian

2013-01-01

AIM: To investigate the expression of the hepatitis B virus (HBV) 1.3-fold genome plasmid (pHBV1.3) in an immortalized mouse hepatic cell line induced by SV40 T-antigen (SV40T) expression. METHODS: Mouse hepatic cells were isolated from mouse liver tissue fragments from 3-5 d old Kunming mice by the direct collagenase digestion method and cultured in vitro. The pRSV-T plasmid was transfected into mouse hepatic cells to establish an SV40LT-immortalized mouse hepatic cell line. The SV40LT-immortalized mouse hepatic cells were identified and transfected with the pHBV1.3 plasmid. The levels of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in the supernatant were determined by an electrochemiluminescence immunoassay at 24, 48, 72 and 96 h after transfection. The expressions of HBsAg and hepatitis B c antigen (HBcAg) in the cells were investigated by indirect immunofluorescence analysis. The presence of HBV DNA replication intermediates in the transfected cells and viral particles in the supernatant of the transfected cell cultures was monitored using the Southern hybridization assay and transmission electronic microscopy, respectively. RESULTS: The pRSV-T plasmid was used to immortalize mouse hepatocytes and an SV40LT-immortalized mouse hepatic cell line was successfully established. SV40LT-immortalized mouse hepatic cells have the same morphology and growth characteristics as primary mouse hepatic cells can be subcultured and produce albumin and cytokeratin-18 in vitro. Immortalized mouse hepatic cells did not show the characteristics of tumor cells, as alpha-fetoprotein levels were comparable (0.58 ± 0.37 vs 0.61 ± 0.31, P = 0.37). SV40LT-immortalized mouse hepatic cells were then transfected with the pHBV1.3 plasmid, and it was found that the HBV genome replicated in SV40LT-immortalized mouse hepatic cells. The levels of HBsAg and HBeAg continuously increased in the supernatant after the transfection of pHBV1.3, and began to decrease 72 h after transfection. The expressions of HBsAg and HBcAg were observed in the pHBV1.3-transfected cells. HBV DNA replication intermediates were also observed at 72 h after transfection, including relaxed circular DNA, double-stranded DNA and single-stranded DNA. Furthermore, a few 42 nm Dane particles, as well as many 22 nm subviral particles with a spherical or filamentous shape, were detected in the supernatant. CONCLUSION: SV40T expression can immortalize mouse hepatic cells, and the pHBV1.3-transfected SV40T-immortalized mouse hepatic cell line can be a new in vitro cell model. PMID:24307795

Expression of hepatitis B virus 1.3-fold genome plasmid in an SV40 T-antigen-immortalized mouse hepatic cell line.

PubMed

Song, Xiu-Guang; Bian, Peng-Fei; Yu, Shu-Li; Zhao, Xiu-Hua; Xu, Wei; Bu, Xue-Hui; Li, Xia; Ma, Li-Xian

2013-11-28

To investigate the expression of the hepatitis B virus (HBV) 1.3-fold genome plasmid (pHBV1.3) in an immortalized mouse hepatic cell line induced by SV40 T-antigen (SV40T) expression. Mouse hepatic cells were isolated from mouse liver tissue fragments from 3-5 d old Kunming mice by the direct collagenase digestion method and cultured in vitro. The pRSV-T plasmid was transfected into mouse hepatic cells to establish an SV40LT-immortalized mouse hepatic cell line. The SV40LT-immortalized mouse hepatic cells were identified and transfected with the pHBV1.3 plasmid. The levels of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in the supernatant were determined by an electrochemiluminescence immunoassay at 24, 48, 72 and 96 h after transfection. The expressions of HBsAg and hepatitis B c antigen (HBcAg) in the cells were investigated by indirect immunofluorescence analysis. The presence of HBV DNA replication intermediates in the transfected cells and viral particles in the supernatant of the transfected cell cultures was monitored using the Southern hybridization assay and transmission electronic microscopy, respectively. The pRSV-T plasmid was used to immortalize mouse hepatocytes and an SV40LT-immortalized mouse hepatic cell line was successfully established. SV40LT-immortalized mouse hepatic cells have the same morphology and growth characteristics as primary mouse hepatic cells can be subcultured and produce albumin and cytokeratin-18 in vitro. Immortalized mouse hepatic cells did not show the characteristics of tumor cells, as alpha-fetoprotein levels were comparable (0.58 ± 0.37 vs 0.61 ± 0.31, P = 0.37). SV40LT-immortalized mouse hepatic cells were then transfected with the pHBV1.3 plasmid, and it was found that the HBV genome replicated in SV40LT-immortalized mouse hepatic cells. The levels of HBsAg and HBeAg continuously increased in the supernatant after the transfection of pHBV1.3, and began to decrease 72 h after transfection. The expressions of HBsAg and HBcAg were observed in the pHBV1.3-transfected cells. HBV DNA replication intermediates were also observed at 72 h after transfection, including relaxed circular DNA, double-stranded DNA and single-stranded DNA. Furthermore, a few 42 nm Dane particles, as well as many 22 nm subviral particles with a spherical or filamentous shape, were detected in the supernatant. SV40T expression can immortalize mouse hepatic cells, and the pHBV1.3-transfected SV40T-immortalized mouse hepatic cell line can be a new in vitro cell model.

Renal Failure in Mice with Gsα Deletion in Juxtaglomerular Cells

PubMed Central

Chen, Limeng; Faulhaber-Walter, Robert; Wen, Yubing; Huang, Yuning; Mizel, Diane; Chen, Min; Sequeira Lopez, Maria Luisa; Weinstein, Lee S.; Gomez, R. Ariel; Briggs, Josephine P.; Schnermann, Jurgen

2010-01-01

Background Mice with deletion of Gsα in renin-producing cells (RC/FF mice) have been shown to have greatly reduced renin production and lack of responsiveness of renin secretion to acute stimuli. In addition, young RC/FF mice are hypotensive and have a vasopressin-resistant concentrating defect. In the present study we have determined the long-term effect on renal function, blood pressure, and renal pathology in this low renin and diuretic mouse model. Methods and Results Urine osmolarity of RC/FF mice was decreased in all age groups. GFR measured at 7, 14 and 20 weeks of age declined progressively. Single nephron GFR similarly declined while fractional proximal fluid absorption was maintained. Expression levels of extracellular matrix proteins (collagen I, IV and fibronectin) and α-smooth muscle actin were increased in kidneys of RC/FF mice at 20 weeks, and this was accompanied by focal segmental glomerulosclerosis and periglomerular interstitial fibrosis. RC/FF mice showed a progressive reduction of body weight, an increase in urine albumin excretion, and an increase of blood pressure with aging. Conclusion A chronic reduction of renin production in mice may be a risk factor in its own right, and does not protect renal function against the profibrotic influence of a chronically elevated urine flow. PMID:20551626

Role of medullary progenitor cells in epithelial cell migration and proliferation

PubMed Central

Chen, Dong; Chen, Zhiyong; Zhang, Yuning; Park, Chanyoung; Al-Omari, Ahmed

2014-01-01

This study is aimed at characterizing medullary interstitial progenitor cells and to examine their capacity to induce tubular epithelial cell migration and proliferation. We have isolated a progenitor cell side population from a primary medullary interstitial cell line. We show that the medullary progenitor cells (MPCs) express CD24, CD44, CXCR7, CXCR4, nestin, and PAX7. MPCs are CD34 negative, which indicates that they are not bone marrow-derived stem cells. MPCs survive >50 passages, and when grown in epithelial differentiation medium develop phenotypic characteristics of epithelial cells. Inner medulla collecting duct (IMCD3) cells treated with conditioned medium from MPCs show significantly accelerated cell proliferation and migration. Conditioned medium from PGE2-treated MPCs induce tubule formation in IMCD3 cells grown in 3D Matrigel. Moreover, most of the MPCs express the pericyte marker PDGFR-b. Our study shows that the medullary interstitium harbors a side population of progenitor cells that can differentiate to epithelial cells and can stimulate tubular epithelial cell migration and proliferation. The findings of this study suggest that medullary pericyte/progenitor cells may play a critical role in collecting duct cell injury repair. PMID:24808539

Nitric oxide decreases the excitability of interstitial cells of Cajal through activation of the BK channel

PubMed Central

Zhu, Yaohui; Huizinga, Jan D

2008-01-01

Abstract Nitrergic nerves are structurally and functionally associated with ICC. To further understand mechanisms of communication, the hypothesis was investigated that NO might affect large conductance K channels. To that end, we searched for IbTX-sensitive currents in ICC obtained through explant cultures from the mouse small intestine and studied effects of the NOS inhibitor omega N-nitro-L-arginine (LNNA) and the NO donor sodium nitroprusside (SNP). IbTX-sensitive currents acquired in the whole-cell configuration through nystatin perforated patches exhibited high noise levels but relatively low amplitude, whereas currents obtained in the conventional whole-cell configuration exhibited less noise and higher amplitudes; depolarization from −80 to + 40 mV evoked 357 ± 159 pA current in the nystatin perforated patch configuration and 1075 ± 597 pA using the conventional whole-cell configuration. Immunohistochemistry showed that ICC associated with ganglia and Auerbach's plexus nerve fibers were immunoreactive to BK antibodies. The IbTX-sensitive currents were increased by SNP and inhibited by LNNA. BK blockers suppressed spontaneous transit outward currents in ICC. After block of BK currents, or before these currents became prominent, calcium currents were activated by depolarization in the same cells. Their peak amplitude occurred at −25 mV and the currents were increased with increasing extracellular calcium and inhibited by cobalt. The hypothesis is warranted that nitrergic innervation inhibits ICC excitability in part through activation of BK channels. In addition, NO is an intracellular regulator of ICC excitability. PMID:18194464

Alterations in biosynthetic accumulation of collagen types I and III during growth and morphogenesis of embryonic mouse salivary glands

NASA Technical Reports Server (NTRS)

Hardman, P.; Spooner, B. S.

1992-01-01

We examined the biosynthetic patterns of interstitial collagens in mouse embryonic submandibular and sublingual glands cultured in vitro. Rudiments explanted on day 13 of gestation and cultured for 24, 48, and 72 h all synthesized collagen types I, III, and V. However, while the total incorporation of label into collagenous proteins did not change over the three-day culture period, the rate of accumulation of newly synthesized types I and III did change. At 24 h, the ratio of newly synthesized collagen types I:III was approximately 2, whereas at 72 h, the ratio was approximately 5. These data suggest that collagen types I and III may be important in initiation of branching in this organ, but that type I may become dominant in the later stages of development and in maintenance of the adult organ.

The human uterotubal junction: a scanning electron microscope study during different phases of the menstrual cycle.

PubMed

Fadel, H E; Berns, D; Zaneveld, L J; Wilbanks, G D; Brueschke, E E

1976-10-01

Uterotubal junctions from surgically extirpated human uteri were examined. The specimens were obtained during different phases of the menstrual cycle. The interstitial portions of the tubes together with the cornual areas were dissected, excised, and their luminal surfaces exposed. The specimens were then processed for scanning electron microscopy. The surface epithelium of both the cornual endometrium and interstitial endosalpins. Ciliated cells were more numerous in the endosalpinx. Cyclic changes in ciliated cells were minimal, while cyclic secretory activity was demonstrated, especially in the endometrium. The transitional area between the endometrium and the endosalpinx was characterized by a marked increase in the number of ciliated cells, and a tendency of the secretory cells to assume a flattened, polygonal shape. These morphologic features suggest a possible role in the transport and/or maintenance of spermatozoa and/or ova.

Immunofluorescence reveals unusual patterns of labelling for connexin43 localized to calbindin-D28K-positive interstitial cells in the pineal gland.

PubMed

Tsao, D D; Wang, S G; Lynn, B D; Nagy, J I

2017-06-01

Gap junctions between cells in the pineal gland have been described ultrastructurally, but their connexin constituents have not been fully characterized. We used immunofluorescence in combination with markers of pineal cells to document the cellular localization of connexin43 (Cx43). Immunofluorescence labelling of Cx43 with several different antibodies was widely distributed throughout the pineal, whereas another connexin examined, connexin26, was not found in pineal but only in surrounding leptomeninges. Labelling apparently associated with plasma membranes was visualized either as fine Cx43-puncta (1-2 μm) or as unusually large pools of Cx43 ranging up to 4-7 μm in diameter or length. These puncta and pools were highly concentrated in perivascular spaces, where they were associated with numerous cells devoid of labelling for markers of pinealocytes (e.g. tryptophan hydroxylase and serotonin), and where they were minimally associated with blood vessels and lacked association with resident macrophages. Astrocytes labelled for glial fibrillary acidic protein were largely restricted to the anterior pole of the pineal gland, where they displayed only fine and sparse Cx43-puncta along their processes. Labelling for Cx43 was localized largely though not exclusively to the somata and long processes of a subpopulation of perivascular interstitial cells that were immunopositive for calbindin-D28K. These cells were often located among dense bundles or termination areas of sympathetic fibres labelled for tyrosine hydroxylase or serotonin. The results indicate that interstitial cells form abundant gap junctions composed of Cx43, and suggest that gap junction-mediated intracellular communication by these cells supports the activities of pinealocytes. © 2017 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

Inflammatory myofibroblastic tumors of the lung carrying a chimeric A2M-ALK gene: report of 2 infantile cases and review of the differential diagnosis of infantile pulmonary lesions.

PubMed

Tanaka, Mio; Kohashi, Kenichi; Kushitani, Kei; Yoshida, Misa; Kurihara, Sho; Kawashima, Masumi; Ueda, Yuka; Souzaki, Ryota; Kinoshita, Yoshiaki; Oda, Yoshinao; Takeshima, Yukio; Hiyama, Eiso; Taguchi, Tomoaki; Tanaka, Yukichi

2017-08-01

We report 2 infantile cases of pulmonary tumor carrying a chimeric A2M-ALK gene. A2M-ALK is a newly identified anaplastic lymphoma kinase (ALK)-related chimeric gene from a tumor diagnosed as fetal lung interstitial tumor (FLIT). FLIT is a recently recognized infantile pulmonary lesion defined as a mass-like lesion that morphologically resembles the fetal lung. Grossly, FLIT characteristically appears as a well-circumscribed spongy mass, whereas the tumors in these patients were solid and firm. Histologically, the tumors showed intrapulmonary lesions composed of densely proliferating polygonal or spindle-shaped mesenchymal cells with diffuse and dense infiltrations of inflammatory cells forming microcystic or micropapillary structures lined by thyroid transcription factor 1-positive pneumocytes, favoring inflammatory myofibroblastic tumor rather than FLIT. The proliferating cells were immunoreactive for ALK, and A2M-ALK was identified in both tumors with reverse-transcription polymerase chain reaction. The dense infiltration of inflammatory cells, immunoreactivity for ALK, and identification of an ALK-related chimeric gene suggested a diagnosis of inflammatory myofibroblastic tumor. Histologically, most reported FLITs show sparse inflammatory infiltrates and a relatively low density of interstitial cells in the septa, although prominent infiltration of inflammatory cells and high cellularity of interstitial cells are seen in some FLITs. The present cases suggest that ALK rearrangements, including the chimeric A2M-ALK gene, may be present in these infantile pulmonary lesions, especially those with inflammatory cell infiltration. We propose that these infantile pulmonary lesions containing a chimeric A2M-ALK gene be categorized as a specific type of inflammatory myofibroblastic tumor that develops exclusively in neonates and infants. Copyright © 2017 Elsevier Inc. All rights reserved.

Origin of the chromosomal radiation of Madeiran house mice: a microsatellite analysis of metacentric chromosomes

PubMed Central

Förster, D W; Mathias, M L; Britton-Davidian, J; Searle, J B

2013-01-01

Chromosome races of Mus musculus domesticus are characterised by particular sets of metacentric chromosomes formed by Robertsonian fusions and whole-arm reciprocal translocations. The Atlantic island of Madeira is inhabited by six chromosome races of house mice with 6–9 pairs of metacentric chromosomes. Three of these races are characterised by the metacentric 3.8 also found elsewhere in the distribution of M. m. domesticus, including Denmark and Spain. We investigated the possibility that metacentric 3.8 was introduced to Madeira during the initial colonisation, as this could have ‘seeded' the cascade of chromosomal mutation that is the basis of the extraordinary chromosomal radiation observed on the island. Variation at 24 microsatellite loci mapping to three different chromosomal regions (proximal, interstitial and distal) of mouse chromosomes 3 and 8 was investigated in 179 mice from Madeira, Denmark, Portugal, Spain, Italy and Scotland. Analyses of microsatellite loci closely linked to the centromeres of these chromosomes (‘proximal loci') do not support a common evolutionary origin of metacentric 3.8 among Madeiran, Danish and Spanish mouse populations. Our results suggest that Madeiran mice are genetically more similar to standard karyotype mice from Portugal than to metacentric mice from elsewhere. There is expected to be an interruption to gene flow between hybridising metacentric races on Madeira, particularly in the chromosomal regions close to the rearrangement breakpoints. Consistent with this, relating to differentiation involving chromosomes 3 and 8 on Madeira, we found greater genetic structure among races for proximal than interstitial or distal loci. PMID:23232832

Interstitial fluid drainage is impaired in ischemic stroke and Alzheimer’s disease mouse models

PubMed Central

Arbel-Ornath, Michal; Hudry, Eloise; Eikermann-Haerter, Katharina; Hou, Steven; Gregory, Julia L.; Zhao, Lingzhi; Betensky, Rebecca A.; Frosch, Matthew P.; Greenberg, Steven M.; Bacskai, Brian J.

2013-01-01

The interstitial fluid (ISF) drainage pathway has been hypothesized to underlie the clearance of solutes and metabolites from the brain. Previous work has implicated the perivascular spaces along arteries as the likely route for ISF clearance, however it has never been demonstrated directly. The accumulation of amyloid β (Aβ) peptides in brain parenchyma is one of the pathological hallmarks of Alzheimer disease (AD), and it is likely related to an imbalance between production and clearance of the peptide. Aβ drainage along perivascular spaces has been postulated to be one of the mechanisms that mediates the peptide clearance from the brain. We therefore devised a novel method to visualize solute clearance in real time in the living mouse brain using laser guided bolus dye injections and multiphoton imaging. This methodology allows high spatial and temporal resolution and revealed the kinetics of ISF clearance. We found that the ISF drains along perivascular spaces of arteries and capillaries but not veins, and its clearance exhibits a bi-exponential profile. ISF drainage requires a functional vasculature, as solute clearance decreased when perfusion was impaired. In addition, reduced solute clearance was observed in transgenic mice with significant vascular amyloid deposition; we suggest the existence of a feed-forward mechanism, by which amyloid deposition promotes further amyloid deposition. This important finding provides a mechanistic link between cerebrovascular disease and Alzheimer disease and suggests that facilitation of Aβ clearance along the perivascular pathway should be considered as a new target for therapeutic approaches to AD and CAA. PMID:23818064

In vitro three-dimensional coculturing poly3-hydroxybutyrate-co-3-hydroxyhexanoate with mouse-induced pluripotent stem cells for myocardial patch application.

PubMed

Shijun, Xu; Junsheng, Mu; Jianqun, Zhang; Ping, Bo

2016-03-01

Identifying a suitable polymeric biomaterial for myocardial patch repair following myocardial infarction, cerebral infarction, and cartilage injury is essential. This study aimed to investigate the effect of the novel polymer material, poly3-hydroxybutyrate-co-3-hydroxyhexanoate, on the adhesion, proliferation, and differentiation of mouse-induced pluripotent stem cells in vitro. Mouse-induced pluripotent stem cells were isolated, expanded, and cultured on either two-dimensional or three-dimensional poly3-hydroxybutyrate-co-3-hydroxyhexanoate films (membranes were perforated to imitate three-dimensional space). Following attachment onto the films, mouse-induced pluripotent stem cell morphology was visualized using scanning electron microscopy. Cell vitality was detected using the Cell Counting Kit-8 assay and cell proliferation was observed using fluorescent 4',6-diamidino-2-phenylindole (DAPI) staining. Mouse-induced pluripotent stem cells were induced into cardiomyocytes by differentiation medium containing vitamin C. A control group in the absence of an inducer was included. Mouse-induced pluripotent stem cell survival and differentiation were observed using immunofluorescence and flow cytometry, respectively. Mouse-induced pluripotent stem cells growth, proliferation, and differentiation were observed on both two-dimensional and three-dimensional poly3-hydroxybutyrate-co-3-hydroxyhexanoate films. Vitamin C markedly improved the efficiency of mouse-induced pluripotent stem cells differentiation into cardiomyocytes on poly3-hydroxybutyrate-co-3-hydroxyhexanoate films. Three-dimensional culture was better at promoting mouse-induced pluripotent stem cell proliferation and differentiation compared with two-dimensional culture. © The Author(s) 2016.

A Lifetime's Adventure in Extracellular K+ Regulation: The Scottish Connection.

PubMed

Brown, Angus M

2017-09-01

In a career that has spanned 45 years and shows no signs of slowing down, Dr Bruce Ransom has devoted considerable time and energy to studying regulation of interstitial K + . When Bruce commenced his studies in 1969 virtually nothing was known of the functions of glial cells, but Bruce's research contributed to the physiological assignation of function to mammalian astrocytes, namely interstitial K + buffering. The experiments that I describe in this review concern the response of the membrane potential (Em) of in vivo cat cortical astrocytes to changes in [K + ] o , an experimental manoeuvre that was achieved in two different ways. The first involved recording the Em of an astrocyte while the initial aCSF was switched to one with different K + , whereas in the second series of experiments the cortex was stimulated and the response of the astrocyte Em to the K + released from neighbouring neurons was recorded. The astrocytes responded in a qualitatively predictable manner, but quantitatively the changes were not as predicted by the Nernst equation. Elevations in interstitial K + are not sustained and K + returns to baseline rapidly due to the buffering capacity of astrocytes, a phenomenon studied by Bruce, and his son Chris, published 27 years after Bruce's initial publications. Thus, a lifetime spent investigating K + buffering has seen enormous advances in glial research, from the time cells were identified as 'presumed' glial cells or 'silent cells', to the present day, where glial cells are recognised as contributing to every important physiological brain function.

Expression of peroxisome proliferator-activated receptor alpha messenger ribonucleic acid and protein in human and rat testis.

PubMed

Schultz, R; Yan, W; Toppari, J; Völkl, A; Gustafsson, J A; Pelto-Huikko, M

1999-07-01

Peroxisome proliferator-activated receptor a (PPARalpha), a member of the steroid hormone receptor superfamily, has been linked to lipid homeostasis and tumorigenesis in tissues with high expression of receptor protein. On the other hand, the role of PPARalpha in tissues with a lower expression is not well known. Here we demonstrate the localization of PPARalpha messenger RNA (mRNA) and protein in developing and adult rat testis. Additionally, we demonstrate the expression of PPARalpha protein in adult human testis. Our experiments with Northern analysis, in situ hybridization and immunocytochemistry reveal a complex distribution of PPARalpha in tubular and interstitial cells of both adult and developing rat testis. The overall expression is rather low but may be modified by exogenous or endogenous stimuli. An up-regulation of PPARalpha mRNA could be observed after stimulation with FSH. In the developing rat testis, a clear expression of PPARalpha mRNA was present from the first days after birth. Additionally, PPARalpha mRNA and protein increased toward adulthood. In adult human testis PPARalpha immunoreactivity (IR) was present in interstitial Leydig cells and tubular cells. In the seminiferous epithelium of adult human testis the expression of PPARalpha-IR could be seen in meiotic spermatocytes, spermatids and myoid peritubular cells. The findings of our study suggest that PPARalpha may be involved in the regulation of growth and differentiation of tubular and interstitial cells in rat and human testis.

Overview of Methods for Overcoming Hindrance to Drug Delivery to Tumors, with Special Attention to Tumor Interstitial Fluid

PubMed Central

Baronzio, Gianfranco; Parmar, Gurdev; Baronzio, Miriam

2015-01-01

Every drug used to treat cancer (chemotherapeutics, immunological, monoclonal antibodies, nanoparticles, radionuclides) must reach the targeted cells through the tumor environment at adequate concentrations, in order to exert their cell-killing effects. For any of these agents to reach the goal cells, they must overcome a number of impediments created by the tumor microenvironment (TME), beginning with tumor interstitial fluid pressure (TIFP), and a multifactorial increase in composition of the extracellular matrix (ECM). A primary modifier of TME is hypoxia, which increases the production of growth factors, such as vascular endothelial growth factor and platelet-derived growth factor. These growth factors released by both tumor cells and bone marrow recruited myeloid cells form abnormal vasculature characterized by vessels that are tortuous and more permeable. Increased leakiness combined with increased inflammatory byproducts accumulates fluid within the tumor mass (tumor interstitial fluid), ultimately creating an increased pressure (TIFP). Fibroblasts are also up-regulated by the TME, and deposit fibers that further augment the density of the ECM, thus, further worsening the TIFP. Increased TIFP with the ECM are the major obstacles to adequate drug delivery. By decreasing TIFP and ECM density, we can expect an associated rise in drug concentration within the tumor itself. In this overview, we will describe all the methods (drugs, nutraceuticals, and physical methods of treatment) able to lower TIFP and to modify ECM used for increasing drug concentration within the tumor tissue. PMID:26258072

Hymyc1 downregulation promotes stem cell proliferation in Hydra vulgaris.

PubMed

Ambrosone, Alfredo; Marchesano, Valentina; Tino, Angela; Hobmayer, Bert; Tortiglione, Claudia

2012-01-01

Hydra is a unique model for studying the mechanisms underlying stem cell biology. The activity of the three stem cell lineages structuring its body constantly replenishes mature cells lost due to normal tissue turnover. By a poorly understood mechanism, stem cells are maintained through self-renewal while concomitantly producing differentiated progeny. In vertebrates, one of many genes that participate in regulating stem cell homeostasis is the protooncogene c-myc, which has been recently identified also in Hydra, and found expressed in the interstitial stem cell lineage. In the present paper, by developing a novel strategy of RNA interference-mediated gene silencing (RNAi) based on an enhanced uptake of small interfering RNAi (siRNA), we provide molecular and biological evidence for an unexpected function of the Hydra myc gene (Hymyc1) in the homeostasis of the interstitial stem cell lineage. We found that Hymyc1 inhibition impairs the balance between stem cell self renewal/differentiation, as shown by the accumulation of stem cell intermediate and terminal differentiation products in genetically interfered animals. The identical phenotype induced by the 10058-F4 inhibitor, a disruptor of c-Myc/Max dimerization, demonstrates the specificity of the RNAi approach. We show the kinetic and the reversible feature of Hymyc1 RNAi, together with the effects displayed on regenerating animals. Our results show the involvement of Hymyc1 in the control of interstitial stem cell dynamics, provide new clues to decipher the molecular control of the cell and tissue plasticity in Hydra, and also provide further insights into the complex myc network in higher organisms. The ability of Hydra cells to uptake double stranded RNA and to trigger a RNAi response lays the foundations of a comprehensive analysis of the RNAi response in Hydra allowing us to track back in the evolution and the origin of this process.

Hymyc1 Downregulation Promotes Stem Cell Proliferation in Hydra vulgaris

PubMed Central

Ambrosone, Alfredo; Marchesano, Valentina; Tino, Angela; Hobmayer, Bert; Tortiglione, Claudia

2012-01-01

Hydra is a unique model for studying the mechanisms underlying stem cell biology. The activity of the three stem cell lineages structuring its body constantly replenishes mature cells lost due to normal tissue turnover. By a poorly understood mechanism, stem cells are maintained through self-renewal while concomitantly producing differentiated progeny. In vertebrates, one of many genes that participate in regulating stem cell homeostasis is the protooncogene c-myc, which has been recently identified also in Hydra, and found expressed in the interstitial stem cell lineage. In the present paper, by developing a novel strategy of RNA interference-mediated gene silencing (RNAi) based on an enhanced uptake of small interfering RNAi (siRNA), we provide molecular and biological evidence for an unexpected function of the Hydra myc gene (Hymyc1) in the homeostasis of the interstitial stem cell lineage. We found that Hymyc1 inhibition impairs the balance between stem cell self renewal/differentiation, as shown by the accumulation of stem cell intermediate and terminal differentiation products in genetically interfered animals. The identical phenotype induced by the 10058-F4 inhibitor, a disruptor of c-Myc/Max dimerization, demonstrates the specificity of the RNAi approach. We show the kinetic and the reversible feature of Hymyc1 RNAi, together with the effects displayed on regenerating animals. Our results show the involvement of Hymyc1 in the control of interstitial stem cell dynamics, provide new clues to decipher the molecular control of the cell and tissue plasticity in Hydra, and also provide further insights into the complex myc network in higher organisms. The ability of Hydra cells to uptake double stranded RNA and to trigger a RNAi response lays the foundations of a comprehensive analysis of the RNAi response in Hydra allowing us to track back in the evolution and the origin of this process. PMID:22292012

Metabolic coupling of glutathione between mouse and quail cardiac myocytes and its protective role against oxidative stress.

PubMed

Nakamura, T Y; Yamamoto, I; Kanno, Y; Shiba, Y; Goshima, K

1994-05-01

Cultured quail myocytes were much more resistant to H2O2 toxicity than cultured mouse myocytes. The intracellular concentration of glutathione ([GSH]i) and the activity of gamma-glutamylcysteine synthetase (gamma-GCS) in quail heart cells were about five and three times higher, respectively, than in mouse heart cells, although catalase and glutathione peroxidase (GSHpx) activity was similar in both. Preloading of gamma-glutamylcysteine monoethyl ester (gamma-GCE), a membrane-permeating GSH precursor, increased the H2O2 resistance of cultured mouse myocytes. These observations suggest that the high [GSH]i and the high activity of gamma-GCS in quail myocytes are responsible for their high resistance to H2O2. Both H2O2 sensitivity and [GSH]i of mosaic sheets composed of equal amounts of mouse and quail myocytes approximated those of sheets composed entirely of quail myocytes. From these observations, it is hypothesized that GSH was transferred from quail myocytes to mouse myocytes, probably through gap junctions between them, and that quail myocytes resynthesized GSH by a feedback mechanism, thus maintaining their intracellular GSH levels. When the fluorescent dye lucifer yellow was injected into a beating quail myocyte in a mosaic sheet, it spread to neighboring mouse myocytes but not to neighboring L cells (a cell line derived from mouse connective tissue). These observations indicate that existence of gap junctions in the region of cell contact between mouse and quail myocytes but not between quail myocytes and L cells. When quail myocytes preloaded with [3H]gamma-GCE were cocultured with mouse myocytes and L cells, the radioactivity was transmitted to neighboring mouse myocytes but not L cells. These observations show that GSH and/or its precursors can be transmitted from quail myocytes to mouse myocytes through gap junctions and that this can protect mouse myocytes from H2O2 toxicity. Mouse myocyte sheets composed of 10(4) cells or more showed higher resistance to H2O2 toxicity than single isolated mouse myocytes. Metabolic coupling of GSH between myocytes may contribute at least in part to this high resistance of the cell sheets.

Populations of subplate and interstitial neurons in fetal and adult human telencephalon.

PubMed

Judaš, Miloš; Sedmak, Goran; Pletikos, Mihovil; Jovanov-Milošević, Nataša

2010-10-01

In the adult human telencephalon, subcortical (gyral) white matter contains a special population of interstitial neurons considered to be surviving descendants of fetal subplate neurons [Kostovic & Rakic (1980) Cytology and the time of origin of interstitial neurons in the white matter in infant and adult human and monkey telencephalon. J Neurocytol9, 219]. We designate this population of cells as superficial (gyral) interstitial neurons and describe their morphology and distribution in the postnatal and adult human cerebrum. Human fetal subplate neurons cannot be regarded as interstitial, because the subplate zone is an essential part of the fetal cortex, the major site of synaptogenesis and the 'waiting' compartment for growing cortical afferents, and contains both projection neurons and interneurons with distinct input-output connectivity. However, although the subplate zone is a transient fetal structure, many subplate neurons survive postnatally as superficial (gyral) interstitial neurons. The fetal white matter is represented by the intermediate zone and well-defined deep periventricular tracts of growing axons, such as the corpus callosum, anterior commissure, internal and external capsule, and the fountainhead of the corona radiata. These tracts gradually occupy the territory of transient fetal subventricular and ventricular zones.The human fetal white matter also contains distinct populations of deep fetal interstitial neurons, which, by virtue of their location, morphology, molecular phenotypes and advanced level of dendritic maturation, remain distinct from subplate neurons and neurons in adjacent structures (e.g. basal ganglia, basal forebrain). We describe the morphological, histochemical (nicotinamide-adenine dinucleotide phosphate-diaphorase) and immunocytochemical (neuron-specific nuclear protein, microtubule-associated protein-2, calbindin, calretinin, neuropeptide Y) features of both deep fetal interstitial neurons and deep (periventricular) interstitial neurons in the postnatal and adult deep cerebral white matter (i.e. corpus callosum, anterior commissure, internal and external capsule and the corona radiata/centrum semiovale). Although these deep interstitial neurons are poorly developed or absent in the brains of rodents, they represent a prominent feature of the significantly enlarged white matter of human and non-human primate brains. © 2010 The Authors. Journal of Anatomy © 2010 Anatomical Society of Great Britain and Ireland.

Early history of subplate and interstitial neurons: from Theodor Meynert (1867) to the discovery of the subplate zone (1974)

PubMed Central

Judaš, Miloš; Sedmak, Goran; Pletikos, Mihovil

2010-01-01

In this historical review, we trace the early history of research on the fetal subplate zone, subplate neurons and interstitial neurons in the white matter of the adult nervous system. We arrive at several general conclusions. First, a century of research clearly testifies that interstitial neurons, subplate neurons and the subplate zone were first observed and variously described in the human brain – or, in more general terms, in large brains of gyrencephalic mammals, characterized by an abundant white matter and slow and protracted prenatal and postnatal development. Secondly, the subplate zone cannot be meaningfully defined using a single criterion – be it a specific population of cells, fibres or a specific molecular or genetic marker. The subplate zone is a highly dynamic architectonic compartment and its size and cellular composition do not remain constant during development. Thirdly, it is important to make a clear distinction between the subplate zone and the subplate (and interstitial) neurons. The transient existence of the subplate zone (as a specific architectonic compartment of the fetal telencephalic wall) should not be equated with the putative transient existence of subplate neurons. It is clear that in rodents, and to an even greater extent in humans and monkeys, a significant number of subplate cells survive and remain functional throughout life. PMID:20979585

Retinoid agonist isotretinoin ameliorates obstructive renal injury.

PubMed

Schaier, Matthias; Jocks, Thomas; Grone, Hermann-Josef; Ritz, Eberhard; Wagner, Juergen

2003-10-01

Interstitial fibrosis is a major cause of end stage renal failure. Retinoids, which are involved in tissue repair and fibrosis, inhibit inflammatory and proliferative pathways. Therefore, we studied the dose dependent effects of the retinoid receptor agonist isotretinoin 13-cis retinoic acid in the unilateral ureteral obstruction model (UUO). Sham operated control rats were compared with UUO rats treated with vehicle (UUO-Veh), or low (5 mg/kg body weight (UUO-LD) or high (25 mg/kg) (UUO-HD) dose isotretinoin. Kidneys were evaluated using reverse transcriptase-polymerase chain reaction and immunohistology 7 days after UUO. Renal injury and fibrosis were quantified by immunostaining and expression measurements of the genes involved in renal fibrosis. In UUO-Veh kidneys the interstitial area was expanded 5-fold but only 3-fold in UUO-HD and 3.5-fold in UUO-LD rats. Interstitial cell counts were 3-fold higher in UUO-Veh rats but significantly less in UUO-HD or UUO-LD animals. Tubular and interstitial cell proliferation was significantly higher in UUO-Veh rats compared with sham operated control plus vehicle animals but less so in UUO-LD and UUO-HD rats. In UUO-Veh rats interstitial infiltration by monocytes/macrophages was higher compared with unobstructed controls. It was significantly less after isotretinoin treatment. In UUO-Veh rats mRNA for procollagen I, and transforming growth factor-beta1 and II receptor was significantly increased. It was significantly less after treatment with isotretinoin. Fibronectin and collagen I immunostaining was also decreased by isotretinoin. Since isotretinoin limits proliferation, inflammation and fibrosis after UUO, retinoids should be further investigated as potentially promising therapeutic agents for renal disease.

Mouse neuroblastoma cell based model and the effect of epileptic events on calcium oscillations and neural spikes

NASA Astrophysics Data System (ADS)

Kim, Suhwan; Baek, Juyeong; Jung, Unsang; Lee, Sangwon; Jung, Woonggyu; Kim, Jeehyun; Kang, Shinwon

2013-05-01

Recently, Mouse neuroblastoma cells are considered as an attractive model for the study of human neurological and prion diseases, and intensively used as a model system in different areas. Among those areas, differentiation of neuro2a (N2A) cells, receptor mediated ion current, and glutamate induced physiological response are actively investigated. The reason for the interest to mouse neuroblastoma N2A cells is that they have a fast growing rate than other cells in neural origin with a few another advantages. This study evaluated the calcium oscillations and neural spikes recording of mouse neuroblastoma N2A cells in an epileptic condition. Based on our observation of neural spikes in mouse N2A cell with our proposed imaging modality, we report that mouse neuroblastoma N2A cells can be an important model related to epileptic activity studies. It is concluded that the mouse neuroblastoma N2A cells produce the epileptic spikes in vitro in the same way as produced by the neurons or the astrocytes. This evidence advocates the increased and strong level of neurotransmitters release by enhancement in free calcium using the 4-aminopyridine which causes the mouse neuroblastoma N2A cells to produce the epileptic spikes and calcium oscillation.

Telocytes and putative stem cells in ageing human heart

PubMed Central

Popescu, Laurentiu M; Curici, Antoanela; Wang, Enshi; Zhang, Hao; Hu, Shengshou; Gherghiceanu, Mihaela

2015-01-01

Tradition considers that mammalian heart consists of about 70% non-myocytes (interstitial cells) and 30% cardiomyocytes (CMs). Anyway, the presence of telocytes (TCs) has been overlooked, since they were described in 2010 (visit http://www.telocytes.com). Also, the number of cardiac stem cells (CSCs) has not accurately estimated in humans during ageing. We used electron microscopy to identify and estimate the number of cells in human atrial myocardium (appendages). Three age-related groups were studied: newborns (17 days–1 year), children (6–17 years) and adults (34–60 years). Morphometry was performed on low-magnification electron microscope images using computer-assisted technology. We found that interstitial area gradually increases with age from 31.3 ± 4.9% in newborns to 41 ± 5.2% in adults. Also, the number of blood capillaries (per mm2) increased with several hundreds in children and adults versus newborns. CMs are the most numerous cells, representing 76% in newborns, 88% in children and 86% in adults. Images of CMs mitoses were seen in the 17-day newborns. Interestingly, no lipofuscin granules were found in CMs of human newborns and children. The percentage of cells that occupy interstitium were (depending on age): endothelial cells 52–62%; vascular smooth muscle cells and pericytes 22–28%, Schwann cells with nerve endings 6–7%, fibroblasts 3–10%, macrophages 1–8%, TCs about 1% and stem cells less than 1%. We cannot confirm the popular belief that cardiac fibroblasts are the most prevalent cell type in the heart and account for about 20% of myocardial volume. Numerically, TCs represent a small fraction of human cardiac interstitial cells, but because of their extensive telopodes, they achieve a 3D network that, for instance, supports CSCs. The myocardial (very) low capability to regenerate may be explained by the number of CSCs, which decreases fivefold by age (from 0.5% to 0.1% in newborns versus adults). PMID:25545142

The Terminator mouse is a diphtheria toxin-receptor knock-in mouse strain for rapid and efficient enrichment of desired cell lineages.

PubMed

Guo, Jian-Kan; Shi, Hongmei; Koraishy, Farrukh; Marlier, Arnaud; Ding, Zhaowei; Shan, Alan; Cantley, Lloyd G

2013-11-01

Biomedical research often requires primary cultures of specific cell types, which are challenging to obtain at high purity in a reproducible manner. Here we engineered the murine Rosa26 locus by introducing the diphtheria toxin receptor flanked by loxP sites. The resultant strain was nicknamed the Terminator mouse. This approach results in diphtheria toxin-receptor expression in all non-Cre expressing cell types, making these cells susceptible to diphtheria toxin exposure. In primary cultures of kidney cells derived from the Terminator mouse, over 99.99% of cells were dead within 72 h of diphtheria toxin treatment. After crossing the Terminator with the podocin-Cre (podocyte specific) mouse or the Ggt-Cre (proximal tubule specific) mouse, diphtheria toxin treatment killed non-Cre expressing cells but spared podocytes and proximal tubule cells, respectively, enriching the primary cultures to over 99% purity, based on both western blotting and immunostaining of marker proteins. Thus, the Terminator mouse can be a useful tool to selectively and reproducibly obtain even low-abundant cell types at high quantity and purity.

Differential biological effects of dehydroepiandrosterone (DHEA) between mouse (B16F10) and human melanoma (BLM) cell lines.

PubMed

Joshi, Kumud; Hassan, Sherif S; Ramaraj, Pandurangan

2017-01-01

Dehydroepiandrosterone (DHEA) is a weak androgen and had been shown to have anti-cancer, anti-adipogenic and anti-inflammatory effects on mouse and other rodent models, but not on humans, suggesting a systemic level difference between mouse and human. Our previous study on DHEA biological functions involving a variety of cell lines, suggested that the functional differences between mouse and human existed even at the cellular level. Hence, using mouse and human melanoma cell models, in-vitro effects of DHEA on cell growth, mechanism of cell death and mechanism of DHEA action were studied. Results indicated a differential biological effects of DHEA between mouse and human melanoma cell lines. These in-vitro studies also suggested that the differential biological effects observed between these two cell lines could be due to the difference in the way DHEA was processed or metabolized inside the cell.

Evidence for the involvement of fibroblast growth factor 10 in lipofibroblast formation during embryonic lung development

PubMed Central

Al Alam, Denise; El Agha, Elie; Sakurai, Reiko; Kheirollahi, Vahid; Moiseenko, Alena; Danopoulos, Soula; Shrestha, Amit; Schmoldt, Carole; Quantius, Jennifer; Herold, Susanne; Chao, Cho-Ming; Tiozzo, Caterina; De Langhe, Stijn; Plikus, Maksim V.; Thornton, Matthew; Grubbs, Brendan; Minoo, Parviz; Rehan, Virender K.; Bellusci, Saverio

2015-01-01

Lipid-containing alveolar interstitial fibroblasts (lipofibroblasts) are increasingly recognized as an important component of the epithelial stem cell niche in the rodent lung. Although lipofibroblasts were initially believed merely to assist type 2 alveolar epithelial cells in surfactant production during neonatal life, recent evidence suggests that these cells are indispensable for survival and growth of epithelial stem cells during adulthood. Despite increasing interest in lipofibroblast biology, little is known about their cellular origin or the molecular pathways controlling their formation during embryonic development. Here, we show that a population of lipid-droplet-containing stromal cells emerges in the developing mouse lung between E15.5 and E16.5. This is accompanied by significant upregulation, in the lung mesenchyme, of peroxisome proliferator-activated receptor gamma (master switch of lipogenesis), adipose differentiation-related protein (marker of mature lipofibroblasts) and fibroblast growth factor 10 (previously shown to identify a subpopulation of lipofibroblast progenitors). We also demonstrate that although only a subpopulation of total embryonic lipofibroblasts derives from Fgf10+ progenitor cells, in vivo knockdown of Fgfr2b ligand activity and reduction in Fgf10 expression lead to global reduction in the expression levels of lipofibroblast markers at E18.5. Constitutive Fgfr1b knockouts and mutants with conditional partial inactivation of Fgfr2b in the lung mesenchyme reveal the involvement of both receptors in lipofibroblast formation and suggest a possible compensation between the two receptors. We also provide data from human fetal lungs to demonstrate the relevance of our discoveries to humans. Our results reveal an essential role for Fgf10 signaling in the formation of lipofibroblasts during late lung development. PMID:26511927

Vascular Endothelial Cell-Specific Connective Tissue Growth Factor (CTGF) Is Necessary for Development of Chronic Hypoxia-Induced Pulmonary Hypertension.

PubMed

Pi, Liya; Fu, Chunhua; Lu, Yuanquing; Zhou, Junmei; Jorgensen, Marda; Shenoy, Vinayak; Lipson, Kenneth E; Scott, Edward W; Bryant, Andrew J

2018-01-01

Chronic hypoxia frequently complicates the care of patients with interstitial lung disease, contributing to the development of pulmonary hypertension (PH), and premature death. Connective tissue growth factor (CTGF), a matricellular protein of the Cyr61/CTGF/Nov (CCN) family, is known to exacerbate vascular remodeling within the lung. We have previously demonstrated that vascular endothelial-cell specific down-regulation of CTGF is associated with protection against the development of PH associated with hypoxia, though the mechanism for this effect is unknown. In this study, we generated a transgenic mouse line in which the Ctgf gene was floxed and deleted in vascular endothelial cells that expressed Cre recombinase under the control of VE-Cadherin promoter (eCTGF KO mice). Lack of vascular endothelial-derived CTGF protected against the development of PH secondary to chronic hypoxia, as well as in another model of bleomycin-induced pulmonary hypertension. Importantly, attenuation of PH was associated with a decrease in infiltrating inflammatory cells expressing CD11b or integrin α M (ITGAM), a known adhesion receptor for CTGF, in the lungs of hypoxia-exposed eCTGF KO mice. Moreover, these pathological changes were associated with activation of-Rho GTPase family member-cell division control protein 42 homolog (Cdc42) signaling, known to be associated with alteration in endothelial barrier function. These data indicate that endothelial-specific deletion of CTGF results in protection against development of chronic-hypoxia induced PH. This protection is conferred by both a decrease in inflammatory cell recruitment to the lung, and a reduction in lung Cdc42 activity. Based on our studies, CTGF inhibitor treatment should be investigated in patients with PH associated with chronic hypoxia secondary to chronic lung disease.

Cell-type specific role of the RNA-binding protein, NONO, in the DNA double-strand break response in the mouse testes.

PubMed

Li, Shuyi; Shu, Feng-Jue; Li, Zhentian; Jaafar, Lahcen; Zhao, Shourong; Dynan, William S

2017-03-01

The tandem RNA recognition motif protein, NONO, was previously identified as a candidate DNA double-strand break (DSB) repair factor in a biochemical screen for proteins with end-joining stimulatory activity. Subsequent work showed that NONO and its binding partner, SFPQ, have many of the properties expected for bona fide repair factors in cell-based assays. Their contribution to the DNA damage response in intact tissue in vivo has not, however, been demonstrated. Here we compare DNA damage sensitivity in the testes of wild-type mice versus mice bearing a null allele of the NONO homologue (Nono gt ). In wild-type mice, NONO protein was present in Sertoli, peritubular myoid, and interstitial cells, with an increase in expression following induction of DNA damage. As expected for the product of an X-linked gene, NONO was not detected in germ cells. The Nono gt/0 mice had at most a mild testis developmental phenotype in the absence of genotoxic stress. However, following irradiation at sublethal, 2-4 Gy doses, Nono gt/0 mice displayed a number of indicators of radiosensitivity as compared to their wild-type counterparts. These included higher levels of persistent DSB repair foci, increased numbers of apoptotic cells in the seminiferous tubules, and partial degeneration of the blood-testis barrier. There was also an almost complete loss of germ cells at later times following irradiation, evidently arising as an indirect effect reflecting loss of stromal support. Results demonstrate a role for NONO protein in protection against direct and indirect biological effects of ionizing radiation in the whole animal. Copyright © 2017 Elsevier B.V. All rights reserved.

Evidence for the involvement of fibroblast growth factor 10 in lipofibroblast formation during embryonic lung development.

PubMed

Al Alam, Denise; El Agha, Elie; Sakurai, Reiko; Kheirollahi, Vahid; Moiseenko, Alena; Danopoulos, Soula; Shrestha, Amit; Schmoldt, Carole; Quantius, Jennifer; Herold, Susanne; Chao, Cho-Ming; Tiozzo, Caterina; De Langhe, Stijn; Plikus, Maksim V; Thornton, Matthew; Grubbs, Brendan; Minoo, Parviz; Rehan, Virender K; Bellusci, Saverio

2015-12-01

Lipid-containing alveolar interstitial fibroblasts (lipofibroblasts) are increasingly recognized as an important component of the epithelial stem cell niche in the rodent lung. Although lipofibroblasts were initially believed merely to assist type 2 alveolar epithelial cells in surfactant production during neonatal life, recent evidence suggests that these cells are indispensable for survival and growth of epithelial stem cells during adulthood. Despite increasing interest in lipofibroblast biology, little is known about their cellular origin or the molecular pathways controlling their formation during embryonic development. Here, we show that a population of lipid-droplet-containing stromal cells emerges in the developing mouse lung between E15.5 and E16.5. This is accompanied by significant upregulation, in the lung mesenchyme, of peroxisome proliferator-activated receptor gamma (master switch of lipogenesis), adipose differentiation-related protein (marker of mature lipofibroblasts) and fibroblast growth factor 10 (previously shown to identify a subpopulation of lipofibroblast progenitors). We also demonstrate that although only a subpopulation of total embryonic lipofibroblasts derives from Fgf10(+) progenitor cells, in vivo knockdown of Fgfr2b ligand activity and reduction in Fgf10 expression lead to global reduction in the expression levels of lipofibroblast markers at E18.5. Constitutive Fgfr1b knockouts and mutants with conditional partial inactivation of Fgfr2b in the lung mesenchyme reveal the involvement of both receptors in lipofibroblast formation and suggest a possible compensation between the two receptors. We also provide data from human fetal lungs to demonstrate the relevance of our discoveries to humans. Our results reveal an essential role for Fgf10 signaling in the formation of lipofibroblasts during late lung development. © 2015. Published by The Company of Biologists Ltd.

A developmentally plastic adult mouse kidney cell line spontaneously generates multiple adult kidney structures

DOE Office of Scientific and Technical Information (OSTI.GOV)

Webb, Carol F., E-mail: carol-webb@omrf.org; Immunobiology and Cancer Research, Oklahoma Medical Research Foundation, Oklahoma City, OK; Department of Microbiology and Immunology, University of Oklahoma Health Sciences Center, Oklahoma City, OK

Despite exciting new possibilities for regenerative therapy posed by the ability to induce pluripotent stem cells, recapitulation of three-dimensional kidneys for repair or replacement has not been possible. ARID3a-deficient mouse tissues generated multipotent, developmentally plastic cells. Therefore, we assessed the adult mouse ARID3a−/− kidney cell line, KKPS5, which expresses renal progenitor surface markers as an alternative cell source for modeling kidney development. Remarkably, these cells spontaneously developed into multicellular nephron-like structures in vitro, and engrafted into immunocompromised medaka mesonephros, where they formed mouse nephron structures. These data implicate KKPS5 cells as a new model system for studying kidney development. - Highlights:more » • An ARID3a-deficient mouse kidney cell line expresses multiple progenitor markers. • This cell line spontaneously forms multiple nephron-like structures in vitro. • This cell line formed mouse kidney structures in immunocompromised medaka fish kidneys. • Our data identify a novel model system for studying kidney development.« less

Are gadolinium-based contrast media nephrotoxic? A renal biopsy study.

PubMed

Akgun, Hulya; Gonlusen, Gulfiliz; Cartwright, Joiner; Suki, Wadi N; Truong, Luan D

2006-09-01

Gadolinium-based contrast media were originally introduced as alternatives to iodinated media for magnetic resonance imaging. Although originally thought to be nonnephrotoxic, gadolinium-based contrast media have recently been reported to be associated with acute renal failure; the mechanism and the underlying renal injury are not completely understood. We report what is, to our knowledge, the first renal biopsy in this context. A 56-year-old patient underwent 2 consecutive vascular imaging procedures in conjunction with gadolinium-based contrast medium administration. A few days later, the patient developed acute renal failure. A renal biopsy showed acute tubular cell injury including patchy tubular cell necrosis, tubular cell degeneration, and marked proliferation of tubular cells, together with mild interstitial edema and interstitial inflammation, but without significant glomerular or vascular changes. During supportive therapy, renal function was partially regained. This case emphasizes the potential nephrotoxicity of gadolinium-based contrast media and suggests that the nephrotoxicity is related to potentially reversible acute tubular cell injury.

Release of enzymes from cells: transport and distribution within the extracellular space.

PubMed

Mattenheimer, H; Friedel, R

1977-01-01

The distribution in the extracellular space of enzymes released from organ cells was investigated using three models: (1) comparison of enzyme activities in blood plasma and lymph of the ductus thoracicus (dog) and plasma and intestinal lymph (rat); (2) i.v. injection of heterologous, homologous and autologous enzymes in order to increase acutely the activities and to measure the rate constants for the distribution and elimination of the enzymes (rat); or (3) plasmapheresis in order to create an enzyme activity gradient from the interstitial space and to determine the rate constants for the reestablishment of the equilibrium between the extra and intravascular compartments (rat). The results suggest that the enzymes are mainly released into the interstitial fluid and transported via the lymph into the intravascular compartment. From there the enzymes diffuse back into the interstitial compartment and are eliminated by a yet unknown mechanism. Transport of enzymes across the capillary membranes in both directions depends on (1) the permeability of the capillary membranes, which varies from region to region and (2) the molecular seizes of the enzymes.

Cellular complexity in subcortical white matter: a distributed control circuit?

PubMed

Colombo, Jorge A

2018-03-01

The subcortical white matter (SWM) has been traditionally considered as a site for passive-neutral-information transfer through cerebral cortex association and projection fibers. Yet, the presence of subcortical neuronal and glial "interstitial" cells expressing immunolabelled neurotransmitters/neuromodulators and synaptic vesicular proteins, and recent immunohistochemical and electrophysiological observations on the rat visual cortex as well as interactive regulation of myelinating processes support the possibility that SWM nests subcortical, regionally variable, distributed neuronal-glial circuits, that could influence information transfer. Their hypothetical involvement in regulating the timing and signal transfer probability at the SWM axonal components ought to be considered and experimentally analysed. Thus, the "interstitial" neuronal cells-associated with local glial cells-traditionally considered to be vestigial and functionally inert under normal conditions, they may well turn to be critical in regulating information transfer at the SWM.

Effects of Arg-Gly-Asp sequence peptide and hyperosmolarity on the permeability of interstitial matrix and fenestrated endothelium in joints.

PubMed

Poli, A; Mason, R M; Levick, J R

2004-09-01

The aims were to assess the contribution of arg-gly-asp (RGD) mediated cell integrin-matrix bonds to interstitial hydraulic resistance and to fenestrated endothelial permeability in joints. Joint fluid is generated by filtration from fenestrated capillaries and drains through a fibronectin-rich synovial intercellular matrix. The role of parenchymal cell-matrix bonding in determining tissue hydraulic resistance is unknown. The knee cavity of anesthetized rabbits was infused with saline or the competitive hexapeptide blocker GRGDTP, with or without added osmotic stress (600 mosm saline). Intra-articular pressure Pj, net trans-synovial drainage rate s, and the permeation of Evans blue-labeled albumin (EVA) from plasma into the joint cavity were measured. GRGDTP increased the hydraulic conductance of the synovial drainage pathway, ds/dPj, by 71% (p =.02, paired t test, n = 6 animals). Synovial plasma EVA clearance (control 7.1 +/- 0.8 microL h-1, mean +/- SEM, n = 15) was unaffected by GRGDTP (7.0 +/- 2.3 microL h(-1), n = 6) or hyperosmolarity (4.9 +/- 1.5 microL h(-1), n = 8) but was increased by GRGDTP and hyperosmolarity together (15.9 +/- 4.8 microL h(-1), n = 5) (p =.01, ANOVA). Changes in dPj/dt evoked by GRGDTP plus hyperosmolarity, but neither alone, demonstrated increased microvascular filtration into the joint cavity (p <.001, ANOVA), as did changes in fluid absorption from the infusion system at fixed Pj. RGD-mediated bonds between the parenchymal cells and interstitial polymers reduce the interstitial hydraulic conductance by 42%. This helps to retain the lubricating fluid inside a joint cavity. RGD-mediated bonds also support the macromolecular barrier function of fenestrated endothelium, but in vivo this is evident only in stressed endothelium (cf. in vitro).

A case of pneumocystis pneumonia associated with everolimus therapy for renal cell carcinoma.

PubMed

Saito, Yoshinobu; Nagayama, Mikie; Miura, Yukiko; Ogushi, Satoko; Suzuki, Yasutomo; Noro, Rintaro; Minegishi, Yuji; Kimura, Go; Kondo, Yukihiro; Gemma, Akihiko

2013-05-01

A 76-year-old female with advanced renal cell carcinoma had been treated with everolimus for 3 months. She visited our hospital because of a cough and fever lasting a few days. Chest X-rays showed bilateral infiltrative shadows, and a chest computed tomography scan showed homogeneous ground-glass opacities with mosaic patterns, especially in the apical region. The laboratory results revealed a decreased white blood cell count with lymphocytopenia and high levels of lactate dehydrogenase, C-reactive protein and KL-6. Pneumonitis was suspected and, therefore, everolimus therapy was interrupted. At that time, the pneumonitis was thought to be drug-induced interstitial lung disease. However, it was not possible to rule out pneumocystis pneumonia, because the patient was immunocompromised and the computed tomography findings suggested the possibility of pneumocystis pneumonia. The pneumonitis progressed rapidly and the patient developed respiratory failure, so we performed bronchoalveolar lavage to make a definitive diagnosis, and simultaneously started treatment with prednisolone and trimethoprim-sulfamethoxazole to cover both interstitial lung disease and pneumocystis pneumonia. A polymerase chain reaction assay of the bronchoalveolar lavage fluid was positive for Pneumocystis carinii DNA, and the serum level of β-d-glucan was significantly elevated. Thus, the patient was diagnosed with pneumocystis pneumonia, which was cured by the treatment. Interstitial lung disease is a major adverse drug reaction associated with everolimus, and interstitial lung disease is the first condition suspected when a patient presents with pneumonitis during everolimus therapy. Pneumocystis pneumonia associated with everolimus therapy is rare, but our experience suggests that pneumocystis pneumonia should be considered as a differential diagnosis when pneumonitis is encountered in patients receiving everolimus therapy.

Mouse IgA inhibits cell growth by stimulating tumor necrosis factor-alpha production and apoptosis of macrophage cell lines.

PubMed

Reljic, Rajko; Crawford, Carol; Challacombe, Stephen; Ivanyi, Juraj

2004-04-01

Potent Fcalpha-mediated actions of IgA have previously been shown for myeloid cells from man, but much less is known in relation to murine cells. Here, we report that mouse monoclonal IgA, irrespective of their antigenic specificity, inhibit the proliferation of mouse macrophage cell lines. The anti-proliferative activity was manifested by both monomeric and polymeric mouse IgA, but not by mouse monoclonal IgG and IgM. Growth of J774 cells was significantly inhibited during the 4-8 days of logarithmic growth, followed by a subsequent recovery of cell numbers prior to the stationary phase. We demonstrated that IgA binds to J774 cells, stimulates tumor necrosis factor (TNF)-alpha production and induces apoptosis which is not dependent on NO or FAS/CD95. We also demonstrated that IgA, in synergy with IFN-gamma, induced TNF-alpha production and apoptosis of thioglycollate-elicited mouse peritoneal macrophages. Thus, the in vitro actions of IgA described may also play a regulatory role for mouse macrophages in vivo.

Collagen in organ development

NASA Technical Reports Server (NTRS)

Hardman, P.; Spooner, B. S.

1992-01-01

It is important to know whether microgravity will adversely affect developmental processes. Collagens are macromolecular structural components of the extracellular matrix (ECM) which may be altered by perturbations in gravity. Interstitial collagens have been shown to be necessary for normal growth and morphogenesis in some embryonic organs, and in the mouse salivary gland, the biosynthetic pattern of these molecules changes during development. Determination of the effects of microgravity on epithelial organ development must be preceded by crucial ground-based studies. These will define control of normal synthesis, secretion, and deposition of ECM macromolecules and the relationship of these processes to morphogenesis.

HBV life cycle is restricted in mouse hepatocytes expressing human NTCP.

PubMed

Li, Hanjie; Zhuang, Qiuyu; Wang, Yuze; Zhang, Tianying; Zhao, Jinghua; Zhang, Yali; Zhang, Junfang; Lin, Yi; Yuan, Quan; Xia, Ningshao; Han, Jiahuai

2014-03-01

Recent studies have revealed that human sodium taurocholate cotransporting polypeptide (SLC10A1 or NTCP) is a functional cellular receptor for hepatitis B virus (HBV). However, whether human NTCP can support HBV infection in mouse hepatocyte cell lines has not been clarified. Because an HBV-permissible mouse model would be helpful for the study of HBV pathogenesis, it is necessary to investigate whether human NTCP supports the susceptibility of mouse hepatocyte cell lines to HBV. The results show that exogenous human NTCP expression can render non-susceptible HepG2 (human), Huh7 (human), Hepa1-6 (mouse), AML-12 (mouse) cell lines and primary mouse hepatocyte (PMH) cells susceptible to hepatitis D virus (HDV) which employs HBV envelope proteins. However, human NTCP could only introduce HBV susceptibility in human-derived HepG2 and Huh7 cells, but not in mouse-derived Hepa1-6, AML-12 or PMH cells. These data suggest that although human NTCP is a functional receptor that mediates HBV infection in human cells, it cannot support HBV infection in mouse hepatocytes. Our study indicated that the restriction of HBV in mouse hepatocytes likely occurs after viral entry but prior to viral transcription. We have excluded the role of mouse hepatocyte nuclear factors in the restriction of the HBV life cycle and showed that knockdown or inhibition of Sting, TBK1, IRF3 or IRF7, the components of the anti-viral signaling pathways, had no effect on HBV infection in mouse hepatocytes. Therefore, murine restriction factors that limit HBV infection need to be identified before a HBV-permissible mouse line can be created.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guo, Wenjing; Hao, Baixia; Wang, Qian

Extracellular signal-regulated kinases (ERKs) have been implicated to be dispensable for self-renewal of mouse embryonic stem (ES) cells, and simultaneous inhibition of both ERK signaling and glycogen synthase kinase 3 (GSK3) not only allows mouse ES cells to self-renew independent of extracellular stimuli but also enables more efficient derivation of naïve ES cells from mouse and rat strains. Interestingly, some ERKs stay active in mouse ES cells which are maintained in regular medium containing leukemia inhibitory factor (LIF) and bone morphogenetic protein (BMP). Yet, the upstream signaling for ERK activation and their roles in mouse ES cells, other than promotingmore » or priming differentiation, have not been determined. Here we found that mouse ES cells express three forms of Raf kinases, A-Raf, B-Raf, and C-Raf. Knocking-down each single Raf member failed to affect the sustained ERK activity, neither did A-Raf and B-Raf double knockdown or B-Raf and C-Raf double knockdown change it in ES cells. Interestingly, B-Raf and C-Raf double knockdown, not A-Raf and B-Raf knockdown, inhibited the maximal ERK activation induced by LIF, concomitant with the slower growth of ES cells. On the other hand, A-Raf, B-Raf, and C-Raf triple knockdown markedly inhibited both the maximal and sustained ERK activity in ES cells. Moreover, Raf triple knockdown, similar to the treatment of U-0126, an MEK inhibitor, significantly inhibited the survival and proliferation of ES cells, thereby compromising the colony propagation of mouse ES cells. In summary, our data demonstrate that all three Raf members are required for ERK activation in mouse ES cells and are involved in growth and survival of mouse ES cells. - Highlights: ●Mouse ES (mES) cells express all three Raf members, A-Raf, B-Raf, and C-Raf. ●Leukemia inhibitory factor (LIF) temporally activates ERKs in mES cells. ●B-Raf and C-Raf are required for LIF-induced maximal ERKs activity in mES cells. ●All Raf members are required for LIF-induced sustained ERK activity in mES cells. ●All Raf members are required the survival and proliferation of mES cells.« less

The implication of pro-inflammatory cytokines in the impaired production of gonadal androgens by patients with pulmonary tuberculosis.

PubMed

Bini, Estela Isabel; D'Attilio, Luciano; Marquina-Castillo, Brenda; Mata-Espinosa, Dulce; Díaz, Ariana; Marquez-Velasco, Ricardo; Ramos-Espinosa, Octavio; Gamboa-Domínguez, Armando; Bay, Maria Luisa; Hernández-Pando, Rogelio; Bottasso, Oscar

2015-12-01

The chronic nature of tuberculosis and the protracted immuno-inflammatory reactions are implied in a series of metabolic and immune-endocrine changes accompanying the disease. We explored components from the hypothalamous-pituitary-gonadal axis and their relationship with cytokines involved in disease immunopathology, in male TB patients. Plasma samples from 36 active untreated pulmonary TB male patients were used to determine TNF-α, IFN-γ, TGF-β, IL-6, cortisol, dehydroepiandrosterone, testosterone, progesterone, estradiol, luteinizing hormone (LH) and follicle-stimulating hormone (FSH) by ELISA. Healthy controls corresponded to 21 volunteers without contact with TB patients and similar age (40 ± 16,8 years). Testicular histological samples from necropsies of patients dying from TB were immune-stained for IL-1β, TNF-α, IL-6 and IFN-γ. The TM3 mouse Leydig cell line was incubated with recombinants TNF-α, IFN-γ and TGF-β, supernatants were collected and used to measure testosterone by ELISA. Patients showed decreased levels of testosterone in presence of high amounts of LH, together with augmented IFN-γ, IL-6 and TGF-β levels. Testicular histological sections showed abundant presence of IL-1β, TNF-α, IL-6 and IFN-γ in interstitial macrophages, Sertoli cells and some spermatogonia. In vitro treatment of Leydig cells with these cytokines led to a remarkable reduction of testosterone production. Copyright © 2015 Elsevier Ltd. All rights reserved.

Neutrophil migration through preexisting holes in the basal laminae of alveolar capillaries and epithelium during streptococcal pneumonia.

PubMed

Walker, D C; Behzad, A R; Chu, F

1995-11-01

The purpose of this study was to determine whether or not there are preexisting holes in the endothelial and epithelial basal laminae of alveolar walls and to determine the path taken by neutrophils as they migrate from the capillaries to the airspace of the alveoli during inflammation. Using transmission electron microscopy and serial thin sections of normal rabbit and mouse lung, we have demonstrated the presence of slit-like holes in the capillary basal laminae and round holes in the basal laminae of type 2 pneumocytes. The slits in the capillary basal laminae were observed at the intersection of the thick and thin walls where endothelium, pericytes, and fibroblasts make close contact. The round holes in the type 2 cell basal laminae were observed at sites of close contact with fibroblasts. Neutrophils were observed to migrate through these slits and holes during streptococcal pneumonia in rabbit lungs. We conclude that during inflammation in the lung, migrating neutrophils displace pericytes and fibroblasts from the slits in the capillary basal lamina and then crawl through these slits into the alveolar interstitium. We postulate that neutrophils find their way to type 2 pneumocytes by following interstitial fibroblasts. We believe that neutrophils displace fibroblasts from their close contacts with the type 2 cells and then crawl through the holes in the basal lamina into the basal lateral space of the type 2 cells. From there, neutrophils migrate into the alveolar airspace.

Integrated external and interstitial radiation therapy for primary carcinoma of the vagina

DOE Office of Scientific and Technical Information (OSTI.GOV)

Puthawala, A.; Syed, A.M.; Nalick, R.

1983-09-01

Twenty-seven patients with the diagnosis of primary carcinoma of the vagina were treated by definitive radiotherapy. Twenty-three patients received a combination of external and interstitial iridium-192 implant irradiation and four patients received only interstitial irradiation. Twenty-one patients had squamous cell carcinoma and six had adenocarcinoma. All patients were staged according to the FIGO classification. More than 70% of patients had relatively advanced local disease and conventional intracavitary irradiation was unsuitable. Local tumor control was observed in 85% (23 of 27 patients), and 56% of the patients remain alive and free of disease for a median follow-up period of 50 monthsmore » to a maximum follow-up period of 84 months. Fifteen percent of the patients suffered from treatment-related complications.« less

Somatic cell hybrid mapping on mouse chromosome 11 (MMU11): Assignment of markers relative to two breakpoints in band D

DOE Office of Scientific and Technical Information (OSTI.GOV)

Morris, D.J.; Robinson, T.J.; Adler, I.D.

1993-02-01

Mouse [times] rat somatic cell hybrids were generated by fusing mouse cell lines that are heterozygous for reciprocal translocations involving the T42H and T9Ad breakpoints on mouse chromosome 11 (MMU11) to a thymidine kinase-negative (Tk[sup [minus

[Distribution diversity of integrins and calcium channels on major human and mouse host cells of Leptospira species].

PubMed

Li, Cheng-xue; Zhao, Xin; Qian, Jing; Yan, Jie

2012-07-01

To determine the distribution of integrins and calcium channels on major human and mouse host cells of Leptospira species. The expression of β1, β2 and β3 integrins was detected with immunofluorescence assay on the surface of human monocyte line THP-1, mouse mononuclear-macrophage-like cell line J774A.1, human vascular endothelial cell line HUVEC, mouse vascular endothelial cell EOMA, human hepatocyte line L-02, mouse hepatocyte line Hepa1-6, human renal tubular epithelial cell line HEK-293, mouse glomerular membrane epithelial cell line SV40-MES13, mouse collagen blast line NIH/3T3, human and mouse platelets. The distribution of voltage gate control calcium channels Cav3.1, Cav3.2, Cav3.3 and Cav2.3, and receptor gate calcium channels P(2)X(1), P(2)2X(2), P(2)X(3), P(2)X(4), P(2)X(5), P(2)X(6) and P(2)X(7) were determined with Western blot assay. β1 integrin proteins were positively expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, L-02, Hepa1-6 and HEK-239 cells as well as human and mouse platelets. β2 integrin proteins were expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, and NIH/3T3 cells. β3 integrin proteins were expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, Hepa1-6, HEK-239 and NIH/3T3 cells as well as human and mouse platelets. P(2)X(1) receptor gate calcium channel was expressed on the membrane surface of human and mouse platelets, while P(2)X(5) receptor gate calcium channel was expressed on the membrane surface of J774A.1, THP-1, L-02, Hepa1-6, HEK-239 and HUVEC cells. However, the other calcium channels were not detected on the tested cell lines or platelets. There is a large distribution diversity of integrins and calcium channel proteins on the major human and mouse host cells of Leptospira species, which may be associated with the differences of leptospira-induced injury in different host cells.

Abnormal myocardial fluid retention as an early manifestation of ischemic injury.

PubMed Central

Willerson, J. T.; Scales, F.; Mukherjee, A.; Platt, M.; Templeton, G. H.; Fink, G. S.; Buja, L. M.

1977-01-01

Fifty-seven isolated, blood perfused, continuously weighed canine hearts have been utilized to study the development of abnormal myocardial fluid retention during early myocardial ischemic injury. Inflatable balloon catheters were positioned around the left anterior descending coronary arteries (LAD) of 54 hearts or the proximal left circumflex coronary arteries of three hearts for study of the following intervals of coronary occlusion: a) 10 minutes followed by 20 minutes of reflow, b) 40 minutes followed by either no reflow or by 20 minutes of reflow, and c) 60 minutes without reflow. After 60 minutes of fixed coronary occlusion, histologic and ultrastructural examination revealed mild swelling of many ischemic cardiac muscle cells in the absence of interstitial edema, cardiac weight gain, and obvious structural defects in cell membrane integrity. After 40 minutes of coronary occlusion and 20 minutes of reflow, significant cardiac weight gain occurred in association with characteristic alterations in the ischemic region, including widespread interstitial edema and focal vascular congestion and hemorrhage and swelling of cardiac muscle cells. Focal structural defects in cell membrane integrity were also noted. The development of abnormal myocardial fluid retention after 40 minutes of LAD occlusion occurred in association with a significant reduction in sodium-potassium-ATPase activity in the ischemic area, but with no significant alteration in either creatine phosphokinase or citrate synthase activity in the same region. Despite the abnormal myocardial fluid retention in these hearts, it was possible pharmacologically to vasodilate coronary vessels with adenosine and nitroglycerin infusion to maintain a consistently high coronary flow following release of the coronary occlusion after 40 minutes and to even exceed initial hyperemic flow values following release of the occlusion when adenosine and nitroglycerin infusion was delayed until 15 minutes after reflow. Thus, the data indicate that impaired cell volume regulation and interstitial fluid accumulation and focal structural defects in cell membrane integrity are early manifestations of ischemic injury followed by reflow, but fail to establish a major role for the abnormal fluid retention in altering coronary blood flow prior to the development of extensive myocardial necrosis. In contrast, fixed coronary occlusion for 60 minutes results in mild intracellular swelling but no significant interstitial edema and no obvious structural defects in cell membrane integrity. Images Figure 1 Figure 5 Figure 6 Figure 2 Figure 3 Figure 4 PMID:139829

Development of experimental tumors formed by mouse and human embryonic stem and teratocarcinoma cells after subcutaneous and intraperitoneal transplantations into immunodeficient and immunocompetent mice.

PubMed

Gordeeva, O F; Nikonova, T M

2013-01-01

Pluripotent stem cells represent an attractive cell source for regenerative medicine. However, the risk of teratoma formation after transplantation restricts their clinical application. Therefore, to adequately evaluate the potential risk of tumorigenicity after cell transplantation into human tissues, effective animal transplantation assays need to be developed. We performed a multiparameter (cell number, transplantation site, cell type, host) comparative analysis of the efficiency of tumor development after transplantation of mouse and human embryonic stem (ES) cells and their malignant counterparts, teratocarcinoma (EC) cells, into animal recipients and revealed several key correlations. We found that the efficiency of tumor growth was higher after intraperitoneal than after subcutaneous transplantations of all cell lines studied. The minimal cell numbers sufficient for tumor growth in immunodeficient nude mice were 100-fold lower for intraperitoneal than for subcutaneous transplantations of mouse and human ES cells (10(3) vs. 10(5) and 10(4) vs. 10(6), respectively). Moreover, mouse ES and EC cells formed tumors in immunodeficient and immunocompetent mice more effectively than human ES and EC cells. After intraperitoneal transplantation of 10(3), 10(4), and 10(5) mouse ES cells, teratomas developed in 83%, 100%, and 100% of nude mice, whereas after human ES cell transplantation, teratomas developed in 0%, 17%, and 60%, respectively. In addition, malignant mouse and human EC cells initiated tumor growth after intraperitoneal transplantation significantly faster and more effectively than ES cells. Mouse and human ES cells formed different types of teratomas containing derivatives of three germ layers but different numbers of undifferentiated cells. ES cell-like sublines with differentiation potential similar to the parental cell line were recloned only from mouse, but not from human, ES cell teratomas. These findings provide new information about the possibility and efficiency of tumor growth after transplantation of pluripotent stem cells. This information allows one to predict and possibly prevent the possible risks of tumorigenicity that could arise from stem cell therapeutics.

Interstitial Photoacoustic Sensor for the Measurement of Tissue Temperature during Interstitial Laser Phototherapy

PubMed Central

Li, Zhifang; Chen, Haiyu; Zhou, Feifan; Li, Hui; Chen, Wei R.

2015-01-01

Photothermal therapy is an effective means to induce tumor cell death, since tumor tissue is more sensitive to temperature increases than normal tissue. Biological responses depend on tissue temperature; target tissue temperature needs to be precisely measured and controlled to achieve desired thermal effects. In this work, a unique photoacoustic (PA) sensor is proposed for temperature measurement during interstitial laser phototherapy. A continuous-wave laser light and a pulsed laser light, for photothermal irradiation and photoacoustic temperature measurement, respectively, were delivered to the target tissue through a fiber coupler. During laser irradiation, the PA amplitude was measured. The Grüneisen parameter and the bioheat equation were used to determine the temperature in strategic positions in the target tissue. Our results demonstrate that the interstitial PA amplitude is a linear function of temperature in the range of 22 to 55 °C, as confirmed by thermocouple measurement. Furthermore, by choosing appropriate laser parameters, the maximum temperature surrounding the active diffuse fiber tip in tissue can be controlled in the range of 41 to 55 °C. Thus, this sensor could potentially be used for fast, accurate, and convenient three-dimensional temperature measurement, and for real-time feedback and control of interstitial laser phototherapy in cancer treatment. PMID:25756865

Heterophilic binding of the adhesion molecules poliovirus receptor and immunoglobulin superfamily 4A in the interaction between mouse spermatogenic and Sertoli cells.

PubMed

Wakayama, Tomohiko; Sai, Yoshimichi; Ito, Akihiko; Kato, Yukio; Kurobo, Miho; Murakami, Yoshinori; Nakashima, Emi; Tsuji, Akira; Kitamura, Yukihiko; Iseki, Shoichi

2007-06-01

The cell adhesion protein immunoglobulin superfamily 4A (IGSF4A) is expressed on the surfaces of spermatogenic cells in the mouse testis. During spermatogenesis, IGSF4A is considered to bind to the surface of Sertoli cells in a heterophilic manner. To identify this unknown partner of IGSF4A, we generated rat monoclonal antibodies against the membrane proteins of mouse Sertoli cells grown in primary culture. Using these monoclonal antibodies, we isolated a clone that immunostained Sertoli cells and reacted with the product of immunoprecipitation of the homogenate of mouse testis with anti-IGSF4A antibody. Subsequently, to identify the Sertoli cell membrane protein that is recognized by this monoclonal antibody, we performed expression cloning of a cDNA library from the mouse testis. As a result, we identified poliovirus receptor (PVR), which is another IGSF-type cell adhesion molecule, as the binding partner of IGSF4A. The antibodies raised against PVR and IGSF4A immunoprecipitated both antigens in the homogenate of mouse testis. Immunoreactivity for PVR was present in Sertoli cells but not in spermatogenic cells at all stages of spermatogenesis. Overexpression of PVR in TM4, a mouse Sertoli cell line, increased more than three-fold its capacity to adhere to Tera-2, which is a human cell line that expresses IGSF4A. These findings suggest that the heterophilic binding of PVR to IGSF4A is responsible, at least in part, for the interaction between Sertoli and spermatogenic cells during mouse spermatogenesis.

Slow wave contraction frequency plateaus in the small intestine are composed of discrete waves of interval increase associated with dislocations.

PubMed

Parsons, Sean P; Huizinga, Jan D

2018-06-03

What is the central question of this study? What is the nature of slow wave driven contraction frequency gradients in the small intestine? What is the main finding and its importance? Frequency plateaus are composed of discrete waves of increased interval, each wave associated with a contraction dislocation. Smooth frequency gradients are generated by localised neural modulation of wave frequency, leading to functionally important wave turbulence. Both patterns are emergent properties of a network of coupled oscillators, the interstitial cells of Cajal. A gut-wide network of interstitial cells of Cajal (ICC) generate electrical oscillations (slow waves) that orchestrate waves of muscle contraction. In the small intestine there is a gradient in slow wave frequency from high at the duodenum to low at the terminal ileum. Time-averaged measurements of frequency have suggested either a smooth or stepped (plateaued) gradient. We measured individual contraction intervals from diameter maps of the mouse small intestine to create interval maps (IMaps). IMaps showed that each frequency plateau was composed of discrete waves of increased interval. Each interval wave originated at a terminating contraction wave, a "dislocation", at the plateau's proximal boundary. In a model chain of coupled phase oscillators, interval wave frequency increased as coupling decreased or as the natural frequency gradient or noise increased. Injuring the intestine at a proximal point to destroy coupling, suppressed distal steps which then reappeared with gap junction block by carbenoxolone. This lent further support to our previous hypothesis that lines of dislocations were fixed by points of low coupling strength. Dislocations induced by electrical field pulses in the intestine and by equivalent phase shift in the model, were associated with interval waves. When the enteric nervous system was active, IMaps showed a chaotic, turbulent pattern of interval change with no frequency steps or plateaus. This probably resulted from local, stochastic release of neurotransmitters. Plateaus, dislocations, interval waves and wave turbulence arise from a dynamic interplay between natural frequency and coupling in the ICC network. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Leptospira Interrogans Induces Fibrosis in the Mouse Kidney through Inos-Dependent, TLR- and NLR-Independent Signaling Pathways

PubMed Central

Fanton d'Andon, Martine; Quellard, Nathalie; Fernandez, Béatrice; Ratet, Gwenn; Lacroix-Lamandé, Sonia; Vandewalle, Alain; Boneca, Ivo G.; Goujon, Jean-Michel; Werts, Catherine

2014-01-01

Background Leptospira (L.) interrogans are bacteria responsible for a worldwide reemerging zoonosis. Rodents carry L. interrogans asymptomatically in their kidneys and excrete bacteria in the urine, contaminating the environment. Humans get infected through skin contact and develop a mild or severe leptospirosis that may lead to renal failure and fibrosis. L. interrogans provoke an interstitial nephritis, but the induction of fibrosis caused by L. interrogans has not been studied in murine models. Innate immune receptors from the TLR and NLR families have recently been shown to play a role in the development and progression of tissue fibrosis in the lung, liver and kidneys under different pathophysiological situations. We recently showed that TLR2, TLR4, and NLRP3 receptors were crucial in the defense against leptospirosis. Moreover, infection of a human cell line with L. interrogans was shown to induce TLR2-dependent production of fibronectin, a component of the extracellular matrix. Therefore, we thought to assess the presence of renal fibrosis in L. interrogans infected mice and to analyze the contribution of some innate immune pathways in this process. Methodology/principal findings Here, we characterized by immunohistochemical studies and quantitative real-time PCR, a model of Leptospira-infected C57BL/6J mice, with chronic carriage of L. interrogans inducing mild renal fibrosis. Using various strains of transgenic mice, we determined that the renal infiltrates of T cells and, unexpectedly, TLR and NLR receptors, are not required to generate Leptospira-induced renal fibrosis. We also show that the iNOS enzyme, known to play a role in Leptospira-induced interstitial nephritis, also plays a role in the induction of renal fibrosis. Conclusion/significance To our knowledge, this work provides the first experimental murine model of sustained renal fibrosis induced by a chronic bacterial infection that may be peculiar, since it does not rely on TLR or NLR receptors. This model may prove useful to test future therapeutic strategies to combat Leptospira-induced renal lesions. PMID:24498450

Transcellular Pathways in Lymphatic Endothelial Cells Regulate Changes in Solute Transport by Fluid Stress.

PubMed

Triacca, Valentina; Güç, Esra; Kilarski, Witold W; Pisano, Marco; Swartz, Melody A

2017-04-28

The transport of interstitial fluid and solutes into lymphatic vessels is important for maintaining interstitial homeostasis and delivering antigens and soluble factors to the lymph node for immune surveillance. Transendothelial transport across lymphatic endothelial cells (LECs) is commonly considered to occur paracellularly, or between cell-cell junctions, and driven by local pressure and concentration gradients. However, emerging evidence suggests that LECs also play active roles in regulating interstitial solute balance and can scavenge and store antigens, raising the possibility that vesicular or transcellular pathways may be important in lymphatic solute transport. The aim of this study was to determine the relative importance of transcellular (vesicular) versus paracellular transport pathways by LECs and how mechanical stress (ie, fluid flow conditioning) alters either pathway. We demonstrate that transcellular transport mechanisms substantially contribute to lymphatic solute transport and that solute uptake occurs in both caveolae- and clathrin-coated vesicles. In vivo, intracelluar uptake of fluorescently labeled albumin after intradermal injection by LECs was similar to that of dermal dendritic cells. In vitro, we developed a method to differentially quantify intracellular solute uptake versus transendothelial transport by LECs. LECs preconditioned to 1 µm/s transmural flow demonstrated increased uptake and basal-to-apical solute transport, which could be substantially reversed by blocking dynamin-dependent vesicle formation. These findings reveal the importance of intracellular transport in steady-state lymph formation and suggest that LECs use transcellular mechanisms in parallel to the well-described paracellular route to modulate solute transport from the interstitium according to biomechanical cues. © 2017 American Heart Association, Inc.

Fibroblast growth factor represses Smad-mediated myofibroblast activation in aortic valvular interstitial cells

PubMed Central

Cushing, Melinda C.; Mariner, Peter D.; Liao, Jo-Tsu; Sims, Evan A.; Anseth, Kristi S.

2008-01-01

This study aimed to identify signaling pathways that oppose connective tissue fibrosis in the aortic valve. Using valvular interstitial cells (VICs) isolated from porcine aortic valve leaflets, we show that basic fibroblast growth factor (FGF-2) effectively blocks transforming growth factor-β1 (TGF-β1)-mediated myofibroblast activation. FGF-2 prevents the induction of α-smooth muscle actin (αSMA) expression and the exit of VICs from the cell cycle, both of which are hallmarks of myofibroblast activation. By blocking the activity of the Smad transcription factors that serve as the downstream nuclear effectors of TGF-β1, FGF-2 treatment inhibits fibrosis in VICs. Using an exogenous Smad-responsive transcriptional promoter reporter, we show that Smad activity is repressed by FGF-2, likely an effect of the fact that FGF-2 treatment prevents the nuclear localization of Smads in these cells. This appears to be a direct effect of FGF signaling through mitogen-activated protein kinase (MAPK) cascades as the treatment of VICs with the MAPK/extracellular regulated kinase (MEK) inhibitor U0126 acted to induce fibrosis and blocked the ability of FGF-2 to inhibit TGF-β1 signaling. Furthermore, FGF-2 treatment of VICs blocks the development of pathological contractile and calcifying phenotypes, suggesting that these pathways may be utilized in the engineering of effective treatments for valvular disease.—Cushing, M. C., Mariner, P. D., Liao, J. T., Sims, E. A., Anseth, K. S. Fibroblast growth factor represses Smad-mediated myofibroblast activation in aortic valvular interstitial cells. PMID:18218921

Flow through internal elastic lamina affects shear stress on smooth muscle cells (3D simulations).

PubMed

Tada, Shigeru; Tarbell, John M

2002-02-01

We describe a three-dimensional numerical simulation of interstitial flow through the medial layer of an artery accounting for the complex entrance condition associated with fenestral pores in the internal elastic lamina (IEL) to investigate the fluid mechanical environment around the smooth muscle cells (SMCs) right beneath the IEL. The IEL was modeled as an impermeable barrier to water flow except for the fenestral pores, which were assumed to be uniformly distributed over the IEL. The medial layer was modeled as a heterogeneous medium composed of a periodic array of cylindrical SMCs embedded in a continuous porous medium representing the interstitial proteoglycan and collagen matrix. Depending on the distance between the IEL bottom surface and the upstream end of the proximal layer of SMCs, the local shear stress on SMCs right beneath the fenestral pore could be more than 10 times higher than that on the cells far removed from the IEL under the conditions that the fenestral pore diameter and area fraction of pores were kept constant at 1.4 microm and 0.05, respectively. Thus these proximal SMCs may experience shear stress levels that are even higher than endothelial cells exposed to normal blood flow (order of 10 dyn/cm(2)). Furthermore, entrance flow through fenestral pores alters considerably the interstitial flow field in the medial layer over a spatial length scale of the order of the fenestral pore diameter. Thus the spatial gradient of shear stress on the most superficial SMC is noticeably higher than computed for endothelial cell surfaces.

Renal denervation prevents long-term sequelae of ischemic renal injury

PubMed Central

Kim, Jinu; Padanilam, Babu J.

2014-01-01

Signals that drive interstitial fibrogenesis after renal ischemia reperfusion injury remain undefined. Sympathetic activation is manifest even in the early clinical stages of chronic kidney disease and is directly related to disease severity. A role for renal nerves in renal interstitial fibrogenesis in the setting of ischemia reperfusion injury has not been studied. In male 129S1/SvImJ mice, ischemia reperfusion injury induced tubulointerstitial fibrosis as indicated by collagen deposition and profibrotic protein expression 4 to 16 days after the injury.. Leukocyte influx, proinflammatory protein expression, oxidative stress, apoptosis, and cell cycle arrest at G2/M phase were enhanced after ischemia reperfusion injury. Renal denervation at the time of injury or up to 1 day post-injury improved histology, decreased proinflammatory/profibrotic responses and apoptosis, and prevented G2/M cell cycle arrest in the kidney. Treatment with afferent nerve-derived calcitonin gene-related peptide (CGRP) or efferent nerve-derived norepinephrine in denervated and ischemia reperfusion injury-induced kidneys mimicked innervation, restored inflammation and fibrosis, induced G2/M arrest, and enhanced TGF-β1 activation. Blocking norepinephrine or CGRP function using respective receptor blockers prevented these effects. Consistent with the in vivo study, treatment with either norepinephrine or CGRP induced G2/M cell cycle arrest in HK-2 proximal tubule cells, whereas antagonists against their respective receptors prevented G2/M arrest. Thus, renal nerve stimulation is a primary mechanism and renal nerve-derived factors drive epithelial cell cycle arrest and the inflammatory cascade causing interstitial fibrogenesis after ischemia reperfusion injury. PMID:25207878

The replication of a mouse adapted SARS-CoV in a mouse cell line stably expressing the murine SARS-CoV receptor mACE2 efficiently induces the expression of proinflammatory cytokines

PubMed Central

Regla-Nava, Jose A.; Jimenez-Guardeño, Jose M.; Nieto-Torres, Jose L.; Gallagher, Thomas M.; Enjuanes, Luis; DeDiego, Marta L.

2013-01-01

Infection of conventional mice with a mouse adapted (MA15) severe acute respiratory syndrome (SARS) coronavirus (CoV) reproduces many aspects of human SARS such as pathological changes in lung, viremia, neutrophilia, and lethality. However, established mouse cell lines highly susceptible to mouse-adapted SARS-CoV infection are not available. In this work, efficiently transfectable mouse cell lines stably expressing the murine SARS-CoV receptor angiotensin converting enzyme 2 (ACE2) have been generated. These cells yielded high SARS-CoV-MA15 titers and also served as excellent tools for plaque assays. In addition, in these cell lines, SARS-CoV-MA15 induced the expression of proinflammatory cytokines and IFN-β, mimicking what has been observed in experimental animal models infected with SARS-CoV and SARS patients. These cell lines are valuable tools to perform in vitro studies in a mouse cell system that reflects the species used for in vivo studies of SARS-CoV-MA15 pathogenesis. PMID:23911968

Pathogenesis of Idiopathic Pulmonary Fibrosis

PubMed Central

Wolters, Paul J.; Collard, Harold R.; Jones, Kirk D.

2014-01-01

Idiopathic pulmonary fibrosis (IPF) is a fibrosing interstitial lung disease associated with aging that is characterized by the histopathological pattern of usual interstitial pneumonia. Although an understanding of the pathogenesis of IPF is incomplete, recent advances delineating specific clinical and pathologic features of IPF have led to better definition of the molecular pathways that are pathologically activated in the disease. In this review we highlight several of these advances, with a focus on genetic predisposition to IPF and how genetic changes, which occur primarily in epithelial cells, lead to activation of profibrotic pathways in epithelial cells. We then discuss the pathologic changes within IPF fibroblasts and the extracellular matrix, and we conclude with a summary of how these profibrotic pathways may be interrelated. PMID:24050627

MOLECULAR ANALYSIS OF MUTATIONS INDUCED BY MUTAGENS IN THE TK GENE OF MOUSE LYMPHOMA CELLS

EPA Science Inventory

MOLECULAR ANALYSIS OF MUTATIONS INDUCED BY BROMATE AND N- ETHYL-N-NITROSOUREA IN THE TK GENE OF MOUSE L YMPHOMA CELLS

The mouse lymphoma assay is widely used to identify chemical mutagens The Tk +1- gene located on an autosome in mouse lymphoma cells may recover a wide ra...

Glial molecular alterations with mouse brain development and aging: up-regulation of the Kir4.1 and aquaporin-4.

PubMed

Gupta, Rajaneesh Kumar; Kanungo, Madhusudan

2013-02-01

Glial cells, besides participating as passive supporting matrix, are also proposed to be involved in the optimization of the interstitial space for synaptic transmission by tight control of ionic and water homeostasis. In adult mouse brain, inwardly rectifying K+ (Kir4.1) and aquaporin-4 (AQP4) channels localize to astroglial endfeets in contact with brain microvessels and glutamate synapses, optimizing clearance of extracellular K(+) and water from the synaptic layers. However, it is still unclear whether there is an age-dependent difference in the expressions of Kir4.1 and AQP4 channels specifically during postnatal development and aging when various marked changes occur in brain and if these changes region specific. RT-PCR and immunoblotting was conducted to compare the relative expression of Kir4.1 and AQP4 mRNA and protein in the early and mature postnatal (0-, 15-, 45-day), adult (20-week), and old age (70-week) mice cerebral and cerebellar cortices. Expressions of Kir4.1 and AQP4 mRNA and protein are very low at 0-day. A pronounced and continuous increase was observed by mature postnatal ages (15-, 45-days). However, in the 70-week-old mice, expressions are significantly up-regulated as compared to 20-week-old mice. Both genes follow the same age-related pattern in both cerebral and cerebellar cortices. The time course and expression pattern suggests that Kir4.1 and AQP4 channels may play an important role in brain K(+) and water homeostasis in early postnatal weeks after birth and during aging.

Human mammary microenvironment better regulates the biology of human breast cancer in humanized mouse model.

PubMed

Zheng, Ming-Jie; Wang, Jue; Xu, Lu; Zha, Xiao-Ming; Zhao, Yi; Ling, Li-Jun; Wang, Shui

2015-02-01

During the past decades, many efforts have been made in mimicking the clinical progress of human cancer in mouse models. Previously, we developed a human breast tissue-derived (HB) mouse model. Theoretically, it may mimic the interactions between "species-specific" mammary microenvironment of human origin and human breast cancer cells. However, detailed evidences are absent. The present study (in vivo, cellular, and molecular experiments) was designed to explore the regulatory role of human mammary microenvironment in the progress of human breast cancer cells. Subcutaneous (SUB), mammary fat pad (MFP), and HB mouse models were developed for in vivo comparisons. Then, the orthotopic tumor masses from three different mouse models were collected for primary culture. Finally, the biology of primary cultured human breast cancer cells was compared by cellular and molecular experiments. Results of in vivo mouse models indicated that human breast cancer cells grew better in human mammary microenvironment. Cellular and molecular experiments confirmed that primary cultured human breast cancer cells from HB mouse model showed a better proliferative and anti-apoptotic biology than those from SUB to MFP mouse models. Meanwhile, primary cultured human breast cancer cells from HB mouse model also obtained the migratory and invasive biology for "species-specific" tissue metastasis to human tissues. Comprehensive analyses suggest that "species-specific" mammary microenvironment of human origin better regulates the biology of human breast cancer cells in our humanized mouse model of breast cancer, which is more consistent with the clinical progress of human breast cancer.

Interstitial lung disease associated with Equine Infectious Anemia Virus infection in horses

PubMed Central

2013-01-01

EIA (Equine Infectious Anemia) is a blood-borne disease primarily transmitted by haematophagous insects or needle punctures. Other routes of transmission have been poorly explored. We evaluated the potential of EIAV (Equine Infectious Anemia Virus) to induce pulmonary lesions in naturally infected equids. Lungs from 77 EIAV seropositive horses have been collected in Romania and France. Three types of lesions have been scored on paraffin-embedded lungs: lymphocyte infiltration, bronchiolar inflammation, and thickness of the alveolar septa. Expression of the p26 EIAV capsid (CA) protein has been evaluated by immunostaining. Compared to EIAV-negative horses, 52% of the EIAV-positive horses displayed a mild inflammation around the bronchioles, 22% had a moderate inflammation with inflammatory cells inside the wall and epithelial bronchiolar hyperplasia and 6.5% had a moderate to severe inflammation, with destruction of the bronchiolar epithelium and accumulation of smooth muscle cells within the pulmonary parenchyma. Changes in the thickness of the alveolar septa were also present. Expression of EIAV capsid has been evidenced in macrophages, endothelial as well as in alveolar and bronchiolar epithelial cells, as determined by their morphology and localization. To summarize, we found lesions of interstitial lung disease similar to that observed during other lentiviral infections such as FIV in cats, SRLV in sheep and goats or HIV in children. The presence of EIAV capsid in lung epithelial cells suggests that EIAV might be responsible for the broncho-interstitial damages observed. PMID:24289102

Interstitial lung disease associated with Equine Infectious Anemia Virus infection in horses.

PubMed

Bolfa, Pompei; Nolf, Marie; Cadoré, Jean-Luc; Catoi, Cornel; Archer, Fabienne; Dolmazon, Christine; Mornex, Jean-François; Leroux, Caroline

2013-12-01

EIA (Equine Infectious Anemia) is a blood-borne disease primarily transmitted by haematophagous insects or needle punctures. Other routes of transmission have been poorly explored. We evaluated the potential of EIAV (Equine Infectious Anemia Virus) to induce pulmonary lesions in naturally infected equids. Lungs from 77 EIAV seropositive horses have been collected in Romania and France. Three types of lesions have been scored on paraffin-embedded lungs: lymphocyte infiltration, bronchiolar inflammation, and thickness of the alveolar septa. Expression of the p26 EIAV capsid (CA) protein has been evaluated by immunostaining. Compared to EIAV-negative horses, 52% of the EIAV-positive horses displayed a mild inflammation around the bronchioles, 22% had a moderate inflammation with inflammatory cells inside the wall and epithelial bronchiolar hyperplasia and 6.5% had a moderate to severe inflammation, with destruction of the bronchiolar epithelium and accumulation of smooth muscle cells within the pulmonary parenchyma. Changes in the thickness of the alveolar septa were also present. Expression of EIAV capsid has been evidenced in macrophages, endothelial as well as in alveolar and bronchiolar epithelial cells, as determined by their morphology and localization. To summarize, we found lesions of interstitial lung disease similar to that observed during other lentiviral infections such as FIV in cats, SRLV in sheep and goats or HIV in children. The presence of EIAV capsid in lung epithelial cells suggests that EIAV might be responsible for the broncho-interstitial damages observed.

In-vitro analysis of early calcification in aortic valvular interstitial cells using Laser-Induced Breakdown Spectroscopy (LIBS).

PubMed

Davari, Seyyed Ali; Masjedi, Shirin; Ferdous, Zannatul; Mukherjee, Dibyendu

2018-01-01

Calcific aortic valve disease (CAVD) is a major cardiovascular disorder caused by osteogenic differentiation of valvular interstitial cells (VICs) within aortic valves. Conventional methods like colorimetric assays and histology fail to detect small calcium depositions during in-vitro VIC cultures. Laser-induced breakdown spectroscopy (LIBS) is a robust analytical tool used for inorganic materials characterizations, but relatively new to biomedical applications. We employ LIBS, for the first time, for quantitative in-vitro detection of calcium depositions in VICs at various osteogenic differentiation stages. VICs isolated from porcine aortic valves were cultured in osteogenic media over various days. Colorimetric calcium assays based on arsenazo dye and Von Kossa staining measured the calcium depositions within VICs. Simultaneously, LIBS signatures for Ca I (422.67 nm) atomic emission lines were collected for estimating calcium depositions in lyophilized VIC samples. Our results indicate excellent linear correlation between the calcium assay and our LIBS measurements. Furthermore, unlike the assay results, the LIBS results could resolve calcium signals from cell samples with as early as 2 days of osteogenic culture. Quantitatively, the LIBS measurements establish the limit of detection for calcium content in VICs to be ∼0.17±0.04 μg which indicates a 5-fold improvement over calcium assay. Picture: Quantitative LIBS enables in-vitro analysis for early stage detection of calcium deposition within aortic valvular interstitial cells (VICs). © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Renalase Protects against Renal Fibrosis by Inhibiting the Activation of the ERK Signaling Pathways

PubMed Central

Wu, Yiru; Wang, Liyan; Deng, Dai; Zhang, Qidong; Liu, Wenhu

2017-01-01

Renal interstitial fibrosis is a common pathway for the progression of chronic kidney disease (CKD) to end-stage renal disease. Renalase, acting as a signaling molecule, has been reported to have cardiovascular and renal protective effects. However, its role in renal fibrosis remains unknown. In this study, we evaluated the therapeutic efficacy of renalase in rats with complete unilateral ureteral obstruction (UUO) and examined the inhibitory effects of renalase on transforming growth factor-β1 (TGF-β1)-induced epithelial–mesenchymal transition (EMT) in human proximal renal tubular epithelial (HK-2) cells. We found that in the UUO model, the expression of renalase was markedly downregulated and adenoviral-mediated expression of renalase significantly attenuated renal interstitial fibrosis, as evidenced by the maintenance of E-cadherin expression and suppressed expression of α-smooth muscle actin (α-SMA), fibronectin and collagen-I. In vitro, renalase inhibited TGF-β1-mediated upregulation of α-SMA and downregulation of E-cadherin. Increased levels of Phospho-extracellular regulated protein kinases (p-ERK1/2) in TGF-β1-stimulated cells were reversed by renalase cotreatment. When ERK1 was overexpressed, the inhibition of TGF-β1-induced EMT and fibrosis mediated by renalase was attenuated. Our study provides the first evidence that renalase can ameliorate renal interstitial fibrosis by suppression of tubular EMT through inhibition of the ERK pathway. These results suggest that renalase has potential renoprotective effects in renal interstitial fibrosis and may be an effective agent for slowing CKD progression. PMID:28448446

Surface immunoglobulin on cultured foetal mouse thymocytes

PubMed Central

Haustein, D.; Mandel, T. E.

1979-01-01

Organ cultures of 14–15 day foetal mouse thymus were used as a source of non-neoplastic differentiating T cells, free of contaminating B cells. Viable cells obtained from such cultured thymuses were radio-iodinated and immunoglobulins (Ig) were isolated by co-precipitation from the 125I-labelled cell-surface proteins released during 1 h of incubation at 37°. The precipitates, both reduced and unreduced, were then analysed by polyacrylamide gel electrophoresis. The unreduced material migrated in a 5% gel as a single peak with a mobility slightly faster than that of mouse IgG. After reduction, however, two peaks were obtained (in a 10% gel), one corresponding in migration to mouse light chain and the other which moved slightly faster than mouse μ chain. This pattern was identical with that previously seen for both surface Ig of normal mouse thymocytes and neoplastic T lymphoma cells. Uncultured, 15 day foetal thymocytes did not produce any detectable co-precipitated cell surface material. Ig detected in these experiments was therefore produced during in vitro culture by non-neoplastic T cells in a system free of contaminating B cells and mouse serum proteins. PMID:315364

Xenogeneic spermatogenesis following transplantation of hamster germ cells to mouse testes.

PubMed

Ogawa, T; Dobrinski, I; Avarbock, M R; Brinster, R L

1999-02-01

It was recently demonstrated that rat spermatogenesis can occur in the seminiferous tubules of an immunodeficient recipient mouse after transplantation of testis cells from a donor rat. In the present study, hamster donor testis cells were transplanted to mice to determine whether xenogeneic spermatogenesis would result. The hamster diverged at least 16 million years ago from the mouse and produces spermatozoa that are larger than, and have a shape distinctly different from, those of the mouse. In four separate experiments with a total of 13 recipient mice, hamster spermatogenesis was identified in the testes of each mouse. Approximately 6% of the tubules examined demonstrated xenogeneic spermatogenesis. In addition, cryopreserved hamster testis cells generated spermatogenesis in recipients. However, abnormalities were noted in hamster spermatids and acrosomes in seminiferous tubules of recipient mice. Hamster spermatozoa were also found in the epididymis of recipient animals, but these spermatozoa generally lacked acrosomes, and heads and tails were separated. Thus, defects in spermiogenesis occur in hamster spermatogenesis in the mouse, which may reflect a limited ability of endogenous mouse Sertoli cells to support fully the larger and evolutionarily distant hamster germ cell. The generation of spermatogenesis from frozen hamster cells now adds this species to the mouse and rat, in which spermatogonial stem cells also can be cryopreserved. This finding has immediate application to valuable animals of many species, because the cells could be stored until suitable recipients are identified or culture techniques devised to expand the stem cell population.

Effects of different feeder layers on culture of bovine embryonic stem cell-like cells in vitro.

PubMed

Cong, Shan; Cao, Guifang; Liu, Dongjun

2014-12-01

To find a suitable feeder layer is important for successful culture conditions of bovine embryonic stem cell-like cells. In this study, expression of pluripotency-related genes OCT4, SOX2 and NANOG in bovine embryonic stem cell-like cells on mouse embryonic fibroblast feeder layers at 1-5 passages were monitored in order to identify the possible reason that bovine embryonic stem cell-like cells could not continue growth and passage. Here, we developed two novel feeder layers, mixed embryonic fibroblast feeder layers of mouse and bovine embryonic fibroblast at different ratios and sources including mouse fibroblast cell lines. The bovine embryonic stem cell-like cells generated in our study displayed typical stem cell morphology and expressed specific markers such as OCT4, stage-specific embryonic antigen 1 and 4, alkaline phosphatase, SOX2, and NANOG mRNA levels. When feeder layers and cell growth factors were removed, the bovine embryonic stem cell-like cells formed embryoid bodies in a suspension culture. Furthermore, we compared the expression of the pluripotent markers during bovine embryonic stem cell-like cell in culture on mixed embryonic fibroblast feeder layers, including mouse fibroblast cell lines feeder layers and mouse embryonic fibroblast feeder layers by real-time quantitative polymerase chain reaction. Results suggested that mixed embryonic fibroblast and sources including mouse fibroblast cell lines feeder layers were more suitable for long-term culture and growth of bovine embryonic stem cell-like cells than mouse embryonic fibroblast feeder layers. The findings may provide useful experimental data for the establishment of an appropriate culture system for bovine embryonic stem cell lines.

N-acetylcysteine attenuates the development of cardiac fibrosis and remodeling in a mouse model of heart failure.

PubMed

Giam, Beverly; Chu, Po-Yin; Kuruppu, Sanjaya; Smith, A Ian; Horlock, Duncan; Kiriazis, Helen; Du, Xiao-Jun; Kaye, David M; Rajapakse, Niwanthi W

2016-04-01

Oxidative stress plays a central role in the pathogenesis of heart failure. We aimed to determine whether the antioxidantN-acetylcysteine can attenuate cardiac fibrosis and remodeling in a mouse model of heart failure. Minipumps were implanted subcutaneously in wild-type mice (n = 20) and mice with cardiomyopathy secondary to cardiac specific overexpression of mammalian sterile 20-like kinase 1 (MST-1;n = 18) to administerN-acetylcysteine (40 mg/kg per day) or saline for a period of 8 weeks. At the end of this period, cardiac remodeling and function was assessed via echocardiography. Fibrosis, oxidative stress, and expression of collagen types I andIIIwere quantified in heart tissues. Cardiac perivascular and interstitial fibrosis were greater by 114% and 209%, respectively, inMST-1 compared to wild type (P ≤ 0.001). InMST-1 mice administeredN-acetylcysteine, perivascular and interstitial fibrosis were 40% and 57% less, respectively, compared to those treated with saline (P ≤ 0. 03). Cardiac oxidative stress was 119% greater inMST-1 than in wild type (P < 0.001) andN-acetylcysteine attenuated oxidative stress inMST-1 by 42% (P = 0.005). These data indicate thatN-acetylcysteine can blunt cardiac fibrosis and related remodeling in the setting of heart failure potentially by reducing oxidative stress. This study provides the basis to investigate the role ofN-acetylcysteine in chronic heart failure. © 2016 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, M; Finlay, J; Zhu, T

Purpose: Photosensitizer concentration during photodynamic therapy (PDT) is an important parameter for accurate dosimetry. Fluorescence signal can be used as a measure of photosensitizer concentration. Two methods of data acquisition were compared to an ex vivo study both for in vivo and phantom models. Methods: Fluorescence signal of commonly used photosensitizer benzoporphyrin derivative monoacid ring A (BPD) was obtained in phantoms and mouse tumors using an excitation light of 405 nm. Interstitial fluorescence signal was obtained using a side-cut fiber inserted into the tumor tissue of interest. Using a previously developed multi-fiber probe, tumor surface fluorescence measurements were also collected.more » Signals were calibrated according to optical phantoms with known sensitizer fluorescence. Optical properties for each sample were determined and the influence of different absorption and scattering properties on the fluorescence signals was investigated. Using single value decomposition of the spectra, the sensitizer concentration was determined using the two different measurement geometries. An ex vivo analysis was also performed for tumor samples to determine the sensitizer concentration. Results: The two fluorescence signals obtained from the surface multi-fiber probe and the interstitial measurements were compared and were corresponding for both phantoms and mouse models. The values obtained were comparable to the ex vivo measurements as well. Despite the difference in geometry, the surface probe measurements can still be used as a metric for determining the presence of sensitizer in small volume tumors. Conclusion: The multi-fiber contact probe can be used as a tool to measure fluorescence at the surface of the treatment area for PDT and predict sensitizer concentration throughout the tumor. This is advantageous in that the measurement does not damage any tissue. Future work will include investigating the dependence of these results on intratumor sensitizer distribution.« less

Non-invasive imaging of barriers to drug delivery in tumors.

PubMed

Hassid, Yaron; Eyal, Erez; Margalit, Raanan; Furman-Haran, Edna; Degani, Hadassa

2008-08-01

Solid tumors often develop high interstitial fluid pressure (IFP) as a result of increased water leakage and impaired lymphatic drainage, as well as changes in the extracellular matrix composition and elasticity. This high fluid pressure forms a barrier to drug delivery and hence, resistance to therapy. We have developed techniques based on contrast enhanced magnetic resonance imaging for mapping in tumors the vascular and transport parameters determining the delivery efficiency of blood borne substances. Sequential images are recorded during continuous infusion of a Gd-based contrast agent and analyzed according to a new physiological model, yielding maps of microvascular transfer constants, as well as outward convective interstitial transfer constants and steady state interstitial contrast agent concentrations both reflecting IFP distribution. We further demonstrated in non small cell human lung cancer xenografts the capability of our techniques to monitor in vivo collagenase induced increase in contrast agent delivery as a result of decreased IFP. These techniques can be applied to test drugs that affect angiogenesis and modulate interstitial fluid pressure and has the potential to be extended to cancer patients for assessing resistance to drug delivery.

Non-Invasive Imaging of Barriers to Drug Delivery in Tumors

PubMed Central

Hassid, Yaron; Eyal, Erez; Margalit, Raanan; Furman-Haran, Edna; Degani, Hadassa

2011-01-01

Solid tumors often develop high interstitial fluid pressure (IFP) as a result of increased water leakage and impaired lymphatic drainage, as well as changes in the extracellular matrix composition and elasticity. This high fluid pressure forms a barrier to drug delivery and hence, resistance to therapy. We have developed techniques based on contrast enhanced magnetic resonance imaging for mapping in tumors the vascular and transport parameters determining the delivery efficiency of blood borne substances. Sequential images are recorded during continuous infusion of a Gd-based contrast agent and analyzed according to a new physiological model, yielding maps of microvascular transfer constants, as well as outward convective interstitial transfer constants and steady state interstitial contrast agent concentrations both reflecting IFP distribution. We further demonstrated in non small cell human lung cancer xenografts the capability of our techniques to monitor in vivo collagenase induced increase in contrast agent delivery as a result of decreased IFP. These techniques can be applied to test drugs that affect angiogenesis and modulate interstitial fluid pressure and has the potential to be extended to cancer patients for assessing resistance to drug delivery. PMID:18638494

/sup 125/I interstitial implants in the RIF-1 murine flank tumor: an animal model for brachytherapy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bernstein, M.; Gutin, P.H.; Weaver, D.A.

1982-09-01

The development of a model for interstitial brachytherapy that uses high-activity, removable /sup 125/I sources in the RIF-1 murine flank tumor is reported. Experimental end points are clonogenic cell and tumor regrowth delay assays. For the clonogenic cell assay, interestitial radiation is delivered at total doses of 500-10,000 rad at dose rates of 0.9-2.7 rad/min to cells in annuli of tissue in the tumor. Dose-survival curves are characterized by an initial shoulder followed by a straight (exponential) portion, with D/sub 0/ similar to that of the curve obtained by external irradiation of the RIF-1 tumor in a self-contained cesium irradiatormore » at similar dose rates. Tumor regrowth curves have been obtained for minimum tumor doses of 500-5000 rad; marked tumor regression has been observed with minimum tumor doses as low as 2000 rad, but results are not as reproducible as the results obtained with the clonogenic cell assay.« less

THE LOCALIZATION OF HOMOLGOUS PLASMA PROTEINS IN THE TISSUES OF YOUNG HUMAN BEINGS AS DEMONSTRATED WITH FLUORESCENT ANTIBODIES

PubMed Central

Gitlin, David; Landing, Benjamin H.; Whipple, Ann

1953-01-01

Employing fluorescent antibodies for the detection of homologous plasma proteins in tissue sections, the distribution of plasma albumin, γ-globulin, β-lipoprotein, β1-metal-combining globulin, and fibrinogen has been studied in the tissues of infants and children. Plasma albumin, γ-globulin, and β1-metal-combining globulin were found in many cells and particularly cell nuclei, connective tissues and interstitial spaces, lymphatics, and blood vessels. β-Lipoprotein was found mostly in the nuclei of all cell types while fibrinogen was restricted largely to the lymphatic and vascular channels, connective tissues and the interstitial spaces. The widespread distribution of these plasma proteins in cells and connective tissues indicates the magnitude of the extravascular plasma protein pool which is in equilibrium with circulating plasma. Unfortunately, these results do not permit accurate localization of the sites of production of these plasma proteins, but do give some idea of their intimate relationship to the tissues. PMID:13022871

Mechanisms of leading edge protrusion in interstitial migration

PubMed Central

Wilson, Kerry; Lewalle, Alexandre; Fritzsche, Marco; Thorogate, Richard; Duke, Tom; Charras, Guillaume

2013-01-01

While the molecular and biophysical mechanisms underlying cell protrusion on two-dimensional substrates are well understood, our knowledge of the actin structures driving protrusion in three-dimensional environments is poor, despite relevance to inflammation, development and cancer. Here we report that, during chemotactic migration through microchannels with 5 μm × 5 μm cross-sections, HL60 neutrophil-like cells assemble an actin-rich slab filling the whole channel cross-section at their front. This leading edge comprises two distinct F-actin networks: an adherent network that polymerizes perpendicular to cell-wall interfaces and a ‘free’ network that grows from the free membrane at the cell front. Each network is polymerized by a distinct nucleator and, due to their geometrical arrangement, the networks interact mechanically. On the basis of our experimental data, we propose that, during interstitial migration, medial growth of the adherent network compresses the free network preventing its retrograde movement and enabling new polymerization to be converted into forward protrusion. PMID:24305616

Incorrect strain information for mouse cell lines: sequential influence of misidentification on sublines.

PubMed

Uchio-Yamada, Kozue; Kasai, Fumio; Ozawa, Midori; Kohara, Arihiro

2017-03-01

Misidentification or cross-contamination of cell lines can cause serious issues. Human cell lines have been authenticated by short tandem repeat profiling; however, mouse cell lines have not been adequately assessed. In this study, mouse cell lines registered with the JCRB cell bank were examined by simple sequence length polymorphism (SSLP) analysis to identify their strains. Based on comparisons with 7 major inbred strains, our results revealed their strains in 80 of 90 cell lines. However, 12 of the 80 cell lines (15%) were found to differ from registered information. Of them, 4 cell lines originated from the same mouse, which had been generated through mating between two different inbred strains. The genotype of the mouse sample had not been examined after the backcross, leading to strain misidentification in those cell lines. Although 8 other cell lines had been established as sublines of a BALB/c cell line, their SSLP profiles are similar to a Swiss cell line. This affects differences in genotypes between inbred and outbred strains. Because the use of inbred samples and interbreeding between strains are not involved in human materials, our results suggest that the cause and influence of misidentification in mouse cell lines are different from those in human.

Human amniotic epithelial cell feeder layers maintain mouse embryonic stem cell pluripotency via epigenetic regulation of the c-Myc promoter.

PubMed

Liu, Te; Cheng, Weiwei; Liu, Tianjin; Guo, Lihe; Huang, Qin; Jiang, Lizhen; Du, Xiling; Xu, Fuhui; Liu, Zhixue; Lai, Dongmei

2010-02-01

Mouse embryonic stem cells (ESCs) are typically cultured on a feeder layer of mouse embryonic fibroblasts (MEFs), with leukemia inhibitory factor (LIF) added to maintain them in an undifferentiated state. We have previously shown that human amniotic epithelial cells (hAECs) can be used as feeder cells to maintain mouse ESC pluripotency, but the mechanism for this is unknown. In the present study, we found that CpG islands 5' of the c-Myc gene remain hypomethylated in mouse ESCs cultured on hAECs. In addition, levels of acetylation of histone H3 and trimethylation of histone H3K4 in the c-Myc gene promoter were higher in ES cells cultured on hAECs than those in ES cells cultured on MEFs. These data suggested that hAECs can alter mouse ESC gene expression via epigenetic modification of c-Myc, providing a possible mechanism for the hAEC-induced maintenance of ESCs in an undifferentiated state.

Renal papillary tip extract stimulates BNP production and excretion from cardiomyocytes

PubMed Central

Hashizume, Ryotaro; Suzuki, Noboru; Ito, Rie; Yamanaka, Keiichi; Saito, Hiromitsu; Kiyonari, Hiroshi; Tawara, Isao; Kageyama, Yuki; Ogihara, Yoshito; Ali, Yusuf; Yamada, Norikazu; Katayama, Naoyuki; Ito, Masaaki

2018-01-01

Background Brain natriuretic peptide (BNP) is an important biomarker for patients with cardiovascular diseases, including heart failure, hypertension and cardiac hypertrophy. It is also known that BNP levels are relatively higher in patients with chronic kidney disease and no heart disease; however, the mechanism remains unclear. Methods and results We developed a BNP reporter mouse and occasionally found that this promoter was activated specifically in the papillary tip of the kidneys, and its activation was not accompanied by BNP mRNA expression. No evidence was found to support the existence of BNP isoforms or other nucleotide expression apart from BNP and tdTomato. The pBNP-tdTomato-positive cells were interstitial cells and were not proliferative. Unexpectedly, both the expression and secretion of BNP increased in primary cultured neonatal cardiomyocytes after their treatment with an extract of the renal papillary tip. Intraperitoneal injection of the extract of the papillary tips reduced blood pressure from 210 mmHg to 165 mmHg, the decrease being accompanied by an increase in serum BNP and urinary cGMP production in stroke-prone spontaneously hypertensive (SHR-SP) rats. Furthermore the induction of BNP by the papillary extract from rats with heart failure due to myocardial infarction was increased in cardiomyocytes. Conclusions These results suggested that the papillary tip express a substance that can stimulate BNP production and secretion from cardiomyocytes. PMID:29734386

Endothelial necrosis at 1h post-burn predicts progression of tissue injury

PubMed Central

Hirth, Douglas; McClain, Steve A.; Singer, Adam J.; Clark, Richard A.F.

2013-01-01

Burn injury progression has not been well characterized at the cellular level. To define burn injury progression in terms of cell death, histopathologic spatiotemporal relationships of cellular necrosis and apoptosis were investigated in a validated porcine model of vertical burn injury progression. Cell necrosis was identified by High Mobility Group Box 1 protein and apoptosis by Caspase 3a staining of tissue samples taken 1h, 24h and 7 days post-burn. Level of endothelial cell necrosis at 1h was predictive of level of apoptosis at 24h (Pearson's r=0.87) and of level of tissue necrosis at 7 days (Pearson's r=0.87). Furthermore, endothelial cell necrosis was deeper than interstitial cell necrosis at 1h (p<0.001). Endothelial cell necrosis at 1h divided the zone of injury progression (Jackson's zone of stasis) into an upper subzone with necrotic endothelial cells and initially viable adnexal and interstitial cells at 1h that progressed to necrosis by 24h, and a lower zone with initially viable endothelial cells at 1h, but necrosis and apoptosis of all cell types by 24h. Importantly, this spatiotemporal series of events and rapid progression resembles myocardial infarction and stroke, and implicates mechanisms of these injuries, ischemia, ischemia reperfusion, and programmed cell death, in burn progression. PMID:23627744

Mouse neuroblastoma cell-based model and the effect of epileptic events on calcium oscillations and neural spikes

NASA Astrophysics Data System (ADS)

Kim, Suhwan; Jung, Unsang; Baek, Juyoung; Lee, Sangwon; Jung, Woonggyu; Kim, Jeehyun; Kang, Shinwon

2013-01-01

Recently, mouse neuroblastoma cells have been considered as an attractive model for the study of human neurological and prion diseases, and they have been intensively used as a model system in different areas. For example, the differentiation of neuro2a (N2A) cells, receptor-mediated ion current, and glutamate-induced physiological responses have been actively investigated with these cells. These mouse neuroblastoma N2A cells are of interest because they grow faster than other cells of neural origin and have a number of other advantages. The calcium oscillations and neural spikes of mouse neuroblastoma N2A cells in epileptic conditions are evaluated. Based on our observations of neural spikes in these cells with our proposed imaging modality, we reported that they can be an important model in epileptic activity studies. We concluded that mouse neuroblastoma N2A cells produce epileptic spikes in vitro in the same way as those produced by neurons or astrocytes. This evidence suggests that increased levels of neurotransmitter release due to the enhancement of free calcium from 4-aminopyridine causes the mouse neuroblastoma N2A cells to produce epileptic spikes and calcium oscillations.

Using the mouse embryonic stem cell test (EST) to evaluate the embryotoxicity of haloacetic acids

EPA Science Inventory

The Embryonic Stem Cell Test (EST) is used to predict the embryotoxic potential of a test compound by combining the data from cytotoxicity assays in undifferentiated mouse embryonic stem (mES) cells and differentiated mouse cells with the data from a differentiation assay in mES ...

The roles of ERAS during cell lineage specification of mouse early embryonic development.

PubMed

Zhao, Zhen-Ao; Yu, Yang; Ma, Huai-Xiao; Wang, Xiao-Xiao; Lu, Xukun; Zhai, Yanhua; Zhang, Xiaoxin; Wang, Haibin; Li, Lei

2015-08-01

Eras encodes a Ras-like GTPase protein that was originally identified as an embryonic stem cell-specific Ras. ERAS has been known to be required for the growth of embryonic stem cells and stimulates somatic cell reprogramming, suggesting its roles on mouse early embryonic development. We now report a dynamic expression pattern of Eras during mouse peri-implantation development: its expression increases at the blastocyst stage, and specifically decreases in E7.5 mesoderm. In accordance with its expression pattern, the increased expression of Eras promotes cell proliferation through controlling AKT activation and the commitment from ground to primed state through ERK activation in mouse embryonic stem cells; and the reduced expression of Eras facilitates primitive streak and mesoderm formation through AKT inhibition during gastrulation. The expression of Eras is finely regulated to match its roles in mouse early embryonic development during which Eras expression is negatively regulated by the β-catenin pathway. Thus, beyond its well-known role on cell proliferation, ERAS may also play important roles in cell lineage specification during mouse early embryonic development. © 2015 The Authors.

Differentiation of sow and mouse ovarian granulosa cells exposed to zearalenone in vitro using RNA-seq gene expression.

PubMed

Zhang, Guo-Liang; Song, Jun-Lin; Zhou, Yi; Zhang, Rui-Qian; Cheng, Shun-Feng; Sun, Xiao-Feng; Qin, Guo-Qing; Shen, Wei; Li, Lan

2018-07-01

Zearalenone (ZEA), a natural contaminant found in feed, has been shown to have a negative impact on domestic animal reproduction, particularly in pigs. There are species-specific differences in the ZEA-induced toxicity pattern. Here, we investigated the different biological effects of ZEA exposure on porcine and mouse granulosa cells, using RNA-seq analysis. We treated murine and porcine granulosa cells with 10 μM and 30 μM ZEA during 72 h of culturing, in vitro. The results showed that 10 μM ZEA exposure significantly altered mitosis associated genes in porcine granulosa cells, while the same treatment significantly altered the steroidogenesis associated genes in mouse granulosa cells. Exposure to 30 μM ZEA resulted in significantly up-regulated expression of inflammatory related genes in porcine granulosa cells as well as the cancer related genes in mouse granulosa cells. Similarly, 30 μM ZEA exposure significantly decreased the expression of tumor suppressor factors in the mouse granulosa cells. Furthermore, immunofluorescence, RT-qPCR as well as western-blot analysis verified the different expression of related genes in ZEA exposed porcine and mouse granulosa cells. Collectively, these results illustrate the presence of species differences with regards to ZEA effects between porcine and mouse ovarian granulosa cells, in vitro. Copyright © 2018 Elsevier Inc. All rights reserved.

The replication of a mouse adapted SARS-CoV in a mouse cell line stably expressing the murine SARS-CoV receptor mACE2 efficiently induces the expression of proinflammatory cytokines.

PubMed

Regla-Nava, Jose A; Jimenez-Guardeño, Jose M; Nieto-Torres, Jose L; Gallagher, Thomas M; Enjuanes, Luis; DeDiego, Marta L

2013-11-01

Infection of conventional mice with a mouse adapted (MA15) severe acute respiratory syndrome (SARS) coronavirus (CoV) reproduces many aspects of human SARS such as pathological changes in lung, viremia, neutrophilia, and lethality. However, established mouse cell lines highly susceptible to mouse-adapted SARS-CoV infection are not available. In this work, efficiently transfectable mouse cell lines stably expressing the murine SARS-CoV receptor angiotensin converting enzyme 2 (ACE2) have been generated. These cells yielded high SARS-CoV-MA15 titers and also served as excellent tools for plaque assays. In addition, in these cell lines, SARS-CoV-MA15 induced the expression of proinflammatory cytokines and IFN-β, mimicking what has been observed in experimental animal models infected with SARS-CoV and SARS patients. These cell lines are valuable tools to perform in vitro studies in a mouse cell system that reflects the species used for in vivo studies of SARS-CoV-MA15 pathogenesis. Copyright © 2013 Elsevier B.V. All rights reserved.

Image-guided Interstitial Photodynamic Therapy for Squamous Cell Carcinomas: Preclinical investigation

PubMed Central

Sajisevi, Mirabelle; Rigual, Nestor R; Bellnier, David A.; Seshadri, Mukund

2014-01-01

Objective Photodynamic therapy (PDT) is a clinically approved minimally invasive treatment for cancer. In this preclinical study, using an imaging-guided approach, we examined the potential utility of PDT in the management of bulky squamous cell carcinomas (SCCs). Methods To mimic bulky oropharyngeal cancers seen in the clinical setting, intramuscular SCCs were established in six-to-eight week old female C3H mice. Animals were injected with the photosensitizer, 2-[hexyloxyethyl]-2-devinyl pyropheophorbide-a (HPPH; 0.4 μmol/kg, i.v.) and tumors were illuminated 24 hours post injection with 665 nm light. PDT as a single treatment modality was administered by surface illumination or by interstitial placement of fibers (iPDT). Magnetic resonance imaging was used to guide treatment and assess tumor response to PDT along with correlative histopathologic assessment. Results Interstitial HPPH-PDT resulted in a marked change on T2 maps 24 hours post treatment compared to untreated controls or transcutaneous illumination. Corresponding apparent diffusion coefficient maps also showed hyperintense areas in tumors following iPDT suggestive of effective photodynamic cell kill. Histologic sections (H&E) confirmed presence of extensive tumor necrosis following iPDT. Conclusions These results highlight the potential utility of PDT in the treatment of bulky oropharyngeal cancers. The findings of our study also demonstrate the utility of MRI as a non-invasive tool for mapping of early tissue response to PDT. PMID:25750858

Cinnamaldehyde impairs high glucose-induced hypertrophy in renal interstitial fibroblasts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chao, Louis Kuoping; Chang, W.-T.; Shih, Y.-W.

2010-04-15

Cinnamaldehyde is a major and a bioactive compound isolated from the leaves of Cinnamomum osmophloeum kaneh. To explore whether cinnamaldehyde was linked to altered high glucose (HG)-mediated renal tubulointerstitial fibrosis in diabetic nephropathy (DN), the molecular mechanisms of cinnamaldehyde responsible for inhibition of HG-induced hypertrophy in renal interstitial fibroblasts were examined. We found that cinnamaldehyde caused inhibition of HG-induced cellular mitogenesis rather than cell death by either necrosis or apoptosis. There were no changes in caspase 3 activity, cleaved poly(ADP-ribose) polymerase (PARP) protein expression, and mitochondrial cytochrome c release in HG or cinnamaldehyde treatments in these cells. HG-induced extracellular signal-regulatedmore » kinase (ERK)/c-Jun N-terminal kinase (JNK)/p38 mitogen-activated protein kinase (MAPK) (but not the Janus kinase 2/signal transducers and activators of transcription) activation was markedly blocked by cinnamaldehyde. The ability of cinnamaldehyde to inhibit HG-induced hypertrophy was verified by the observation that it significantly decreased cell size, cellular hypertrophy index, and protein levels of collagen IV, fibronectin, and alpha-smooth muscle actin (alpha-SMA). The results obtained in this study suggest that cinnamaldehyde treatment of renal interstitial fibroblasts that have been stimulated by HG reduces their ability to proliferate and hypertrophy through mechanisms that may be dependent on inactivation of the ERK/JNK/p38 MAPK pathway.« less

Physiological water model development

NASA Technical Reports Server (NTRS)

Doty, Susan

1993-01-01

The water of the human body can be categorized as existing in two main compartments: intracellular water and extracellular water. The intracellular water consists of all the water within the cells and constitutes over half of the total body water. Since red blood cells are surrounded by plasma, and all other cells are surrounded by interstitial fluid, the intracellular compartment has been subdivided to represent these two cell types. The extracellular water, which includes all of the fluid outside of the cells, can be further subdivided into compartments which represent the interstitial fluid, circulating blood plasma, lymph, and transcellular water. The interstitial fluid surrounds cells outside of the vascular system whereas plasma is contained within the blood vessels. Avascular tissues such as dense connective tissue and cartilage contain interstitial water which slowly equilibrates with tracers used to determine extracellular fluid volume. For this reason, additional compartments are sometimes used to represent these avascular tissues. The average size of each compartment, in terms of percent body weight, has been determined for adult males and females. These compartments and the forces which cause flow between them are presented. The kidneys, a main compartment, receive about 25 percent of the cardiac output and filters out a fluid similar to plasma. The composition of this filtered fluid changes as it flows through the kidney tubules since compounds are continually being secreted and reabsorbed. Through this mechanism, the kidneys eliminate wastes while conserving body water, electrolytes, and metabolites. Since sodium accounts for over 90 percent of the cations in the extracellular fluid, and the number of cations is balanced by the number of anions, considering the renal handling sodium and water only should sufficiently describe the relationship between the plasma compartment and kidneys. A kidney function model is presented which has been adapted from a previous model of normal renal function in man. To test the validity of the proposed kidney model, results predicted by the model will be compared to actual data involving injected or ingested fluids and subsequent urine flow rates. Comparison of the model simulation to actual data following the ingestion of 1 liter of water is shown. The model simulation is also shown with actual data following the intravenous infusion of hypertonic saline.

Preventing light-induced degradation in multicrystalline silicon

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lindroos, J., E-mail: jeanette.lindroos@aalto.fi; Boulfrad, Y.; Yli-Koski, M.

2014-04-21

Multicrystalline silicon (mc-Si) is currently dominating the silicon solar cell market due to low ingot costs, but its efficiency is limited by transition metals, extended defects, and light-induced degradation (LID). LID is traditionally associated with a boron-oxygen complex, but the origin of the degradation in the top of the commercial mc-Si brick is revealed to be interstitial copper. We demonstrate that both a large negative corona charge and an aluminum oxide thin film with a built-in negative charge decrease the interstitial copper concentration in the bulk, preventing LID in mc-Si.

Preventing light-induced degradation in multicrystalline silicon

NASA Astrophysics Data System (ADS)

Lindroos, J.; Boulfrad, Y.; Yli-Koski, M.; Savin, H.

2014-04-01

Multicrystalline silicon (mc-Si) is currently dominating the silicon solar cell market due to low ingot costs, but its efficiency is limited by transition metals, extended defects, and light-induced degradation (LID). LID is traditionally associated with a boron-oxygen complex, but the origin of the degradation in the top of the commercial mc-Si brick is revealed to be interstitial copper. We demonstrate that both a large negative corona charge and an aluminum oxide thin film with a built-in negative charge decrease the interstitial copper concentration in the bulk, preventing LID in mc-Si.

[Effect of CsA bleomycin-induced interstitial pulmonary disease in mice].

PubMed

Ren, Ying; Yang, Hui; Zhu, Ping; Fan, Chun-mei; Wang, Yan-hong; Li, Jia; Liu, Hui

2012-03-01

To observe the therapeutic effect of cyclosporine A (CsA) on bleomycin (BLM) induced pulmonary fibrosis and to investigate its mechanism. One hundred and twenty C57BL/6 female mice were divided randomly into five groups: BLM model group, control saline group, CsA30 mg treatment group, CsA50 mg treatment group and control treatment group. Treatment groups and model groups were administrated BLM intratracheally to induce interstitial pulmonary disease model, with control saline group administrated with equal volume of normal saline instead. Mice in treatment groups were intraperitoneal injected with CsA, while control treatment group were injected with equal volume of normal saline instead. On the 4th, 7th and 14th day after administration, 8 mice of each group were sacrificed, and the peripheral blood was obtained to count total leucocytes with counting chamber and quantify CD4(+); T cells, CD14(+); monocytes and CD19(+); B cells by flow cytometry (FCM). Bronchoalveolar levage fluid was harvested for cell counting and Giemsa staining. Lung tissues were harvested for immunohistochemical staining and pathological examination. The quantity of total leucocyte was higher in BLM model group than those in control saline group.The proportion of CD14(+); T cells and CD19(+);B cells in BLM model group were increased markedly than those in control saline group on the 4th, 7th and 14th day post BLM. With CsA treatment, The proportion of CD14(+); T cells was lower than BLM model group at the same time point, especially on the 4th day. The proportion of CD19(+); B cells were significantly lower than those of BLM model group at the same time point(7 d, 14 d). The total and classification of cells of BLM model group were increased markedly than those in control saline group, and decreased obviously in the treatment groups at the same time point. Examination of lung tissues: With the prolonged time of BLM administration, it showed wider alveolar septum, more collagen deposition, as well as more infiltrating inflammatory cells which consisted of generous lymphocyte and few mononuclear macrophages than those in saline control group. With the prolonged time of CsA injection, the interstitial pulmonary inflammation was remissive, and there was less fibroblast infiltration and collagen deposition in pulmonary interstitium and periphery of bronchiole. Alveolar epithelial cells, bronchiolar epithelial cells, mononuclear macrophages, neutrophils and lymphocytes were demonstrated to express CD147, there was higher CD147 expression in BLM model group than those in CsA treatment groups. CsA may heal BLM induced interstitial pulmonary disease by blocking CD147-CypA interaction, then decreasing chemotaxis for the immunocyte, and reducing migration of immunocytes to the lung and collagen deposition in the lung.

The terminator mouse: salvation for primary cell culture.

PubMed

Kabgani, Nazanin; Moeller, Marcus J

2013-11-01

The Terminator had to come back from the future already several times in an effort to bring salvation to mankind. In the present issue of Kidney International, Guo et al. brought us a novel transgenic mouse model: the terminator mouse. This highly elegant mouse may facilitate significantly the derivation of primary cultures of a specific cell type from a tissue containing multiple cell populations.

Luteinizing hormone (LH) blood test

MedlinePlus

ICSH - blood test; Luteinizing hormone - blood test; Interstitial cell stimulating hormone - blood test ... to temporarily stop medicines that may affect the test results. Be sure to tell your provider about ...

Response of a mouse hybridoma cell line to heat shock, agitation, and sparging

NASA Technical Reports Server (NTRS)

Passini, Cheryl A.; Goochee, Charles F.

1989-01-01

A mouse hybridoma cell line is used as a model system for studying the effect of environmental stress on attachment-independent mammalian cells. The full time course of recovery for a mouse hybridoma cell line from both a mild and intermediate heat shock is examined. The pattern of intracellular synthesis is compared for actively growing, log phase cells and nondividing, stationary phase cells.

[Low expression of activin A in mouse and human embryonic teratocarcinoma cells].

PubMed

Gordeeva, O F

2014-01-01

TGFP3 family factors play an important role in regulating the balance of self-renewal and differentiation of mouse and human pluripotent stem and embryonic teratocarcinoma cells. The expression patterns of TGFbeta family signaling ligands and functional roles of these signaling pathways differ significantly in mouse and human embryonic stem cells, but the activity and functional role of these factors in mouse and human embryonic teratocarcinoma cells were not sufficiently investigated. Comparative quantitative real-time PCR analysis of the expression of TGF@[beta] family factors in mouse embryonic stem, embryonic germ, and embryonic teratocarcinoma cells showed that embryonic teratocarcinoma cells express lower ActivinA than pluripotent stem cells but similar levels of factors Nodal, Lefty 1, TGFbeta1, BMP4, and GDF3. In human nullipotent embryonic teratocarcinoma PA-1 cells, most factors of the TGFbeta family (ACTIVINA, NODAL, LEFTY 1, BMP4, and GDF3) are expressed at lower levels than in human embryonic stem cells: Thus, in mouse and human nullipotent teratocarcinoma cells, theexpression of ActivinA is significantly reduced com- pared ivith embryonic stem cells. Presumably, these differences may be associated with changes in the functional activity of the respective signaling pathways and deregulation of proliferative and antiproliferative mechanisms in embryonic teratocarcinoma cells.

In vivo measurement of apolipoprotein E from the brain interstitial fluid using microdialysis

PubMed Central

2013-01-01

Background The APOE4 allele variant is the strongest known genetic risk factor for developing late-onset Alzheimer’s disease. The link between apolipoprotein E (apoE) and Alzheimer’s disease is likely due in large part to the impact of apoE on the metabolism of amyloid β (Aβ) within the brain. Manipulation of apoE levels and lipidation within the brain has been proposed as a therapeutic target for the treatment of Alzheimer’s disease. However, we know little about the dynamic regulation of apoE levels and lipidation within the central nervous system. We have developed an assay to measure apoE levels in the brain interstitial fluid of awake and freely moving mice using large molecular weight cut-off microdialysis probes. Results We were able to recover apoE using microdialysis from human cerebrospinal fluid (CSF) in vitro and mouse brain parenchyma in vivo. Microdialysis probes were inserted into the hippocampus of wild-type mice and interstitial fluid was collected for 36 hours. Levels of apoE within the microdialysis samples were determined by ELISA. The levels of apoE were found to be relatively stable over 36 hours. No apoE was detected in microdialysis samples from apoE KO mice. Administration of the RXR agonist bexarotene increased ISF apoE levels while ISF Aβ levels were decreased. Extrapolation to zero-flow analysis allowed us to determine the absolute recoverable concentration of apoE3 in the brain ISF of apoE3 KI mice. Furthermore, analysis of microdialysis samples by non-denaturing gel electrophoresis determined lipidated apoE particles in microdialysis samples were consistent in size with apoE particles from CSF. Finally, we found that the concentration of apoE in the brain ISF was dependent upon apoE isoform in human apoE KI mice, following the pattern apoE2>apoE3>apoE4. Conclusions We are able to collect lipidated apoE from the brain of awake and freely moving mice and monitor apoE levels over the course of several hours from a single mouse. Our technique enables assessment of brain apoE dynamics under physiological and pathophysiological conditions and in response to therapeutic interventions designed to affect apoE levels and lipidation within the brain. PMID:23601557

Increased Risk of Interstitial Lung Disease in Children with a Single R288K Variant of ABCA3

PubMed Central

Wittmann, Thomas; Frixel, Sabrina; Höppner, Stefanie; Schindlbeck, Ulrike; Schams, Andrea; Kappler, Matthias; Hegermann, Jan; Wrede, Christoph; Liebisch, Gerhard; Vierzig, Anne; Zacharasiewicz, Angela; Kopp, Matthias Volkmar; Poets, Christian F; Baden, Winfried; Hartl, Dominik; van Kaam, Anton H; Lohse, Peter; Aslanidis, Charalampos; Zarbock, Ralf; Griese, Matthias

2016-01-01

The ABCA3 gene encodes a lipid transporter in type II pneumocytes critical for survival and normal respiratory function. The frequent ABCA3 variant R288K increases the risk for neonatal respiratory distress syndrome among term and late preterm neonates, but its role in children’s interstitial lung disease has not been studied in detail. In a retrospective cohort study of 228 children with interstitial lung disease related to the alveolar surfactant system, the frequency of R288K was assessed and the phenotype of patients carrying a single R288K variant further characterized by clinical course, lung histology, computed tomography and bronchoalveolar lavage phosphatidylcholine PC 32:0. Cell lines stably transfected with ABCA3-R288K were analyzed for intracellular transcription, processing and targeting of the protein. ABCA3 function was assessed by detoxification assay of doxorubicin, and the induction and volume of lamellar bodies. We found nine children with interstitial lung disease carrying a heterozygous R288K variant, a frequency significantly higher than in the general Caucasian population. All identified patients had neonatal respiratory insufficiency, recovered and developed chronic interstitial lung disease with intermittent exacerbations during early childhood. In vitro analysis showed normal transcription, processing, and targeting of ABCA3-R288K, but impaired detoxification function and smaller lamellar bodies. We propose that the R288K variant can underlie interstitial lung disease in childhood due to reduced function of ABCA3, demonstrated by decelerated detoxification of doxorubicin, reduced PC 32:0 content and decreased lamellar body volume. PMID:26928390

Association between renal iron accumulation and renal interstitial fibrosis in a rat model of chronic kidney disease.

PubMed

Naito, Yoshiro; Fujii, Aya; Sawada, Hisashi; Oboshi, Makiko; Iwasaku, Toshihiro; Okuhara, Yoshitaka; Morisawa, Daisuke; Eguchi, Akiyo; Hirotani, Shinichi; Masuyama, Tohru

2015-07-01

Iron accumulation is associated with the pathophysiology of chronic kidney disease (CKD). Renal fibrosis is a final common feature that contributes to the progression of CKD; however, little is known about the association between renal iron accumulation and renal interstitial fibrosis in CKD. Here we investigate the effects of iron chelation on renal interstitial fibrosis in a rat model of CKD. CKD was induced by 5/6 nephrectomy in Sprague-Dawley rats. At 8 weeks after operation, 5/6 nephrectomized rats were administered an oral iron chelator, deferasirox (DFX), in chow for 8 weeks. Other CKD rats were given a normal diet. Sham-operative rats given a normal diet served as a control. CKD rats exhibited hypertension, glomerulosclerosis and renal interstitial fibrosis. Iron chelation with DFX did not change hypertension and glomerulosclerosis; however, renal interstitial fibrosis was attenuated in CKD rats. Consistent with these findings, renal gene expression of collagen type III and transforming growth factor-β was increased in CKD rats compared with the controls, while iron chelation suppressed these increments. In addition, a decrease in vimentin along an increase in E-cadherin in renal gene expression was observed in CKD rats with iron chelation. CKD rats also showed increased CD68-positive cells in the kidney, whereas its increase was attenuated by iron deprivation. Similarly, increased renal gene expression of CD68, tumor necrosis factor-α and monocyte chemoattractant protein-1 was suppressed in CKD rats with iron chelation. Renal iron accumulation seems to be associated with renal interstitial fibrosis in a rat model of CKD.

Interstitial radiation therapy for carcinoma of the penis using iridium 192 wires: the Henri Mondor experience (1970-1979)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mazeron, J.J.; Langlois, D.; Lobo, P.A.

1984-10-01

From 1970 to 1979, a group of 50 patients was treated for squamous cell carcinoma of the penis by interstitial irradiation using an afterloading technique and iridium 192 wires. The group included 9 patients with T1 tumors, 27 with T2 tumors, and 14 with T3 tumors. Forty-five patients presented with no metastatic inguinal nodes (NO), 3 patients with N1 nodes, and 2 patients had N3 nodes. After treatment, 11 patients (1 T1, 6 T2 and 4 T3) developed local recurrences. Three patients developed post-therapeutic necrosis which necessitated partial amputation in 2 cases. Eight patients developed post-therapeutic urethral stenosis, which requiredmore » surgical treatment in three of the cases. Twenty-one percent of the patients died of their disease. The authors advocate interstitial irradiation using iridium 192 wires for the treatment of non-infiltrating or moderately infiltrating squamous cell carcinoma of the penis in which the largest dimension does no exceed 4 cm. When regular follow-up can be assurred, it is reasonable to forgo prophylactic treatment of the inguinal nodes in patients presenting without groin metastasis.« less

Surface Antigens Common to Mouse Cleavage Embryos and Primitive Teratocarcinoma Cells in Culture

PubMed Central

Artzt, Karen; Dubois, Philippe; Bennett, Dorothea; Condamine, Hubert; Babinet, Charles; Jacob, François

1973-01-01

Syngeneic antisera have been produced in mouse strain 129/Sv-CP males against the primitive cells of teratocarcinoma. These sera react specifically with the primitive cells and are negative on various types of differentiated teratoma cells derived from the same original tumor. They are negative on all other mouse cells tested, with the exception of male germ cells and cleavage-stage embryos. Thus, teratoma cells possess cell-surface antigens in common with normal cleavage-stage embryos. Images PMID:4355379

Selective binding and transcytosis of Ulex europaeus 1 lectin by mouse Peyer's patch M-cells in vivo.

PubMed

Clark, M A; Jepson, M A; Simmons, N L; Hirst, B H

1995-12-01

The in vivo interaction of the lectin Ulex europaeus agglutinin 1 with mouse Peyer's patch follicle-associated epithelial cells was studied in the mouse Peyer's patch gut loop model by immunofluorescence and electron microscopy. The lectin targets to mouse Peyer's patch M-cells and is rapidly endocytosed and transcytosed. These processes are accompanied by morphological changes in the M-cell microvilli and by redistribution of polymerised actin. The demonstration of selective binding and uptake of a lectin by intestinal M-cells in vivo suggests that M-cell-specific surface glycoconjugates might act as receptors for the selective adhesion/uptake of microorganisms.

Intermediate-grade mammary gland adenocarcinoma in an 18-year-old female black leopard (Panthera pardus) with acute pancreatic necrosis and chronic interstitial nephropathy.

PubMed

Nakamura, Misato; Yoshida, Toshinori; Eguchi, Ayumi; Inohana, Mari; Nagahara, Rei; Shiraki, Ayako; Ito, Nanao; Shibutani, Makoto

2018-03-02

An 18-year-old female black leopard (Panthera pardus) showed renal failure, leukocytosis and presence of subcutaneous masses in the lower abdominal region and right shoulder; she eventually died. Histopathological observations included a mammary gland carcinoma with comedo, solid and tubulopapillary patterns in subcutaneous tissue, and highly proliferated tumor cells in systemic organs. The tumor cells were positive for cytokeratin AE1/AE3. The mammary gland tumor was diagnosed as intermediate-grade adenocarcinoma, based on a previously reported histological grading system of feline mammary carcinomas. Chronic interstitial nephritis was estimated to have been ongoing for 5 years, whilst acute necrotic pancreatitis in relation to tumor metastasis could have been the cause of death.

Disorders of interstitial cells of Cajal in a neonate with segmental dilatation of the intestine.

PubMed

Okada, Tadao; Sasaki, Fumiaki; Honda, Shohei; Cho, Kazutosi; Matsuno, Yoshihiro; Itoh, Tomoo; Kubota, Kanako C; Todo, Satoru

2010-06-01

Localized myopathy of the muscular layers may be an important factor contributing to segmental dilatation of the intestine (SDI). Only one report has described SDI of the jejunum in a neonate showing no abnormality of the interstitial cells of Cajal (ICC). The present report describes the very rare case of a neonatal girl with segmental dilatation of the distal duodenum and proximal jejunum with irregular arrangements of Auerbach's plexus and ICC and the successful surgical treatment of SDI. We review the literature on this type of relationship between abnormality of ICC and SDI and discuss the clinical features of this complication. Furthermore, the possible neuropathic cause of SDI complicated with disorders of ICC was explored in this report. Copyright 2010 Elsevier Inc. All rights reserved.

A Functional Analysis on the Interspecies Interaction between Mouse LFA-1 and Human Intercellular Adhesion Molecule-1 at the Cell Level

PubMed Central

Núñez, David; Comas, Laura; Lanuza, Pilar M.; Sánchez-Martinez, Diego; Pérez-Hernández, Marta; Catalán, Elena; Domingo, María Pilar; Velázquez-Campoy, Adrián; Pardo, Julián; Gálvez, Eva M.

2017-01-01

The interaction between intercellular adhesion molecules (ICAM) and the integrin leukocyte function-associated antigen-1 (LFA-1) is crucial for the regulation of several physiological and pathophysiological processes like cell-mediated elimination of tumor or virus infected cells, cancer metastasis, or inflammatory and autoimmune processes. Using purified proteins it was reported a species restriction for the interaction of ICAM-1 and LFA-1, being mouse ICAM-1 able to interact with human LFA-1 but not human ICAM-1 with mouse LFA-1. However, in vivo results employing tumor cells transfected with human ICAM-1 suggest that functionally mouse LFA-1 can recognize human ICAM-1. In order to clarify the interspecies cross-reactivity of the ICAM-1/LFA-1 interaction, we have performed functional studies analyzing the ability of human soluble ICAM-1 and human/mouse LFA-1 derived peptides to inhibit cell aggregation and adhesion as well as cell-mediated cytotoxicity in both mouse and human systems. In parallel, the affinity of the interaction between mouse LFA-1-derived peptides and human ICAM-1 was determined by calorimetry assays. According to the results obtained, it seems that human ICAM-1 is able to interact with mouse LFA-1 on intact cells, which should be taking into account when using humanized mice and xenograft models for the study of immune-related processes. PMID:29312326

A Functional Analysis on the Interspecies Interaction between Mouse LFA-1 and Human Intercellular Adhesion Molecule-1 at the Cell Level.

PubMed

Núñez, David; Comas, Laura; Lanuza, Pilar M; Sánchez-Martinez, Diego; Pérez-Hernández, Marta; Catalán, Elena; Domingo, María Pilar; Velázquez-Campoy, Adrián; Pardo, Julián; Gálvez, Eva M

2017-01-01

The interaction between intercellular adhesion molecules (ICAM) and the integrin leukocyte function-associated antigen-1 (LFA-1) is crucial for the regulation of several physiological and pathophysiological processes like cell-mediated elimination of tumor or virus infected cells, cancer metastasis, or inflammatory and autoimmune processes. Using purified proteins it was reported a species restriction for the interaction of ICAM-1 and LFA-1, being mouse ICAM-1 able to interact with human LFA-1 but not human ICAM-1 with mouse LFA-1. However, in vivo results employing tumor cells transfected with human ICAM-1 suggest that functionally mouse LFA-1 can recognize human ICAM-1. In order to clarify the interspecies cross-reactivity of the ICAM-1/LFA-1 interaction, we have performed functional studies analyzing the ability of human soluble ICAM-1 and human/mouse LFA-1 derived peptides to inhibit cell aggregation and adhesion as well as cell-mediated cytotoxicity in both mouse and human systems. In parallel, the affinity of the interaction between mouse LFA-1-derived peptides and human ICAM-1 was determined by calorimetry assays. According to the results obtained, it seems that human ICAM-1 is able to interact with mouse LFA-1 on intact cells, which should be taking into account when using humanized mice and xenograft models for the study of immune-related processes.

Establishment of mouse neuron and microglial cell co-cultured models and its action mechanism.

PubMed

Zhang, Bo; Yang, Yunfeng; Tang, Jun; Tao, Yihao; Jiang, Bing; Chen, Zhi; Feng, Hua; Yang, Liming; Zhu, Gang

2017-06-27

The objective of this study is to establish a co-culture model of mouse neurons and microglial cells, and to analyze the mechanism of action of oxygen glucose deprivation (OGD) and transient oxygen glucose deprivation (tOGD) preconditioning cell models. Mouse primary neurons and BV2 microglial cells were successfully cultured, and the OGD and tOGD models were also established. In the co-culture of mouse primary neurons and microglial cells, the cell number of tOGD mouse neurons and microglial cells was larger than the OGD cell number, observed by a microscope. CCK-8 assay result showed that at 1h after treatment, the OD value in the control group is lower compared to all the other three groups (P < 0.05). The treatment group exhibited the highest OD value among the four groups. The results observed at 5h were consistent with the results at 1 h. Flow cytometry results showed that at 1h after treatment the apoptosis percentages is higher in the control group compared to other three groups (P < 0.05). Mouse brain tissues were collected and primary neurons cells were cultured. In the meantime mouse BV2 microglia cells were cultured. Two types of cells were co-cultured, and OGD and tOGD cell models were established. There were four groups in the experiment: control group (OGD), treatment group (tOGD+OGD), placebo group (tOGD+OGD+saline) and minocycline intervention group (tOGD+OGD+minocycline). CCK-8 kit was used to detect cell viability and flow cytometry was used to detect apoptosis. In this study, mouse primary neurons and microglial cells were co-cultured. The OGD and tOGD models were established successfully. tOGD was able to effectively protect neurons and microglial cells from damage, and inhibit the apoptosis caused by oxygen glucose deprivation.

Generation and gene expression profiling of 48 transcription-factor-inducible mouse embryonic stem cell lines.

PubMed

Yamamizu, Kohei; Sharov, Alexei A; Piao, Yulan; Amano, Misa; Yu, Hong; Nishiyama, Akira; Dudekula, Dawood B; Schlessinger, David; Ko, Minoru S H

2016-05-06

Mouse embryonic stem cells (ESCs) can differentiate into a wide range - and possibly all cell types in vitro, and thus provide an ideal platform to study systematically the action of transcription factors (TFs) in cell differentiation. Previously, we have generated and analyzed 137 TF-inducible mouse ESC lines. As an extension of this "NIA Mouse ESC Bank," we generated and characterized 48 additional mouse ESC lines, in which single TFs in each line could be induced in a doxycycline-controllable manner. Together, with the previous ESC lines, the bank now comprises 185 TF-manipulable ESC lines (>10% of all mouse TFs). Global gene expression (transcriptome) profiling revealed that the induction of individual TFs in mouse ESCs for 48 hours shifts their transcriptomes toward specific differentiation fates (e.g., neural lineages by Myt1 Isl1, and St18; mesodermal lineages by Pitx1, Pitx2, Barhl2, and Lmx1a; white blood cells by Myb, Etv2, and Tbx6, and ovary by Pitx1, Pitx2, and Dmrtc2). These data also provide and lists of inferred target genes of each TF and possible functions of these TFs. The results demonstrate the utility of mouse ESC lines and their transcriptome data for understanding the mechanism of cell differentiation and the function of TFs.

Compositional changes in lipid microdomains of air-blood barrier plasma membranes in pulmonary interstitial edema.

PubMed

Palestini, Paola; Calvi, Chiara; Conforti, Elena; Daffara, Rossella; Botto, Laura; Miserocchi, Giuseppe

2003-10-01

We evaluated in anesthetized rabbits the compositional changes of plasmalemmal lipid microdomains from lung tissue samples after inducing pulmonary interstitial edema (0.5 ml/kg for 3 h, leading to approximately 5% increase in extravascular water). Lipid microdomains (lipid rafts and caveolae) were present in the detergent-resistant fraction (DRF) obtained after discontinuous sucrose density gradient. DRF was enriched in caveolin-1, flotillin, aquaporin-1, GM1, cholesterol, sphingomyelin, and phosphatidylserine, and their contents significantly increased in interstitial edema. The higher DRF content in caveolin, flotillin, and aquaporin-1 and of the ganglioside GM1 suggests an increase both in caveolar domains and in lipid rafts, respectively. Compositional changes could be ascribed to endothelial and epithelial cells that provide most of plasma membrane surface area in the air-blood barrier. Alterations in lipid components in the plasma membrane may reflect rearrangement of floating lipid platforms within the membrane and/or lipid translocation from intracellular stores. Lipid traffic could be stimulated by the marked increase in hydraulic interstitial pressure after initial water accumulation, from approximately -10 to 5 cmH2O, due to the low compliance of the pulmonary tissue, in particular in the basement membranes and in the interfibrillar substance. Compositional changes in lipid microdomains represent a sign of cellular activation and suggest the potential role of mechanotransduction in response to developing interstitial edema.

Identification and characterization of VEGF and FGF from Hydra.

PubMed

Krishnapati, Lakshmi-Surekha; Ghaskadbi, Surendra

2013-01-01

Vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) play important roles in the formation of the blood vascular system and in axon guidance, nervous system development and function. Here, we report isolation and characterization of VEGF and FGF homologues from Hydra vulgaris Ind-Pune, a Cnidarian which exhibits an organized nervous system and primitive epithelio-muscular cells. VEGF expression was prominent in the endoderm of the peduncle region and tentacles, as evident from in situ hybridization of whole polyps and its transverse sections. High levels of FGF were detected in the ectoderm of the budding region. The expression of VEGF in endodermal and FGF in interstitial cells was confirmed using sf-1 hydra, a temperature-sensitive mutant strain of Hydra magnipapillata. Tissue-specific expression of VEGF and FGF was confirmed by semi quantitative RT-PCR for ectodermal and endodermal tissues in H. vulgaris Ind-Pune. Treatment with SU5416, a specific inhibitor of the VEGF receptor, did not affect the whole polyp, but did delay both budding and head regeneration, suggesting a possible role of VEGF in nerve cell development, tube formation and/or in branching. FGF expression in the ectoderm of budding region, where the majority of interstitial stem cells reside suggests its role in interstitial stem cell maintenance. Further, activation of canonical Wnt signalling with the glycogen synthase kinase-3β (GSK-3β) inhibitor alsterpaullone caused down-regulation of VEGF and FGF, suggesting an antagonistic relationship between the Wnt and VEGF/FGF pathways. Our results indicate that VEGF and FGF evolved early in evolution, before the development of the blood vascular system, and open up the possibility of elucidating the evolutionarily ancient functions of VEGF and FGF.

Urinary Peptides As a Novel Source of T Cell Allergen Epitopes

PubMed Central

da Silva Antunes, Ricardo; Pham, John; McMurtrey, Curtis; Hildebrand, William H.; Phillips, Elizabeth; Mallal, Simon; Sidney, John; Busse, Paula; Peters, Bjoern; Schulten, Véronique; Sette, Alessandro

2018-01-01

Mouse allergy in both laboratory workers and in inner-city children is associated with allergic rhinitis and asthma, posing a serious public health concern. Urine is a major source of mouse allergens, as mice spray urine onto their surroundings, where the proteins dry up and become airborne on dust particles. Here, we tested whether oligopeptides that are abundant in mouse urine may contribute to mouse allergic T cell response. Over 1,300 distinct oligopeptides were detected by mass spectrometry analysis of the low molecular weight filtrate fraction of mouse urine (LoMo). Posttranslationally modified peptides were common, accounting for almost half of total peptides. A pool consisting of 225 unique oligopeptides of 13 residues or more in size identified within was tested for its capacity to elicit T cell reactivity in mouse allergic donors. Following 14-day in vitro stimulation of PBMCs, we detected responses in about 95% of donors tested, directed against 116 distinct peptides, predominantly associated with Th2 cytokines (IL-5). Peptides from non-urine related proteins such as epidermal growth factor, collagen, and Beta-globin accounted for the highest response (15.9, 9.1, and 8.1% of the total response, respectively). Peptides derived from major urinary proteins (MUPs), kidney androgen-regulated protein (KAP), and uromodulin were the main T cell targets from kidney or urine related sources. Further ex vivo analysis of enrichment of 4-1BB expressing cells demonstrated that LoMo pool-specific T cell reactivity can be detected directly ex vivo in mouse allergic but not in non-allergic donors. Further cytometric analysis of responding cells revealed a bone fide memory T cell phenotype and confirmed their Th2 polarization. Overall, these data suggest that mouse urine-derived oligopeptides are a novel target for mouse allergy-associated T cell responses, which may contribute to immunopathological mechanisms in mouse allergy. PMID:29755469

A potential inhibitory function of draxin in regulating mouse trunk neural crest migration.

PubMed

Zhang, Sanbing; Su, Yuhong; Gao, Jinbao; Zhang, Chenbing; Tanaka, Hideaki

2017-01-01

Draxin is a repulsive axon guidance protein that plays important roles in the formation of three commissures in the central nervous system and dorsal interneuron 3 (dI3) in the chick spinal cord. In the present study, we report the expression pattern of mouse draxin in the embryonic mouse trunk spinal cord. In the presence of draxin, the longest net migration length of a migrating mouse trunk neural crest cell was significantly reduced. In addition, the relative number of apolar neural crest cells increased as the draxin treatment time increased. Draxin caused actin cytoskeleton rearrangement in the migrating trunk neural crest cells. Our data suggest that draxin may regulate mouse trunk neural crest cell migration by the rearrangement of cell actin cytoskeleton and by reducing the polarization activity of these cells subsequently.

Hyperbranched Polyglycerol as a Colloid in Cold Organ Preservation Solutions

PubMed Central

Gao, Sihai; Guan, Qiunong; Chafeeva, Irina; Brooks, Donald E.; Nguan, Christopher Y. C.; Kizhakkedathu, Jayachandran N.; Du, Caigan

2015-01-01

Hydroxyethyl starch (HES) is a common colloid in organ preservation solutions, such as in University of Wisconsin (UW) solution, for preventing graft interstitial edema and cell swelling during cold preservation of donor organs. However, HES has undesirable characteristics, such as high viscosity, causing kidney injury and aggregation of erythrocytes. Hyperbranched polyglycerol (HPG) is a branched compact polymer that has low intrinsic viscosity. This study investigated HPG (MW-0.5 to 119 kDa) as a potential alternative to HES for cold organ preservation. HPG was synthesized by ring-opening multibranching polymerization of glycidol. Both rat myocardiocytes and human endothelial cells were used as an in vitro model, and heart transplantation in mice as an in vivo model. Tissue damage or cell death was determined by both biochemical and histological analysis. HPG polymers were more compact with relatively low polydispersity index than HES in UW solution. Cold preservation of mouse hearts ex vivo in HPG solutions reduced organ damage in comparison to those in HES-based UW solution. Both size and concentration of HPGs contributed to the protection of the donor organs; 1 kDa HPG at 3 wt% solution was superior to HES-based UW solution and other HPGs. Heart transplants preserved with HPG solution (1 kDa, 3%) as compared with those with UW solution had a better functional recovery, less tissue injury and neutrophil infiltration in syngeneic recipients, and survived longer in allogeneic recipients. In cultured myocardiocytes or endothelial cells, significantly more cells survived after cold preservation with the HPG solution than those with the UW solution, which was positively correlated with the maintenance of intracellular adenosine triphosphate and cell membrane fluidity. In conclusion, HPG solution significantly enhanced the protection of hearts or cells during cold storage, suggesting that HPG is a promising colloid for the cold storage of donor organs and cells in transplantation. PMID:25706864

Hyperbranched polyglycerol as a colloid in cold organ preservation solutions.

PubMed

Gao, Sihai; Guan, Qiunong; Chafeeva, Irina; Brooks, Donald E; Nguan, Christopher Y C; Kizhakkedathu, Jayachandran N; Du, Caigan

2015-01-01

Hydroxyethyl starch (HES) is a common colloid in organ preservation solutions, such as in University of Wisconsin (UW) solution, for preventing graft interstitial edema and cell swelling during cold preservation of donor organs. However, HES has undesirable characteristics, such as high viscosity, causing kidney injury and aggregation of erythrocytes. Hyperbranched polyglycerol (HPG) is a branched compact polymer that has low intrinsic viscosity. This study investigated HPG (MW-0.5 to 119 kDa) as a potential alternative to HES for cold organ preservation. HPG was synthesized by ring-opening multibranching polymerization of glycidol. Both rat myocardiocytes and human endothelial cells were used as an in vitro model, and heart transplantation in mice as an in vivo model. Tissue damage or cell death was determined by both biochemical and histological analysis. HPG polymers were more compact with relatively low polydispersity index than HES in UW solution. Cold preservation of mouse hearts ex vivo in HPG solutions reduced organ damage in comparison to those in HES-based UW solution. Both size and concentration of HPGs contributed to the protection of the donor organs; 1 kDa HPG at 3 wt% solution was superior to HES-based UW solution and other HPGs. Heart transplants preserved with HPG solution (1 kDa, 3%) as compared with those with UW solution had a better functional recovery, less tissue injury and neutrophil infiltration in syngeneic recipients, and survived longer in allogeneic recipients. In cultured myocardiocytes or endothelial cells, significantly more cells survived after cold preservation with the HPG solution than those with the UW solution, which was positively correlated with the maintenance of intracellular adenosine triphosphate and cell membrane fluidity. In conclusion, HPG solution significantly enhanced the protection of hearts or cells during cold storage, suggesting that HPG is a promising colloid for the cold storage of donor organs and cells in transplantation.

Molecular Strategy to Reduce In Vivo Collagen Barrier Promotes Entry of NCX1 Positive Inducible Pluripotent Stem Cells (iPSCNCX1+) into Ischemic (or Injured) Myocardium

PubMed Central

Millard, Ronald W.; Yu, Xi-Yong; Luther, Kristin; Xu, Meifeng; Zhao, Ting C.; Yang, Huang-Tian; Qi, Zhihua; LaSance, Kathleen; Ashraf, Muhammad; Wang, Yigang

2013-01-01

Objective The purpose of this study was to assess the effect of collagen composition on engraftment of progenitor cells within infarcted myocardium. Background We previously reported that intramyocardial penetration of stem/progenitor cells in epicardial patches was enhanced when collagen was reduced in hearts overexpressing adenylyl cyclase-6 (AC6). In this study we hypothesized an alternative strategy wherein overexpression of microRNA-29b (miR-29b), inhibiting mRNAs that encode cardiac fibroblast proteins involved in fibrosis, would similarly facilitate progenitor cell migration into infarcted rat myocardium. Methods In vitro: A tri-cell patch (Tri-P) consisting of cardiac sodium-calcium exchanger-1 (NCX1) positive iPSC (iPSCNCX1+), endothelial cells (EC), and mouse embryonic fibroblasts (MEF) was created, co-cultured, and seeded on isolated peritoneum. The expression of fibrosis-related genes was analyzed in cardiac fibroblasts (CFb) by qPCR and Western blot. In vivo: Nude rat hearts were administered mimic miRNA-29b (miR-29b), miRNA-29b inhibitor (Anti-29b), or negative mimic (Ctrl) before creation of an ischemically induced regional myocardial infarction (MI). The Tri-P was placed over the infarcted region 7 days later. Angiomyogenesis was analyzed by micro-CT imaging and immunofluorescent staining. Echocardiography was performed weekly. Results The number of green fluorescent protein positive (GFP+) cells, capillary density, and heart function were significantly increased in hearts overexpressing miR-29b as compared with Ctrl and Anti-29b groups. Conversely, down-regulation of miR-29b with anti-29b in vitro and in vivo induced interstitial fibrosis and cardiac remodeling. Conclusion Overexpression of miR-29b significantly reduced scar formation after MI and facilitated iPSCNCX1+ penetration from the cell patch into the infarcted area, resulting in restoration of heart function after MI. PMID:23990893

Electron Microscopic Observations of the Carotid Body of the Cat

PubMed Central

Ross, Leonard L.

1959-01-01

Carotid bodies were removed from cats, fixed in buffered 1 per cent osmic acid, embedded in deaerated, nitrogenated methacrylate, and cut into thin sections for electron microscopic study. The carotid body is seen to be composed of islands of chemoreceptor and sustentacular cells surrounded by wide irregular sinusoids. These cells are separated from the sinusoids by relatively broad interstitial spaces which are filled with collagen, fibroblasts, and many unmyelinated nerve fibers with their Schwann cell sheaths. The chemoreceptor cells are surrounded by the flattened, multiprocessed sustentacular cells which serve to convey the axons from an interstitial to a pericellular location. These sustentacular cells are assumed to be lemmoblastic in origin. Relatively few axons are seen to abut on the chemoreceptor cells. The cytoplasm of the chemoreceptor cell is characterized by numerous small mitochondria, units of granular endoplasmic reticulum, a small Golgi complex, and a variety of vesicles. There are many small vesicles diffusely scattered throughout the cytoplasm. In addition, there is a small number of dark-cored vesicles of the type which has been previously described in the adrenal medulla. These are usually associated with the Golgi complex. These findings are discussed in relation to the concepts of the origin of the chemoreceptor cell and the nature of the synapse. PMID:14439171

Rheologic and hemodynamic characteristics of red cells of mouse, rat and human.

PubMed

Chen, D; Kaul, D K

1994-01-01

The present study compares hematologic, rheologic and hemodynamic characteristics of red cells from mouse, rat and human. Red cells in these species are biconcave discs that show significant differences in diameter and mean corpuscular volume (MCV). However, differences in mean corpuscular hemoglobin concentration (MCHC) are not significant. Viscosity measurement of washed red cell suspensions (in each case the medium osmolarity adjusted to match plasma osmolarity) showed significant interspecies differences at shear rates of 37.5 and 750 sec-1 as follows: Human > rat > mouse. Hemodynamic and microcirculatory behavior of these red cells was investigated in the artificially perfused ex vivo mesocecum vasculature of the rat. Hemodynamic measurements in the whole ex vivo mesocecum preparation revealed maximal increase in the peripheral resistance unit (PRU) for the human red cells followed by the rat and mouse red cells, respectively at a hematocrit (Hct) of 40%. Further, measurements of red cell velocities (Vrbc) in single arterioles of the mesocecum vasculature, during sustained perfusion with washed red cell suspensions, showed that at any given perfusion pressure (Pa), Vrbc for both mouse and rat red cells was higher than that for human red cells, while Vrbc for mouse red cells was higher than that for the rat. These results demonstrate that the microvascular flow behavior of these red cells is likely to be influenced by both physical and rheologic characteristics.

In vitro chronotropic effects of Erythrina senegalensis DC (Fabaceae) aqueous extract on mouse heart slice and pluripotent stem cell-derived cardiomyocytes.

PubMed

Nembo, Erastus Nembu; Atsamo, Albert Donatien; Nguelefack, Télesphore Benoît; Kamanyi, Albert; Hescheler, Jürgen; Nguemo, Filomain

2015-05-13

Erythrina senegalensis DC (Fabaceae) bark is commonly used in sub-Saharan traditional medicine for the treatment of many diseases including gastrointestinal disorders and cardiovascular diseases. In this study, we investigated the effect of the aqueous extract of the stem bark of Erythrina senegalensis on the contractile properties of mouse ventricular slices and human induced pluripotent stem (hiPS) cell-derived cardiomyocytes. We also investigated the cytotoxic effect of the extract on mouse embryonic stem (ES) cells differentiating into cardiomyocytes (CMs). We used well-established electrophysiological technologies to assess the effect of Erythrina senegalensis aqueous extract (ESAE) on the beating activity of mouse ventricular slices, mouse ES and hiPS cell-derived CMs. To study the cytotoxic effect of our extract, differentiating mouse ES cells were exposed to different concentrations of ESAE. EB morphology was assessed by microscopy at different stages of differentiation whereas cell viability was measured by flow cytometry, fluorometry and immunocytochemistry. The electrical activity of CMs and heart slices were respectively captured by the patch clamp technique and microelectrode array (MEA) method following ESAE acute exposure. Our findings revealed that ESAE exhibits a biphasic chronotropic activity on mouse ventricular slices with an initial low dose (0.001 and 0.01 µg/mL) decrease in beating activity followed by a corresponding significant increase in chronotropic activity at higher doses above 10 µg/mL. The muscarinic receptor blocker, atropine abolished the negative chronotropic activity of ESAE, while propranolol successfully blocked its positive chronotropic activity. ESAE showed a significant dose-dependent positive chronotropic activity on hiPS cell-derived CMs. Also, though not significantly, ESAE decreased cell viability and increased total caspase-3/7 activity of mouse ES cells in a concentration-dependent manner. Erythrina senegalensis aqueous extract exhibits a biphasic chronotropic effect on mouse heart and a positive chronotropic activity on hiPS cell-derived CMs, suggesting a possible mechanism through muscarinic and β-adrenergic receptor pathways. Also, ESAE is not cytotoxic on mouse ES cells at concentrations up to 100 µg/mL. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Feasibility of pig and human-derived aortic valve interstitial cells seeding on fixative-free decellularized animal pericardium.

PubMed

Santoro, Rosaria; Consolo, Filippo; Spiccia, Marco; Piola, Marco; Kassem, Samer; Prandi, Francesca; Vinci, Maria Cristina; Forti, Elisa; Polvani, Gianluca; Fiore, Gianfranco Beniamino; Soncini, Monica; Pesce, Maurizio

2016-02-01

Glutaraldehyde-fixed pericardium of animal origin is the elective material for the fabrication of bio-prosthetic valves for surgical replacement of insufficient/stenotic cardiac valves. However, the pericardial tissue employed to this aim undergoes severe calcification due to chronic inflammation resulting from a non-complete immunological compatibility of the animal-derived pericardial tissue resulting from failure to remove animal-derived xeno-antigens. In the mid/long-term, this leads to structural deterioration, mechanical failure, and prosthesis leaflets rupture, with consequent need for re-intervention. In the search for novel procedures to maximize biological compatibility of the pericardial tissue into immunocompetent background, we have recently devised a procedure to decellularize the human pericardium as an alternative to fixation with aldehydes. In the present contribution, we used this procedure to derive sheets of decellularized pig pericardium. The decellularized tissue was first tested for the presence of 1,3 α-galactose (αGal), one of the main xenoantigens involved in prosthetic valve rejection, as well as for mechanical tensile behavior and distensibility, and finally seeded with pig- and human-derived aortic valve interstitial cells. We demonstrate that the decellularization procedure removed the αGAL antigen, maintained the mechanical characteristics of the native pig pericardium, and ensured an efficient surface colonization of the tissue by animal- and human-derived aortic valve interstitial cells. This establishes, for the first time, the feasibility of fixative-free pericardial tissue seeding with valve competent cells for derivation of tissue engineered heart valve leaflets. © 2015 Wiley Periodicals, Inc.

Toll-Like Receptor Signaling and SIGIRR in Renal Fibrosis upon Unilateral Ureteral Obstruction

PubMed Central

Allam, Ramanjaneyulu; Ryu, Mi; Clauss, Sebastian; Susanti, Heni Eka; Römmele, Christoph; Garlanda, Cecilia; Mantovani, Alberto; Anders, Hans-Joachim

2011-01-01

Innate immune activation via IL-1R or Toll-like receptors (TLR) contibutes to acute kidney injury but its role in tissue remodeling during chronic kidney disease is unclear. SIGIRR is an inhibitor of TLR-induced cytokine and chemokine expression in intrarenal immune cells, therefore, we hypothesized that Sigirr-deficiency would aggravate postobstructive renal fibrosis. The expression of TLRs as well as endogenous TLR agonists increased within six days after UUO in obstructed compared to unobstructed kidneys while SIGIRR itself was downregulated by day 10. However, lack of SIGIRR did not affect the intrarenal mRNA expression of proinflammatory and profibrotic mediators as well as the numbers of intrarenal macrophages and T cells or morphometric markers of tubular atrophy and interstitial fibrosis. Because SIGIRR is known to block TLR/IL-1R signaling at the level of the intracellular adaptor molecule MyD88 UUO experiments were also performed in mice deficient for either MyD88, TLR2 or TLR9. After UUO there was no significant change of tubular interstitial damage and interstitial fibrosis in neither of these mice compared to wildtype counterparts. Additional in-vitro studies with CD90+ renal fibroblasts revealed that TLR agonists induce the expression of IL-6 and MCP-1/CCL2 but not of TGF-β, collagen-1α or smooth muscle actin. Together, postobstructive renal interstitial fibrosis and tubular atrophy develop independent of SIGIRR, TLR2, TLR9, and MyD88. These data argue against a significant role of these molecules in renal fibrosis. PMID:21544241

Two Pore Channel 2 Differentially Modulates Neural Differentiation of Mouse Embryonic Stem Cells

PubMed Central

Zhang, Zhe-Hao; Lu, Ying-Ying; Yue, Jianbo

2013-01-01

Nicotinic acid adenine dinucleotide phosphate (NAADP) is an endogenous Ca2+ mobilizing nucleotide presented in various species. NAADP mobilizes Ca2+ from acidic organelles through two pore channel 2 (TPC2) in many cell types and it has been previously shown that NAADP can potently induce neuronal differentiation in PC12 cells. Here we examined the role of TPC2 signaling in the neural differentiation of mouse embryonic stem (ES) cells. We found that the expression of TPC2 was markedly decreased during the initial ES cell entry into neural progenitors, and the levels of TPC2 gradually rebounded during the late stages of neurogenesis. Correspondingly, TPC2 knockdown accelerated mouse ES cell differentiation into neural progenitors but inhibited these neural progenitors from committing to neurons. Overexpression of TPC2, on the other hand, inhibited mouse ES cell from entering the early neural lineage. Interestingly, TPC2 knockdown had no effect on the differentiation of astrocytes and oligodendrocytes of mouse ES cells. Taken together, our data indicate that TPC2 signaling plays a temporal and differential role in modulating the neural lineage entry of mouse ES cells, in that TPC2 signaling inhibits ES cell entry to early neural progenitors, but is required for late neuronal differentiation. PMID:23776607

Identification of transcriptional regulators in the mouse immune system

PubMed Central

Jojic, Vladimir; Shay, Tal; Sylvia, Katelyn; Zuk, Or; Sun, Xin; Kang, Joonsoo; Regev, Aviv; Koller, Daphne

2013-01-01

The differentiation of hematopoietic stem cells into immune cells has been extensively studied in mammals, but the transcriptional circuitry controlling it is still only partially understood. Here, the Immunological Genome Project gene expression profiles across mouse immune lineages allowed us to systematically analyze these circuits. Using a computational algorithm called Ontogenet, we uncovered differentiation-stage specific regulators of mouse hematopoiesis, identifying many known hematopoietic regulators, and 175 new candidate regulators, their target genes, and the cell types in which they act. Among the novel regulators, we highlight the role of ETV5 in γδT cells differntiation. Since the transcriptional program of human and mouse cells is highly conserved1, it is likely that many lessons learned from the mouse model apply to humans. PMID:23624555

Development and function of human innate immune cells in a humanized mouse model.

PubMed

Rongvaux, Anthony; Willinger, Tim; Martinek, Jan; Strowig, Till; Gearty, Sofia V; Teichmann, Lino L; Saito, Yasuyuki; Marches, Florentina; Halene, Stephanie; Palucka, A Karolina; Manz, Markus G; Flavell, Richard A

2014-04-01

Mice repopulated with human hematopoietic cells are a powerful tool for the study of human hematopoiesis and immune function in vivo. However, existing humanized mouse models cannot support development of human innate immune cells, including myeloid cells and natural killer (NK) cells. Here we describe two mouse strains called MITRG and MISTRG, in which human versions of four genes encoding cytokines important for innate immune cell development are knocked into their respective mouse loci. The human cytokines support the development and function of monocytes, macrophages and NK cells derived from human fetal liver or adult CD34(+) progenitor cells injected into the mice. Human macrophages infiltrated a human tumor xenograft in MITRG and MISTRG mice in a manner resembling that observed in tumors obtained from human patients. This humanized mouse model may be used to model the human immune system in scenarios of health and pathology, and may enable evaluation of therapeutic candidates in an in vivo setting relevant to human physiology.

Development and function of human innate immune cells in a humanized mouse model

PubMed Central

Rongvaux, Anthony; Willinger, Tim; Martinek, Jan; Strowig, Till; Gearty, Sofia V.; Teichmann, Lino L.; Saito, Yasuyuki; Marches, Florentina; Halene, Stephanie; Palucka, A. Karolina; Manz, Markus G.; Flavell, Richard A.

2014-01-01

Mice repopulated with human hematopoietic cells are a powerful tool for the study of human hematopoiesis and immune function in vivo. However, existing humanized mouse models are unable to support development of human innate immune cells, including myeloid cells and NK cells. Here we describe a mouse strain, called MI(S)TRG, in which human versions of four genes encoding cytokines important for innate immune cell development are knocked in to their respective mouse loci. The human cytokines support the development and function of monocytes/macrophages and natural killer cells derived from human fetal liver or adult CD34+ progenitor cells injected into the mice. Human macrophages infiltrated a human tumor xenograft in MI(S)TRG mice in a manner resembling that observed in tumors obtained from human patients. This humanized mouse model may be used to model the human immune system in scenarios of health and pathology, and may enable evaluation of therapeutic candidates in an in vivo setting relevant to human physiology. PMID:24633240

Deletions of the long arm of chromosome 5 define subgroups of T-cell acute lymphoblastic leukemia

PubMed Central

La Starza, Roberta; Barba, Gianluca; Demeyer, Sofie; Pierini, Valentina; Di Giacomo, Danika; Gianfelici, Valentina; Schwab, Claire; Matteucci, Caterina; Vicente, Carmen; Cools, Jan; Messina, Monica; Crescenzi, Barbara; Chiaretti, Sabina; Foà, Robin; Basso, Giuseppe; Harrison, Christine J.; Mecucci, Cristina

2016-01-01

Recurrent deletions of the long arm of chromosome 5 were detected in 23/200 cases of T-cell acute lymphoblastic leukemia. Genomic studies identified two types of deletions: interstitial and terminal. Interstitial 5q deletions, found in five cases, were present in both adults and children with a female predominance (chi-square, P=0.012). Interestingly, these cases resembled immature/early T-cell precursor acute lymphoblastic leukemia showing significant down-regulation of five out of the ten top differentially expressed genes in this leukemia group, including TCF7 which maps within the 5q31 common deleted region. Mutations of genes known to be associated with immature/early T-cell precursor acute lymphoblastic leukemia, i.e. WT1, ETV6, JAK1, JAK3, and RUNX1, were present, while CDKN2A/B deletions/mutations were never detected. All patients had relapsed/resistant disease and blasts showed an early differentiation arrest with expression of myeloid markers. Terminal 5q deletions, found in 18 of patients, were more prevalent in adults (chi-square, P=0.010) and defined a subgroup of HOXA-positive T-cell acute lymphoblastic leukemia characterized by 130 up- and 197 down-regulated genes. Down-regulated genes included TRIM41, ZFP62, MAPK9, MGAT1, and CNOT6, all mapping within the 1.4 Mb common deleted region at 5q35.3. Of interest, besides CNOT6 down-regulation, these cases also showed low BTG1 expression and a high incidence of CNOT3 mutations, suggesting that the CCR4-NOT complex plays a crucial role in the pathogenesis of HOXA-positive T-cell acute lymphoblastic leukemia with terminal 5q deletions. In conclusion, interstitial and terminal 5q deletions are recurrent genomic losses identifying distinct subtypes of T-cell acute lymphoblastic leukemia. PMID:27151989

Deletions of the long arm of chromosome 5 define subgroups of T-cell acute lymphoblastic leukemia.

PubMed

La Starza, Roberta; Barba, Gianluca; Demeyer, Sofie; Pierini, Valentina; Di Giacomo, Danika; Gianfelici, Valentina; Schwab, Claire; Matteucci, Caterina; Vicente, Carmen; Cools, Jan; Messina, Monica; Crescenzi, Barbara; Chiaretti, Sabina; Foà, Robin; Basso, Giuseppe; Harrison, Christine J; Mecucci, Cristina

2016-08-01

Recurrent deletions of the long arm of chromosome 5 were detected in 23/200 cases of T-cell acute lymphoblastic leukemia. Genomic studies identified two types of deletions: interstitial and terminal. Interstitial 5q deletions, found in five cases, were present in both adults and children with a female predominance (chi-square, P=0.012). Interestingly, these cases resembled immature/early T-cell precursor acute lymphoblastic leukemia showing significant down-regulation of five out of the ten top differentially expressed genes in this leukemia group, including TCF7 which maps within the 5q31 common deleted region. Mutations of genes known to be associated with immature/early T-cell precursor acute lymphoblastic leukemia, i.e. WT1, ETV6, JAK1, JAK3, and RUNX1, were present, while CDKN2A/B deletions/mutations were never detected. All patients had relapsed/resistant disease and blasts showed an early differentiation arrest with expression of myeloid markers. Terminal 5q deletions, found in 18 of patients, were more prevalent in adults (chi-square, P=0.010) and defined a subgroup of HOXA-positive T-cell acute lymphoblastic leukemia characterized by 130 up- and 197 down-regulated genes. Down-regulated genes included TRIM41, ZFP62, MAPK9, MGAT1, and CNOT6, all mapping within the 1.4 Mb common deleted region at 5q35.3. Of interest, besides CNOT6 down-regulation, these cases also showed low BTG1 expression and a high incidence of CNOT3 mutations, suggesting that the CCR4-NOT complex plays a crucial role in the pathogenesis of HOXA-positive T-cell acute lymphoblastic leukemia with terminal 5q deletions. In conclusion, interstitial and terminal 5q deletions are recurrent genomic losses identifying distinct subtypes of T-cell acute lymphoblastic leukemia. Copyright© Ferrata Storti Foundation.

Assessment of Parylene C Thin Films for Heart Valve Tissue Engineering

PubMed Central

Marei, Isra; Chester, Adrian; Carubelli, Ivan; Prodromakis, Themistoklis; Trantidou, Tatiana

2015-01-01

Background: Scaffolds are a key component of tissue-engineered heart valves (TEHVs). Several approaches had been adopted in the design of scaffolds using both natural and synthetic resources. We have investigated the suitability of parylene C (PC), a vapor deposited polymeric material, for the use as a scaffold in TEHV. Aims: To evaluate the adsorption of extracellular matrix components onto plasma-activated PC and study the biocompatibility of PC by measuring cellular adhesion, viability, apoptosis, and phenotypic expression of valve endothelial and interstitial cells. Finally, the mechanical properties of PC were compared with those of native aortic valve cusp tissue. Methods: PC slides were plasma activated and then coated with gelatin, type I collagen, or fibronectin. Porcine pulmonary valve endothelial and interstitial cells were then grown on plasma oxidized PC with different types of coatings and their adhesion was observed after 20 h of incubation. Cell viability was tested using the MTS assay, and apoptosis was estimated using TUNEL staining. The mechanical properties of PC and valve tissue were measured using a Bose Mechanical Tester. Finally, cell-seeded PC films were exposed to pulsatile pressure and aortic shear stress, respectively, to test their durability in a dynamic environment. Results: Our findings show that collagen and fibronectin could bind to plasma oxidized PC. Both valve endothelial and interstitial cells adhered to protein-coated ECM. PC had a profile of mechanical stiffness and ultimate tensile strength that were comparable with or in excess of those seen in porcine aortic valve cusps. Cells were still attached to PC films after 3 days of exposure to up to 50 mmHg pulsatile pressure or aortic levels of shear stress. Conclusion: PC is a promising candidate for use as a scaffold in tissue engineering heart valves. Additional studies are required to determine both the durability and long-term performance of cell-seeded PC when in a similar hemodynamic environment to that of the aortic valve. PMID:26101808

Radiation-Induce Immune Modulation in Prostate Cancer

DTIC Science & Technology

2005-01-01

Prostate-specific antigen Prostate carcinoma Mammoglobin-A Breast carcinoma Overexpressed Alpha - fetoprotein Hepatocellular carcinoma and yolk-sac tumors...Interleukin-3 cooperates with tumor necrosis factor alpha for the development of human dendritic/Langerhans cells from cord blood CD34+ hematopoietic progenitor...additional subsets, e.g. Langerhans cells of the epidermis, and dermal or interstitial DC. PDC are the major interferon- alpha (IFNca) producing cells

Mouse embryo attachment to substratum and interaction of trophoblast with cultured cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Glass, R.H.; Spindle, A.I.; Pedersen, R.A.

1979-06-01

Hatching, attachment, and trophoblast outgrowth of mouse embryos in vitro were examined as a model for implantation. Mouse embryos attached and grew out on glass cover slips that were partially covered with cultured mouse cells (L cells, liver cells, transformed JLS-V11 cells, and teratocarcinoma cells). Scanning electron microscopy showed that processes of these cells made contact with trophoblast, but there was no evidence of cell lysis or of phagocytosis of the cells by trophoblast. Time-lapse cinematography showed that after contact the cultured mouse cells retracted from the trophoblast, which then spread into the areas vacated by those cells. This suggestsmore » a means by which the trophoblast gains entry into the endometrium without destruction of maternal cells. Neuraminidase (100 or 250 units/ml) had no effect on attachment of mouse embryos to glass. However, attachment was inhibited by trypsin at concentrations of 0.25%, 0.025%, and 0.0025%. Treatment of early blastocysts with diazooxo-norleucine, an inhibitor of glycoprotein synthesis, decreased the number of embryos hatching from the zona pellucida; treatment at the late blastocyst stage decreased hatching to a lesser extent. Among the late blastocysts that did hatch, the number forming trophoblast outgrowths was lower than in controls. These results suggest that glycoproteins may be of importance for embryo hatching, attachment, and outgrowth.« less

Rho kinase inhibitor Y-27632 and Accutase dramatically increase mouse embryonic stem cell derivation.

PubMed

Zhang, Peng; Wu, Xinglong; Hu, Chunchao; Wang, Pengbo; Li, Xiangyun

2012-01-01

Although it has been 30 yr since the development of derivation methods for mouse embryonic stem (ES) cells, the biology of derivation of ES cells is poorly understood and the efficiency varies dramatically between cell lines. Recently, the Rho kinase inhibitor Y-27632 and the cell dissociation reagent Accutase were reported to significantly inhibit apoptosis of human ES cells during passaging. Therefore, in the current study, C57BL/6×129/Sv mouse blastocysts were used to evaluate the effect of the combination of the two reagents instead of using the conventional 129 line in mouse ES cell derivation. The data presented in this study suggests that the combination of Y-27632 and Accutase significantly increases the efficiency of mouse ES cell derivation; furthermore, no negative side effects were observed with Y-27632 and Accutase treatment. The newly established ES cell lines retain stable karyotype, surface markers expression, formed teratomas, and contributed to viable chimeras and germline transmission by tetraploid complementation assay. In addition, Y-27632 improved embryoid body formation of ES cells. During ES cell microinjection, Y-27632 prevented the formation of dissociation-induced cell blebs and facilitates the selection and the capture of intact cells. The methods presented in this study clearly demonstrate that inhibition of Rho kinase with Y-27632 and Accutase dissociation improve the derivation efficiently and reproducibility of mouse ES cell generation which is essential for reducing variability in the results obtained from different cell lines.

Inhibition of WISE preserves renal allograft function.

PubMed

Qian, Xueming; Yuan, Xiaodong; Vonderfecht, Steven; Ge, Xupeng; Lee, Jae; Jurisch, Anke; Zhang, Li; You, Andrew; Fitzpatrick, Vincent D; Williams, Alexia; Valente, Eliane G; Pretorius, Jim; Stevens, Jennitte L; Tipton, Barbara; Winters, Aaron G; Graham, Kevin; Harriss, Lindsey; Baker, Daniel M; Damore, Michael; Salimi-Moosavi, Hossein; Gao, Yongming; Elkhal, Abdallah; Paszty, Chris; Simonet, W Scott; Richards, William G; Tullius, Stefan G

2013-01-01

Wnt-modulator in surface ectoderm (WISE) is a secreted modulator of Wnt signaling expressed in the adult kidney. Activation of Wnt signaling has been observed in renal transplants developing interstitial fibrosis and tubular atrophy; however, whether WISE contributes to chronic changes is not well understood. Here, we found moderate to high expression of WISE mRNA in a rat model of renal transplantation and in kidneys from normal rats. Treatment with a neutralizing antibody against WISE improved proteinuria and graft function, which correlated with higher levels of β-catenin protein in kidney allografts. In addition, treatment with the anti-WISE antibody reduced infiltration of CD68(+) macrophages and CD8(+) T cells, attenuated glomerular and interstitial injury, and decreased biomarkers of renal injury. This treatment reduced expression of genes involved in immune responses and in fibrogenic pathways. In summary, WISE contributes to renal dysfunction by promoting tubular atrophy and interstitial fibrosis.

Role of lipids in the progression of renal disease in systemic lupus erythematosus patients.

PubMed

Luzar, B; Ferluga, D

2000-08-25

Systemic lupus erythematosus (SLE) is an autoimmune connective tissue disease marked by immune-complex mediated lesions in small blood vessels of various organs, especially the kidneys, although other factors may also be implicated in the pathogenesis of the disease. This article focuses on the role of lipids in the progression of glomerular, vascular and tubulo-interstitial lesions in two patients with lupus nephritis associated with pronounced hyper- and dyslipidemia. The pathogenesis of progressive glomerulosclerosis in both patients appears to be multifactorial. In addition to immune complex mediated lupus glomerulonephritis, progressively active in the first patient, severe nephrotic-range persistent proteinuria, arterial hypertension associated with hyperfiltration and hyperperfusion injuries and, to a minor extent, hyper- and dyslipidemia were observed. Immunological and non-immunological factors were shown to contribute to the development of tubulo-interstitial lesions. In both patients, in addition to local immune deposits, prominent tubulo-interstitial lipid deposits were probably causally related to both hyperlipidemia and the increased permeability of the glomerular filtration barrier. Tubular lesions were highlighted by intracytoplasmic lipid droplets as well as small cleft-like spaces found to be impacted in the tubular lumina. They were seen to penetrate tubular epithelial cells and eventually lodge in the interstitium, surrounded by mononuclear cell infiltrates and foam cells. In both patients, hypertensive angiopathy and extraglomerular vascular immune deposits were demonstrated. In addition, in the second patient, arteriolar and small arterial hyaline was found at the age of 28 years to be full of lipids and calcium precipitates, suggesting a peripheral atherosclerosis-like process which never occurs as a natural age-related condition. In conclusion, all parts of the nephron may be involved in the pathogenetic process causally related or influenced by hyper- or dyslipidemia. Associated either with endothelial cell injury and consequent insudation of lipids in the vascular walls, glomerular filtration barrier injury with hyperfiltration, or tubulo-interstitial lipid deposition, the mechanism of tissue damage by lipids in all parts of the nephron shares similarities with the pathogenesis of systemic atherosclerosis.

[Effect of topical application of a recombinant adenovirus carrying promyelocytic leukemia gene in a psoriasis-like mouse model].

PubMed

Wang, Qiongyu; Zhang, Aijun; Ma, Huiqun; Wang, Shijie; Ma, Yunyun; Zou, Xingwei; Li, Ruilian

2013-03-01

To investigate the effects of topical treatment with adenovirus-mediated promyelocytic leukemia gene (PML) gene in a psoriasis-like mouse model. The effect of adenovirus-mediated PML gene on the granular layer of mouse tail scale epidermis and epithelial mitosis were observed on longitudinal histological sections prepared from the tail skin and vaginal epithelium of the mice. Adenovirus-mediated PML gene significantly inhibited mitosis of mouse vaginal epithelial cells and promoted the formation of granular layer in mouse tail scale epidermis. The therapeutic effect of PML gene in the psoriasis-like mouse model may be associated with increased granular cells and suppressed epidemic cell proliferation.

Expression of the Intermediate Filament Nestin in Gastrointestinal Stromal Tumors and Interstitial Cells of Cajal

PubMed Central

Tsujimura, Tohru; Makiishi-Shimobayashi, Chiaki; Lundkvist, Johan; Lendahl, Urban; Nakasho, Keiji; Sugihara, Ayako; Iwasaki, Teruo; Mano, Masayuki; Yamada, Naoko; Yamashita, Kunihiro; Toyosaka, Akihiro; Terada, Nobuyuki

2001-01-01

It has recently been proposed that gastrointestinal stromal tumors (GISTs) originate from stem cells that differentiate toward a phenotype of interstitial cells of Cajal (ICCs). Nestin is a newly identified intermediate filament protein, and is predominantly expressed in immature cells, such as neuroectodermal stem cells and skeletal muscle progenitor cells, and tumors originating from these cells. In this study, we examined, using immunohistochemistry, the nestin expression in GISTs and ICCs to clarify the origin of GISTs. Strong immunoreactivity for nestin was observed in all 18 GISTs, and its expression was confirmed by Western blot and Northern blot analyses. In contrast, three leiomyomas and a schwannoma that developed in the gastrointestinal tract showed no apparent immunoreactivity for nestin. Among 17 mesenchymal tumors (seven leiomyosarcomas, five malignant peripheral nerve sheath tumors, and five fibrosarcomas) that occurred in sites other than the gastrointestinal tract, only two malignant peripheral nerve sheath tumors were moderately immunoreactive for nestin. Furthermore, with fluorescence double immunostaining of the normal small intestine, nestin expression was demonstrated in ICCs. These results show that nestin may be a useful marker for diagnosis of GISTs, and support the current hypothesis that GISTs are tumors of stem cells that differentiate toward an ICC phenotype. PMID:11238030

Molecular cloning of a human Ca2+-dependent cell-cell adhesion molecule homologous to mouse placental cadherin: its low expression in human placental tissues

PubMed Central

1989-01-01

P-cadherin is a subclass of Ca2+-dependent cell-cell adhesion molecules present in mouse placenta, where its localization suggests a function of connecting the embryo to the uterus (Nose, A., and M. Takeichi. 1986. J. Cell Biol. 103:2649-2658). We recently identified a human cadherin detected by an mAb capable of disrupting cell-cell adhesion of A-431 cells, and found that it was closely related immunochemically to mouse P-cadherin. Curiously, this cadherin was undetectable in human placenta by immunohistochemical examination (Shimoyama, Y., S. Hirohashi, S. Hirano, M. Noguchi, Y. Shimosato, M. Takeichi, and O. Abe. 1989. Cancer Res. 49:2128-2133). We here report the cloning and sequencing of cDNA clone encoding the human homologue of mouse P- cadherin. The deduced amino acid sequence of the human P-cadherin consists of 829 amino acid and shows striking homology with mouse P- cadherin. On Northern blot analysis, human P-cadherin was scarcely expressed in human placenta in contrast to mouse P-cadherin, which was abundantly expressed in mouse placenta throughout pregnancy, and it was shown that E-cadherin, but not P-cadherin, was the major cadherin molecule in human placenta. Moreover, NIH3T3 cells transfected with human P-cadherin cDNA expressed the functional cadherin molecule, which was identical to the cadherin we had previously identified using the mAb, showing that this molecule really does mediate cell-cell adhesion and that the cadherin we detected immunochemically is undoubtedly human P-cadherin. The results obtained in this study support the idea that P- cadherin plays little role, if any, in Ca2+-dependent cell-cell binding in human placental tissue at least after several weeks of pregnancy. PMID:2793940

Primary Xenografts of Human Prostate Tissue as a Model to Study Angiogenesis Induced by Reactive Stroma

PubMed Central

Montecinos, Viviana P.; Godoy, Alejandro; Hinklin, Jennifer; Vethanayagam, R. Robert; Smith, Gary J.

2012-01-01

Characterization of the mechanism(s) of androgen-driven human angiogenesis could have significant implications for modeling new forms of anti-angiogenic therapies for CaP and for developing targeted adjuvant therapies to improve efficacy of androgen-deprivation therapy. However, models of angiogenesis by human endothelial cells localized within an intact human prostate tissue architecture are until now extremely limited. This report characterizes the burst of angiogenesis by endogenous human blood vessels in primary xenografts of fresh surgical specimens of benign prostate or prostate cancer (CaP) tissue that occurs between Days 6–14 after transplantation into SCID mice pre-implanted with testosterone pellets. The wave of human angiogenesis was preceded by androgen-mediated up-regulation of VEGF-A expression in the stromal compartment. The neo-vessel network anastomosed to the host mouse vascular system between Days 6–10 post-transplantation, the angiogenic response ceased by Day 15, and by Day 30 the vasculature had matured and stabilized, as indicated by a lack of leakage of serum components into the interstitial tissue space and by association of nascent endothelial cells with mural cells/pericytes. The angiogenic wave was concurrent with the appearance of a reactive stroma phenotype, as determined by staining for α-SMA, Vimentin, Tenascin, Calponin, Desmin and Masson's trichrome, but the reactive stroma phenotype appeared to be largely independent of androgen availability. Transplantation-induced angiogenesis by endogenous human endothelial cells present in primary xenografts of benign and malignant human prostate tissue was preceded by induction of androgen-driven expression of VEGF by the prostate stroma, and was concurrent with and the appearance of a reactive stroma phenotype. Androgen-modulated expression of VEGF-A appeared to be a causal regulator of angiogenesis, and possibly of stromal activation, in human prostate xenografts. PMID:22303438

Intratumoral Delivery of Interferonγ-Secreting Mesenchymal Stromal Cells Repolarizes Tumor-Associated Macrophages and Suppresses Neuroblastoma Proliferation In Vivo.

PubMed

Relation, Theresa; Yi, Tai; Guess, Adam J; La Perle, Krista; Otsuru, Satoru; Hasgur, Suheyla; Dominici, Massimo; Breuer, Christopher; Horwitz, Edwin M

2018-06-01

Neuroblastoma, the most common extracranial solid tumor in childhood, remains a therapeutic challenge. However, one promising patient treatment strategy is the delivery of anti-tumor therapeutic agents via mesenchymal stromal cell (MSC) therapy. MSCs have been safely used to treat genetic bone diseases such as osteogenesis imperfecta, cardiovascular diseases, autoimmune diseases, and cancer. The pro-inflammatory cytokine interferon-gamma (IFNγ) has been shown to decrease tumor proliferation by altering the tumor microenvironment (TME). Despite this, clinical trials of systemic IFNγ therapy have failed due to the high blood concentration required and associated systemic toxicities. Here, we developed an intra-adrenal model of neuroblastoma, characterized by liver and lung metastases. We then engineered MSCs to deliver IFNγ directly to the TME. In vitro, these MSCs polarized murine macrophages to the M1 phenotype. In vivo, we attained a therapeutically active TME concentration of IFNγ without increased systemic concentration or toxicity. The TME-specific IFNγ reduced tumor growth rate and increased survival in two models of T cell deficient athymic nude mice. Absence of this benefit in NOD SCID gamma (NSG) immunodeficient mouse model indicates a mechanism dependent on the innate immune system. IL-17 and IL-23p19, both uniquely M1 polarization markers, transiently increased in the tumor interstitial fluid. Finally, the MSC vehicle did not promote tumor growth. These findings reveal that MSCs can deliver effective cytokine therapy directly to the tumor while avoiding systemic toxicity. This method transiently induces inflammatory M1 macrophage polarization, which reduces tumor burden in our novel neuroblastoma murine model. Stem Cells 2018;36:915-924. © AlphaMed Press 2018.

Absence of Activation-induced Cytidine Deaminase, a Regulator of Class Switch Recombination and Hypermutation in B Cells, Suppresses Aorta Allograft Vasculopathy in Mice.

PubMed

Nakanishi, Tomonori; Xu, Xiaoyan; Wynn, Carmen; Yamada, Toshiko; Pan, Fan; Erickson, Laurie; Teo, Haeman; Nakagawa, Terry; Masunaga, Taro; Abe, Jumpei; Akamatsu, Masahiko; Tamura, Kouichi; Jiang, Hongsi

2015-08-01

Antibody-mediated rejection is caused in part by increasing circulation/production of donor-specific antibody (DSA). Activation-induced cytidine deaminase (AID) is a key regulator of class switch recombination and somatic hypermutation of immunoglobulin in B cells, yet its role in antibody-mediated transplant rejection remains unclear. We show here that AID deficiency in mice enables suppression of allograft vasculopathy (AV) after aorta transplantation, a DSA-mediated process. Splenocytes from C57BL/6 J (B6) AID(−/−) mice were used for determining in vitro proliferation responses, alloreactivity, cell surface marker expression, and antibody production. BALB/c mouse aortas were transplanted into B6 AID(−/−) mice with or without FK506 treatment. Blood and aorta grafts were harvested on day 30 after transplantation and were subjected to DSA, histological, and immunohistological analyses. The AID(−/−) splenocytes were comparable to wild type splenocytes in proliferation responses, alloreactivity, and expression of cell surface markers in vitro. However, they completely failed to produce immunoglobulin G, although they were not impaired in immunoglobulin M production relative to controls. Furthermore, BALB/c aorta grafts from B6 AID(−/−) recipient mice on day 30 after transplantation showed reduced signs of AV compared to the grafts from B6 wild type recipient mice which had severe vascular intimal hyperplasia, interstitial fibrosis, and inflammation. Treatment with FK506 produced a synergistic effect in the grafts from AID(−/−) recipients with further reduction of intimal hyperplasia and fibrosis scores. The AID deficiency inhibits DSA-mediated AV after aorta transplantation in mice. We propose that AID could be a novel molecular target for controlling antibody-mediated rejection in organ transplantation.

The role of lead and excess oxygen in uranite

USGS Publications Warehouse

Berman, Robert Morris

1957-01-01

Analysed samples of uraninite were x-rayed, annealed by heating to 550° and 900° for various times in a nitrogen atmosphere, and x-rayed again. A decrease in unit cell size was generally observed. Calculations on the basis of Vegard's Law showed that the ordering of the interstitial oxygen ions could account for the decrease in cell size on annealing. The interstitial oxygens are not necessarily completely disordered before annealing. The degree of original disorder is dependent on the Rare Earth/ThO2 ratio; for high ThO2 and low rare earths, the interstitial oxygens are completely random. The degree of disorder apparently depends solely on the composition, and not on the past history of the sample; this implies that the oxygens are being continuously disordered, perhaps by alpha particles, to the equilibrium point determined by the R.E./ThO2 ratio. The degree of ordering of the interstitial oxygens also accounts for the difference in cell size between vein pitchblendes and those from the sediments of the Colorado Plateau. A study was also made of the degree of oxidation of uraninites. Although the uranium in many pegmatitic uraninites is more oxidized than can be obtained with the cubic UO2 phase in the laboratory, if the atoms proxying for uranium are calculated into the structural formula, and the lead is assumed to be radiogenic and calculated as original uranium, almost all pegmatitic uraninites fall into the range of interstitial oxygen content obtainable in the laboratory. This fact supports the auto-oxidation hypothesis. Many of the vein and sedimentary pitchblendes have compositions close to U3O8, although they are cubic. They may gave crystallized as U3O8, the decomposed to the cubic phase and a amorphous phase. This suggests that the stability range of U3O8 includes only very exceptional natural conditions. Vegard's Law calculations, studies of zoning in crystals, differential leaching, polished section textures, and other lines of evidence indicate that lead, including radiogenic lead, is exsolved from uraninite. A study of x-ray line intensities indicates that it exsolves as oriented monomolecular layers of orthohombic PbO (massicot) along cube planes in the uraninite, separating the uraninite crystallites so that the x-ray reflections interfere destructively to different degrees for different reflections.

Production of an aminoterminally truncated, stable type of bioactive mouse fibroblast growth factor 4 in Escherichia coli.

PubMed

Sugawara, Saiko; Ito, Toshihiko; Sato, Shiori; Sato, Yuki; Kasuga, Kano; Kojima, Ikuo; Kobayashi, Masayuki

2014-05-01

In mice, fibroblast growth factor 4 (Fgf4) is a crucial gene for the generation of trophectoderm, progenitor cells of the placenta. Therefore, exogenous FGF4 promotes the isolation and maintenance of trophoblast stem cells from preimplantation embryos. We previously produced a 6× histidine (His)-tagged, mouse FGF4 (Pro(31)-Leu(202)) without a secretory signal peptide at the amino-terminus, referred to as HismFGF4, in Escherichia coli. Here, we found that HismFGF4 was unstable, such as in phosphate-buffered saline. In these conditions, site-specific cleavage between Ser(50) and Leu(51) was identified. In order to generate stable mouse FGF4 derivatives, a 6× His-tagged mouse FGF4 (Leu(51)-Leu(202)), termed HismFGF4L, was expressed in E. coli. HismFGF4L could be purified from the supernatant of cell lysates by heparin column chromatography. In phosphate-buffered saline, HismFGF4L was relatively stable. HismFGF4L exerted significant mitogenic activities at concentrations as low as 0.01 nM (P < 0.01) in mouse embryonic fibroblast Balb/c 3T3 cells expressing FGF receptor 2. In the presence of PD173074, an FGF receptor inhibitor, the growth-promoting activity of HismFGF4L was abolished. Taken together, we suggest that aminoterminally truncated HismFGF4L is capable of promoting the proliferation of mouse-derived cells via an authentic FGF signaling pathway. We consider that HismFGF4L is useful as a derivative of mouse FGF4 protein for analyzing the effects of mouse FGF4 and for stimulating cell growth of mouse-derived cells, such as trophoblast stem cells. Our study provides a simple method for the production of a bioactive, stable mouse FGF4 derivative in E. coli. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Developing Novel Therapeutic Approaches in Small Cell Lung Carcinoma Using Genetically Engineered Mouse Models and Human Circulating Tumor Cells

DTIC Science & Technology

2014-10-01

AD_________________ Award Number: W81XWH-13-1-0325 TITLE: Developing Novel Therapeutic Approaches in Small Cell Lung Carcinoma Using ...Genetically Engineered Mouse Models and Human Circulating Tumor Cells PRINCIPAL INVESTIGATOR: Jeffrey Engelman MD PhD CONTRACTING ORGANIZATION ...Novel Therapeutic Approaches in Small Cell Lung 5a. CONTRACT NUMBER W81XWH-13-1-0325 Carcinoma Using Genetically Engineered Mouse Models and 5b

Performance evaluation of the Abbott CELL-DYN Emerald for use as a bench-top analyzer in a research setting.

PubMed

Khoo, T-L; Xiros, N; Guan, F; Orellana, D; Holst, J; Joshua, D E; Rasko, J E J

2013-08-01

The CELL-DYN Emerald is a compact bench-top hematology analyzer that can be used for a three-part white cell differential analysis. To determine its utility for analysis of human and mouse samples, we evaluated this machine against the larger CELL-DYN Sapphire and Sysmex XT2000iV hematology analyzers. 120 human (normal and abnormal) and 30 mouse (normal and abnormal) samples were analyzed on both the CELL-DYN Emerald and CELL-DYN Sapphire or Sysmex XT2000iV analyzers. For mouse samples, the CELL-DYN Emerald analyzer required manual recalibration based on the histogram populations. Analysis of the CELL-DYN Emerald showed excellent precision, within accepted ranges (white cell count CV% = 2.09%; hemoglobin CV% = 1.68%; platelets CV% = 4.13%). Linearity was excellent (R² ≥ 0.99), carryover was minimal (<1%), and overall interinstrument agreement was acceptable for both human and mouse samples. Comparison between the CELL-DYN Emerald and Sapphire analyzers for human samples or Sysmex XT2000iV analyzer for mouse samples showed excellent correlation for all parameters. The CELL-DYN Emerald was generally comparable to the larger reference analyzer for both human and mouse samples. It would be suitable for use in satellite research laboratories or as a backup system in larger laboratories. © 2012 John Wiley & Sons Ltd.

Efficient CRISPR-mediated mutagenesis in primary immune cells using CrispRGold and a C57BL/6 Cas9 transgenic mouse line.

PubMed

Chu, Van Trung; Graf, Robin; Wirtz, Tristan; Weber, Timm; Favret, Jeremy; Li, Xun; Petsch, Kerstin; Tran, Ngoc Tung; Sieweke, Michael H; Berek, Claudia; Kühn, Ralf; Rajewsky, Klaus

2016-11-01

Applying clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9)-mediated mutagenesis to primary mouse immune cells, we used high-fidelity single guide RNAs (sgRNAs) designed with an sgRNA design tool (CrispRGold) to target genes in primary B cells, T cells, and macrophages isolated from a Cas9 transgenic mouse line. Using this system, we achieved an average knockout efficiency of 80% in B cells. On this basis, we established a robust small-scale CRISPR-mediated screen in these cells and identified genes essential for B-cell activation and plasma cell differentiation. This screening system does not require deep sequencing and may serve as a precedent for the application of CRISPR/Cas9 to primary mouse cells.

EGF targeted fluorescence molecular tomography as a predictor of PDT outcomes in pancreas cancer models

NASA Astrophysics Data System (ADS)

Samkoe, Kimberley S.; Davis, Scott C.; Srinivasan, Subhadra; Isabelle, Martin E.; O'Hara, Julia; Hasan, Tayyaba; Pogue, Brian W.

2010-02-01

Verteporfin photodynamic therapy (PDT) is a promising adjuvant therapy for pancreas cancer and investigations for its use are currently underway in both orthotopic xenograft mouse models and in human clinical trials. The mouse models have been studied extensively using magnetic resonance (MR) imaging as a measure of surrogate response to verteporfin PDT and it was found that tumor lines with different levels of aggression respond with varying levels to PDT. MR imaging was successful in determining the necrotic volume caused by PDT but there was difficultly in distinguishing inflamed tissues and regions of surviving tumor. In order to understand the molecular changes within the tumor immediately post-PDT we propose the implementation of MR-guided fluorescence molecular tomography (FMT) in conjunction with an exogenously administered fluorescently labeled epidermal growth factor (EGF-IRDye800CW, LI-COR Biosciences). We have previously shown that MR-guided FMT is feasible in the mouse abdomen when multiple regions of fluorescence are considered from contributing internal organs. In this case the highly aggressive AsPC-1 (+EGFR) orthotopic tumor was implanted in SCID mice, interstitial verteporfin PDT (1mg/kg, 20J/cm) was performed when the tumor reached ~60mm3 and both tumor volume and EGF binding were followed with MR-guided FMT.

Mouse senile amyloid fibrils deposited in skeletal muscle exhibit amyloidosis-enhancing activity.

PubMed

Qian, Jinze; Yan, Jingmin; Ge, Fengxia; Zhang, Beiru; Fu, Xiaoying; Tomozawa, Hiroshi; Sawashita, Jinko; Mori, Masayuki; Higuchi, Keiichi

2010-05-20

Amyloidosis describes a group of protein folding diseases in which amyloid proteins are abnormally deposited in organs and/or tissues as fine fibrils. Mouse senile amyloidosis is a disorder in which apolipoprotein A-II (apoA-II) deposits as amyloid fibrils (AApoAII) and can be transmitted from one animal to another both by the feces and milk excreted by mice with amyloidosis. Thus, mouse AApoAII amyloidosis has been demonstrated to be a "transmissible disease". In this study, to further characterize the transmissibility of amyloidosis, AApoAII amyloid fibrils were injected into transgenic Apoa2(c)Tg(+/-) and normal R1.P1-Apoa2(c) mice to induce AApoAII systemic amyloidosis. Two months later, AApoAII amyloid deposits were found in the skeletal muscles of amyloid-affected mice, primarily in the blood vessels and in the interstitial tissues surrounding muscle fibers. When amyloid fibrils extracted from the skeletal muscles were subjected to Western blot analysis, apoA-II was detected. Amyloid fibril fractions isolated from the muscles not only demonstrated the structure of amyloid fibrils but could also induce amyloidosis in young mice depending on its fibril conformation. These findings present a possible pathogenesis of amyloidosis: transmission of amyloid fibril conformation through muscle, and shed new light on the etiology involved in amyloid disorders.

Radiation damage to the microvasculature in the rabbit ear chamber. An electron microscope study. [X radiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, V.V.; Stearner, S.P.; Dimitrievich, G.S.

1977-04-01

Cell aggregates in increased numbers appear along blood vessel walls within a few days after local x irradiation of the tissue within rabbit ear chambers. At 7 days after irradiation with 400 or 700 rad of 250 kVp of x rays, electron microscopic studies of the microvasculature were carried out to determine the morphological characteristics of the cell types involved in the aggregates and the relation of these cells to vascular repair. The cell aggregates usually occur in the interstitial region subjacent to the endothelium. The cells that make up the aggregates show morphological characteristics of relatively undifferentiated mesenchymal cells;more » they have an irregularly rounded shape and contain large amounts of rough endoplasmic reticulum, Golgi vesicles, and mitochondria. In a few instances, cells of similar morphology also occur as part of the lining of the blood vessels. The perivascular cell aggregates may originate from the pericyte population or from undifferentiated mesenchymal cells that occur in the interstitial region surrounding blood vessels; it is improbable that they are dedifferentiated smooth muscle cells. It is suggested that the cells that make up these aggregates contribute to the repair of the microvasculation after radiation injury. The radiosensitivity of vascular endothelium reported by previous investigators seems to preclude endothelial proliferation as the principal repair mechanism at higher radiation doses.« less

CCL11 elicits secretion of RNases from mouse eosinophils and their cell-free granules

PubMed Central

Shamri, Revital; Melo, Rossana C. N.; Young, Kristen M.; Bivas-Benita, Maytal; Xenakis, Jason J.; Spencer, Lisa A.; Weller, Peter F.

2012-01-01

Rapid secretion of eosinophil-associated RNases (EARs), such as the human eosinophilic cationic protein (ECP), from intracellular granules is central to the role of eosinophils in allergic diseases and host immunity. Our knowledge regarding allergic inflammation has advanced based on mouse experimental models. However, unlike human eosinophils, capacities of mouse eosinophils to secrete granule proteins have been controversial. To study mechanisms of mouse eosinophil secretion and EAR release, we combined an RNase assay of mouse EARs with ultrastructural studies. In vitro, mouse eosinophils stimulated with the chemokine eotaxin-1 (CCL11) secreted enzymatically active EARs (EC50 5 nM) by piecemeal degranulation. In vivo, in a mouse model of allergic airway inflammation, increased airway eosinophil infiltration (24-fold) correlated with secretion of active RNases (3-fold). Moreover, we found that eosinophilic inflammation in mice can involve eosinophil cytolysis and release of cell-free granules. Cell-free mouse eosinophil granules expressed functional CCR3 receptors and secreted their granule proteins, including EAR and eosinophil peroxidase in response to CCL11. Collectively, these data demonstrate chemokine-dependent secretion of EARs from both intact mouse eosinophils and their cell-free granules, findings pertinent to understanding the pathogenesis of eosinophil-associated diseases, in which EARs are key factors.—Shamri, R., Melo, R. C. N., Young, K. M., B.-B, M., Xenakis, J. J., Spencer, L. A., Weller, P. F. CCL11 elicits secretion of RNases from mouse eosinophils and their cell-free granules. PMID:22294786

Membrane-To-Nucleus Signaling Links Insulin-Like Growth Factor-1- and Stem Cell Factor-Activated Pathways

PubMed Central

Hayashi, Yujiro; Asuzu, David T.; Gibbons, Simon J.; Aarsvold, Kirsten H.; Bardsley, Michael R.; Lomberk, Gwen A.; Mathison, Angela J.; Kendrick, Michael L.; Shen, K. Robert; Taguchi, Takahiro; Gupta, Anu; Rubin, Brian P.; Fletcher, Jonathan A.; Farrugia, Gianrico; Urrutia, Raul A.; Ordog, Tamas

2013-01-01

Stem cell factor (mouse: Kitl, human: KITLG) and insulin-like growth factor-1 (IGF1), acting via KIT and IGF1 receptor (IGF1R), respectively, are critical for the development and integrity of several tissues. Autocrine/paracrine KITLG-KIT and IGF1-IGF1R signaling are also activated in several cancers including gastrointestinal stromal tumors (GIST), the most common sarcoma. In murine gastric muscles, IGF1 promotes Kitl-dependent development of interstitial cells of Cajal (ICC), the non-neoplastic counterpart of GIST, suggesting cooperation between these pathways. Here, we report a novel mechanism linking IGF1-IGF1R and KITLG-KIT signaling in both normal and neoplastic cells. In murine gastric muscles, the microenvironment for ICC and GIST, human hepatic stellate cells (LX-2), a model for cancer niches, and GIST cells, IGF1 stimulated Kitl/KITLG protein and mRNA expression and promoter activity by activating several signaling pathways including AKT-mediated glycogen synthase kinase-3β inhibition (GSK3i). GSK3i alone also stimulated Kitl/KITLG expression without activating mitogenic pathways. Both IGF1 and GSK3i induced chromatin-level changes favoring transcriptional activation at the Kitl promoter including increased histone H3/H4 acetylation and H3 lysine (K) 4 methylation, reduced H3K9 and H3K27 methylation and reduced occupancy by the H3K27 methyltransferase EZH2. By pharmacological or RNA interference-mediated inhibition of chromatin modifiers we demonstrated that these changes have the predicted impact on KITLG expression. KITLG knock-down and immunoneutralization inhibited the proliferation of GIST cells expressing wild-type KIT, signifying oncogenic autocrine/paracrine KITLG-KIT signaling. We conclude that membrane-to-nucleus signaling involving GSK3i establishes a previously unrecognized link between the IGF1-IGF1R and KITLG-KIT pathways, which is active in both physiologic and oncogenic contexts and can be exploited for therapeutic purposes. PMID:24116170

Inhibition of Necroptosis Attenuates Kidney Inflammation and Interstitial Fibrosis Induced By Unilateral Ureteral Obstruction.

PubMed

Xiao, Xia; Du, Chunyang; Yan, Zhe; Shi, Yonghong; Duan, Huijun; Ren, Yunzhuo

2017-01-01

Inflammation plays a crucial role in renal interstitial fibrosis, the pathway of chronic kidney diseases. Necroptosis is a novel form of regulated cell death, which plays a potential role in inflammation and renal diseases. The small molecule necrostatin-1 (Nec-1) is a specific inhibitor of necroptosis. This study was aimed at determining the role of necroptosis, RIP1/RIP3/mixed lineage kinase domain-like (MLKL) signaling pathway, in renal inflammation and interstitial fibrosis related to primitive tubulointerstitial injury. It was also aimed at evaluating the effect of Nec-1 in renal fibrosis induced by unilateral ureteral obstruction (UUO). Renal histology, immunohistochemistry, western blot, and real-time polymerase chain reaction were performed using UUO C57BL/6J mice model. Moreover, we tested whether Nec-1 was renal-protective in the interstitial fibrosis kidney. Mice were exposed to UUO and injected intraperitoneal with Nec-1 or vehicle. The levels of RIP1/RIP3/MLKL protein and mRNA were increased in the obstructed kidneys 7 days after UUO; this was accompanied by changes in renal pathological lesions. Renal histological examination showed lesser renal damage in Nec-1-treated UUO mice. Renal inflammation, assessed by tumor necrosis factor-α, interleukin-1β, and monocyte chemotactic protein-1 was markedly attenuated by Nec-1. Furthermore, Nec-1 treatment also significantly reduced TGF-β and α-smooth muscle actin, indicating lesser renal interstitial fibrosis. These findings suggest that the participation of necroptosis in UUO is partly demonstrated. And necroptosis inhibition may have a potential role in the treatment of diseases with increased inflammatory response and interstitial fibrosis in renal. © 2017 S. Karger AG, Basel.

Micropattern differentiation of mouse pluripotent stem cells recapitulates embryo regionalized cell fate patterning

PubMed Central

Morgani, Sophie M; Metzger, Jakob J; Nichols, Jennifer

2018-01-01

During gastrulation epiblast cells exit pluripotency as they specify and spatially arrange the three germ layers of the embryo. Similarly, human pluripotent stem cells (PSCs) undergo spatially organized fate specification on micropatterned surfaces. Since in vivo validation is not possible for the human, we developed a mouse PSC micropattern system and, with direct comparisons to mouse embryos, reveal the robust specification of distinct regional identities. BMP, WNT, ACTIVIN and FGF directed mouse epiblast-like cells to undergo an epithelial-to-mesenchymal transition and radially pattern posterior mesoderm fates. Conversely, WNT, ACTIVIN and FGF patterned anterior identities, including definitive endoderm. By contrast, epiblast stem cells, a developmentally advanced state, only specified anterior identities, but without patterning. The mouse micropattern system offers a robust scalable method to generate regionalized cell types present in vivo, resolve how signals promote distinct identities and generate patterns, and compare mechanisms operating in vivo and in vitro and across species. PMID:29412136

Therapeutic effects of autologous lymphocytes activated with trastuzumab for xenograft mouse models of human breast cancer.

PubMed

Nakagawa, Shinichiro; Matsuoka, Yusuke; Ichihara, Hideaki; Yoshida, Hitoji; Yoshida, Kenshi; Ueoka, Ryuichi

2013-01-01

Trastuzumab (TTZ) is molecular targeted drug used for metastatic breast cancer patients overexpressing human epidermal growth factor receptor 2 (HER2). Therapeutic effects of lymphocytes activated with TTZ (TTZ-LAK) using xenograft mouse models of human breast cancer (MDA-MB-453) cells were examined in vivo. Remarkable reduction of tumor volume in a xenograft mouse models intravenously treated with TTZ-LAK cells after the subcutaneously inoculated of MDA-MB-453 cells was verified in vivo. The migration of TTZ-LAK cells in tumor of mouse models subcutaneously inoculated MDA-MB-453 cells was observed on the basis of histological analysis using immunostaining with CD-3. Induction of apoptosis in tumor of xenograft mice treated with TTZ-LAK cells was observed in micrographs using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) method. It was noteworthy that the therapeutic effects of TTZ-LAK cells along with apoptosis were obtained for xenograft mouse models of human breast tumor in vivo.

Treatment of hypoxic-ischemic encephalopathy in mouse by transplantation of embryonic stem cell-derived cells.

PubMed

Ma, Jie; Wang, Yu; Yang, Jianhua; Yang, Min; Chang, Keun-A; Zhang, Linhua; Jiang, Feng; Li, Yi; Zhang, Zhonggong; Heo, Chaejeong; Suh, Yoo-Hun

2007-07-01

A 7-day-old hypoxic-ischemic encephalopathy (HIE) mouse model was used to study the effect of transplantation of embryonic stem (ES) cell-derived cells on the HIE. After the inducement in vitro, the ES cell-derived cells expressed Nestin and MAP-2, rather than GFAP mRNA. After transplantation, ES cell-derived cells can survive, migrate into the injury site, and specifically differentiate into neurons, showing improvement of the learning ability and memory of the HIE mouse at 8 months post-transplantation. The non-grafted HIE mouse brain showed typical pathological changes in the hippocampus and cerebral cortex, where the number of neurons was reduced, while in the cell graft group, number of the neurons increased in the same regions. Although further study is necessary to elucidate the precise mechanisms responsible for this functional recovery, we believe that ES cells have advantages for use as a donor source in HIE.

Progressive renal papillary calcification and ureteral stone formation in mice deficient for Tamm-Horsfall protein

PubMed Central

Liu, Yan; Mo, Lan; Goldfarb, David S.; Evan, Andrew P.; Liang, Fengxia; Khan, Saeed R.; Lieske, John C.

2010-01-01

Mammalian urine contains a range of macromolecule proteins that play critical roles in renal stone formation, among which Tamm-Horsfall protein (THP) is by far the most abundant. While THP is a potent inhibitor of crystal aggregation in vitro and its ablation in vivo predisposes one of the two existing mouse models to spontaneous intrarenal calcium crystallization, key controversies remain regarding the role of THP in nephrolithiasis. By carrying out a long-range follow-up of more than 250 THP-null mice and their wild-type controls, we demonstrate here that renal calcification is a highly consistent phenotype of the THP-null mice that is age and partially gene dosage dependent, but is gender and genetic background independent. Renal calcification in THP-null mice is progressive, and by 15 mo over 85% of all the THP-null mice develop spontaneous intrarenal crystals. The crystals consist primarily of calcium phosphate in the form of hydroxyapatite, are located more frequently in the interstitial space of the renal papillae than intratubularly, particularly in older animals, and lack accompanying inflammatory cell infiltration. The interstitial deposits of hydroxyapatite observed in THP-null mice bear strong resemblances to the renal crystals found in human kidneys bearing idiopathic calcium oxalate stones. Compared with 24-h urine from the wild-type mice, that of THP-null mice is supersaturated with brushite (calcium phosphate), a stone precursor, and has reduced urinary excretion of citrate, a stone inhibitor. While less frequent than renal calcinosis, renal pelvic and ureteral stones and hydronephrosis occur in the aged THP-null mice. These results provide direct in vivo evidence indicating that normal THP plays an important role in defending the urinary system against calcification and suggest that reduced expression and/or decreased function of THP could contribute to nephrolithiasis. PMID:20591941

SU-E-J-197: A Novel Optical Interstitial Fiber Spectroscopic System for Real-Time Tissue Micro-Vascular Hemodynamics Monitoring.

PubMed

Zhao, D; Campos, D; Yan, Y; Kimple, R; Jacques, S; van der Kogel, A; Kissick, M

2012-06-01

To demonstrate a novel interstitial optical fiber spectroscopic system, based on diffuse optical spectroscopies with spectral fitting, for the simultaneous monitoring of tumor blood volume and oxygen tension. The technique provides real-time, minimally-invasive and quantification of tissue micro-vascular hemodynamics. An optical fiber prototype probe characterizesthe optical transport in tissue between two large Numerical Aperture (NA) fibers of 200μm core diameter (BFH37-200, ThorLabs) spaced 3-mm apart. Two 21-Ga medical needles are used to protect fiber ends and to facilitate tissue penetration with minimum local blunt trauma in nude mice with xenografts. A 20W white light source (HL-2000-HP, Ocean Optics) is coupled to one fiber with SMA adapter. The other fiber is used to collect light, which is coupled into the spectrometer (QE65000 with Spectrasuite Operating software and OmniDriver, Ocean Optics). The wavelength response of the probe depends on the wavelength dependence of the light source, and of the light signal collection that includes considerable scatter, modeled with Monte-Carlo techniques (S. Jacques 2010 J. of Innov. Opt. Health Sci. 2 123-9). Measured spectra of tissue are normalized by a measured spectrum of a white standard, yielding the transmission spectrum. A head-and-neck xenograft on the flank of a live mouse is used for development. The optical fiber probe delivers and collects light at an arbitrary depth in the tumor. By spectral fitting of the measured transmission spectrum, an analysis of blood volume and oxygen tension is obtained from the fitting parameters in real time. A newly developed optical fiber spectroscopic system with an optical fiber probe takes spectroscopic techniques to a much deeper level in a tumor, which has potential applications for real-time monitoring hypoxic cell population dynamics for an eventual adaptive therapy metric of particular use in hypofractionated radiotherapy. © 2012 American Association of Physicists in Medicine.

Establishment of mouse expanded potential stem cells

PubMed Central

Gao, Xuefei; Antunes, Liliana; Yu, Yong; Zhu, Zhexin; Wang, Juexuan; Kolodziejczyk, Aleksandra A.; Campos, Lia S.; Wang, Cui; Yang, Fengtang; Zhong, Zhen; Fu, Beiyuan; Eckersley-Maslin, Melanie A.; Woods, Michael; Tanaka, Yosuke; Chen, Xi; Wilkinson, Adam C.; Bussell, James; White, Jacqui; Ramirez-Solis, Ramiro; Reik, Wolf; Göttgens, Berthold; Teichmann, Sarah A.; Tam, Patrick P. L.; Nakauchi, Hiromitsu; Zou, Xiangang; Lu, Liming; Liu, Pentao

2018-01-01

Mouse embryonic stem cells derived from the epiblast1 contribute to the somatic lineages and the germline but are excluded from the extra-embryonic tissues that are derived from the trophectoderm and the primitive endoderm2 upon reintroduction to the blastocyst. Here we report that cultures of expanded potential stem cells can be established from individual eight-cell blastomeres, and by direct conversion of mouse embryonic stem cells and induced pluripotent stem cells. Remarkably, a single expanded potential stem cell can contribute both to the embryo proper and to the trophectoderm lineages in a chimaera assay. Bona fide trophoblast stem cell lines and extra-embryonic endoderm stem cells can be directly derived from expanded potential stem cells in vitro. Molecular analyses of the epigenome and single-cell transcriptome reveal enrichment for blastomere-specific signature and a dynamic DNA methylome in expanded potential stem cells. The generation of mouse expanded potential stem cells highlights the feasibility of establishing expanded potential stem cells for other mammalian species. PMID:29019987

Testicular leiomyosarcoma and marked alopecia in a cryptorchid ferret (Mustela putorius furo).

PubMed

Kammeyer, P; Ziege, S; Wellhöner, S; Cichowski, S; Baumgärtner, W

2014-01-01

A 3.5-year-old male ferret, bought as male castrated, was presented to the veterinarian with marked alopecia of back, neck, abdomen and tail, a pronounced sexual behaviour and weight loss. An inguinal mass of about 2.5 cm in diameter was diagnosed as potentially tumorous inguinal testicle by ultrasound and fine-needle aspiration. Adrenal glands and prostate were ultrasonographically unremarkable. The surgically removed cryptorchid testicle contained a greyish tumour that was histologically composed of spindle-shaped cells with elongated nuclei, embedded in a fibro-vascular stroma. Up to two mitotic figures per high power field were noted. Additionally, an interstitial cell hyperplasia and marked reactive proliferation of a collagen-rich fibrous tissue were observed. Tumour cells were positive for α-smooth muscle actin, desmin, and occasionally vimentin and S-100, leading to the diagnosis of an intratesticular leiomyosarcoma. As an adrenal-associated endocrinopathy was excluded and a complete fur recovery was observed after removal of the cryptorchid testicle the alopecia was eventually due to hormones produced by the hyperplastic interstitial (Leydig) cells.

38 CFR 4.115b - Ratings of the genitourinary system-diagnoses.

Code of Federal Regulations, 2014 CFR

2014-07-01

.... 7537Interstitial nephritis: Rate as renal dysfunction. 7538Papillary necrosis: Rate as renal dysfunction. 7539Renal... necrosis: Rate as renal dysfunction. 7541Renal involvement in diabetes mellitus, sickle cell anemia...

38 CFR 4.115b - Ratings of the genitourinary system-diagnoses.

Code of Federal Regulations, 2012 CFR

2012-07-01

.... 7537Interstitial nephritis: Rate as renal dysfunction. 7538Papillary necrosis: Rate as renal dysfunction. 7539Renal... necrosis: Rate as renal dysfunction. 7541Renal involvement in diabetes mellitus, sickle cell anemia...

38 CFR 4.115b - Ratings of the genitourinary system-diagnoses.

Code of Federal Regulations, 2013 CFR

2013-07-01

.... 7537Interstitial nephritis: Rate as renal dysfunction. 7538Papillary necrosis: Rate as renal dysfunction. 7539Renal... necrosis: Rate as renal dysfunction. 7541Renal involvement in diabetes mellitus, sickle cell anemia...

Sunitinib-Induced Acute Interstitial Nephritis in a Thrombocytopenic Renal Cell Cancer Patient.

PubMed

Azar, Ibrahim; Esfandiarifard, Saghi; Sinai, Pedram; Wazir, Ali; Foulke, Llewellyn; Mehdi, Syed

2017-01-01

Sunitinib, a multitargeted tyrosine kinase inhibitor (TKI), is currently the standard of care for patients with metastatic renal cell carcinoma. Renal adverse events associated with sunitinib include proteinuria, renal insufficiency secondary to focal segmental glomerulosclerosis (FSGS), and thrombotic microangiopathy. We describe the second reported instance of biopsy-proven sunitinib-induced acute interstitial nephritis (AIN), in a challenging case complicated by thrombocytopenia. The case illustrates the importance of early diagnosis and intervention in ensuring long-term recovery from renal complications. Four other cases of AIN reported along with inhibition of the vascular endothelial growth factor (VEGF) by either TKI (sunitinib and sorafenib) or antibodies (bevacizumab) suggest a possible class effect. Given our experience, we recommend monitoring renal function with VEGF inhibition, and in the case of renal failure in the setting of an unclear diagnosis, we recommend prompt biopsy.

Sudden death of an Indian peafowl (Pavo cristatus) at a zoo due to non-pigmented Serratia marcescens infection

PubMed Central

LEE, Seung-Hun; PARK, Sang-Joon; KWAK, Dongmi; KIM, Kyoo-Tae

2017-01-01

A 16-year-old female Indian peafowl (Pavo cristatus) died two days after recognition of conjunctivitis in the right eye, anorexia and depression. Gross necropsy revealed a thick pseudomembrane under the eyelid and hydropericardium. Histopathological examination revealed hepatocellular necrosis, sinusoidal and vascular congestion and infiltrated inflammatory cells. Infiltration by inflammatory cells was noted in the epicardium. The lungs had mild interstitial pneumonia with the extensive congestion within the capillaries of the air sacs. Tubular interstitial congestion and necrosis was noted in the kidneys. Bacterial culture and nucleotide sequencing of the inflammatory specimens identified the causative agent as Serratia marcescens, an uncommon bacterium in birds. In summary, this study describes the sudden death of an Indian peafowl due to S. marcescens infection, which is rarely seen in animals. PMID:29081475

Mefunidone Attenuates Tubulointerstitial Fibrosis in a Rat Model of Unilateral Ureteral Obstruction

PubMed Central

Liu, Chunyan; Mei, Wenjuan; Tang, Juan; Yuan, Qiongjing; Huang, Ling; Lu, Miaomiao; Wu, Lin; Peng, Zhangzhe; Meng, Jie; Yang, Huixiang; Shen, Hong; Lv, Ben; Hu, Gaoyun; Tao, Lijian

2015-01-01

Background Inflammation has a crucial role in renal interstitial fibrosis, which is the common pathway of chronic kidney diseases. Mefunidone (MFD) is a new compound which could effectively inhibit the proliferation of renal fibroblasts in vitro. However, the overall effect of Mefunidone in renal fibrosis remains unknown. Methods Sprague-Dawley rats were randomly divided intro 6 groups: sham operation, unilateral ureteral obstruction (UUO), UUO/Mefunidone (25, 50, 100mg/kg/day) and UUO/PFD (500mg/kg/day). The rats were sacrificed respectively on days 3, 7, and 14 after the operation. Tubulointerstitial injury index, interstitial collagen deposition, expression of fibronectin (FN), α-smooth muscle actin (α-SMA), type I and III collagen and the number of CD3+ and CD68+ cells were determined. The expressions of proinflammatory cytokines, p-ERK, p-IκB, and p-STAT3 were measured in human renal proximal tubular epithelial cells of HK-2 or macrophages. Results Mefunidone treatment significantly attenuated tubulointerstitial injury, interstitial collagen deposition, expression of FN, α-SMA, type I and III collagen in the obstructive kidneys, which correlated with significantly reduced the number of T cells and macrophages in the obstructive kidneys. Mechanistically, Mefunidone significantly inhibited tumor necrosis factor-α (TNF-α-) or lipopolysaccharide (LPS)-induced production of proinflammatory cytokines. This effect is possibly due to the inhibition of phosphorylation of ERK, IκB, and STAT3. Conclusion Mefunidone treatment attenuated tubulointerstitial fibrosis in a rat model of UUO, at least in part, through inhibition of inflammation. PMID:26042668

Roles of interleukin-9 in the growth and cholecystokinin-induced intracellular calcium signaling of cultured interstitial cells of Cajal.

PubMed

Gong, Yaoyao; Huang, Lei; Cheng, Wenfang; Li, Xueliang; Lu, Jia; Lin, Lin; Si, Xinmin

2014-01-01

Interstitial cells of Cajal (ICC) are pacemaker cells in the gastrointestinal (GI) tract and loss of ICC is associated with many GI motility disorders. Previous studies have shown that ICC have the capacity to regenerate or restore, and several growth factors are critical to their growth, maintenance or regeneration. The present study aimed to investigate the roles of interleukin-9 (IL-9) in the growth, maintenance and pacemaker functions of cultured ICC. Here, we report that IL-9 promotes proliferation of ICC, and culturing ICC with IL-9 enhances cholecystokinin-8-induced Ca²⁺ transients, which is probably caused by facilitating maintenance of ICC functions under culture condition. We also show co-localizations of cholecystokinin-1 receptor and IL-9 receptor with c-kit by double-immunohistochemical labeling. In conclusion, IL-9 can promote ICC growth and help maintain ICC functions; IL-9 probably performs its functions via IL-9 receptors on ICC.

Grape seed extract ameliorates bleomycin-induced mouse pulmonary fibrosis.

PubMed

Liu, Qi; Jiang, Jun-Xia; Liu, Ya-Nan; Ge, Ling-Tian; Guan, Yan; Zhao, Wei; Jia, Yong-Liang; Dong, Xin-Wei; Sun, Yun; Xie, Qiang-Min

2017-05-05

Pulmonary fibrosis is common in a variety of inflammatory lung diseases, such as interstitial pneumonia, chronic obstructive pulmonary disease, and silicosis. There is currently no effective clinical drug treatment. It has been reported that grape seed extracts (GSE) has extensive pharmacological effects with minimal toxicity. Although it has been found that GSE can improve the lung collagen deposition and fibrosis pathology induced by bleomycin in rat, its effects on pulmonary function, inflammation, growth factors, matrix metalloproteinases and epithelial-mesenchymal transition remain to be researched. In the present study, we studied whether GSE provided protection against bleomycin (BLM)-induced mouse pulmonary fibrosis. ICR strain mice were treated with BLM in order to establish pulmonary fibrosis models. GSE was given daily via intragastric administration for three weeks starting at one day after intratracheal instillation. GSE at 50 or 100mg/kg significantly reduced BLM-induced inflammatory cells infiltration, proinflammatory factor protein expression, and hydroxyproline in lung tissues, and improved pulmonary function in mice. Additionally, treatment with GSE also significantly impaired BLM-induced increases in lung fibrotic marker expression (collagen type I alpha 1 and fibronectin 1) and decreases in an anti-fibrotic marker (E-cadherin). Further investigation indicated that the possible molecular targets of GSE are matrix metalloproteinases-9 (MMP-9) and TGF-β1, given that treatment with GSE significantly prevented BLM-induced increases in MMP-9 and TGF-β1 expression in the lungs. Together, these results suggest that supplementation with GSE may improve the quality of life of lung fibrosis patients by inhibiting MMP-9 and TGF-β1 expression in the lungs. Copyright © 2017 Elsevier B.V. All rights reserved.

β-Liddle mutation of the epithelial sodium channel increases alveolar fluid clearance and reduces the severity of hydrostatic pulmonary oedema in mice

PubMed Central

Randrianarison, Nadia; Escoubet, Brigitte; Ferreira, Chrystophe; Fontayne, Alexandre; Fowler-Jaeger, Nicole; Clerici, Christine; Hummler, Edith; Rossier, Bernard C; Planès, Carole

2007-01-01

Transepithelial sodium transport via alveolar epithelial Na+ channels and Na+,K+-ATPase constitutes the driving force for removal of alveolar oedema fluid. Decreased activity of the amiloride-sensitive epithelial Na+ channel (ENaC) in the apical membrane of alveolar epithelial cells impairs sodium-driven alveolar fluid clearance (AFC) and predisposes to pulmonary oedema. We hypothesized that hyperactivity of ENaC in the distal lung could improve AFC and facilitate the resolution of pulmonary oedema. AFC and lung fluid balance were studied at baseline and under conditions of hydrostatic pulmonary oedema in the β-Liddle (L) mouse strain harbouring a gain-of-function mutation (R566stop) within the Scnn1b gene. As compared with wild-type (+/+), baseline AFC was increased by 2- and 3-fold in heterozygous (+/L) and homozygous mutated (L/L) mice, respectively, mainly due to increased amiloride-sensitive AFC. The β2-agonist terbutaline stimulated AFC in +/+ and +/L mice, but not in L/L mice. Acute volume overload induced by saline infusion (40% of body weight over 2 h) significantly increased extravascular (i.e. interstitial and alveolar) lung water as assessed by the bloodless wet-to-dry lung weight ratio in +/+ and L/L mice, as compared with baseline. However, the increase was significantly larger in +/+ than in L/L groups (P= 0.01). Volume overload also increased the volume of the alveolar epithelial lining fluid in +/+ mice, indicating the presence of alveolar oedema, but not in L/L mice. Cardiac function as evaluated by echocardiography was comparable in both groups. These data show that constitutive ENaC activation improved sodium-driven AFC in the mouse lung, and attenuated the severity of hydrostatic pulmonary oedema. PMID:17430990

Cerebellar stem cells do not produce neurons and astrocytes in adult mouse

DOE Office of Scientific and Technical Information (OSTI.GOV)

Su, Xin; Guan, Wuqiang; Yu, Yong-Chun

Highlights: • No new neurons and astrocytes are generated in adult mouse cerebellum. • Very few mash1{sup +} or nestin{sup +} stem cells exist, and most of them are quiescent. • Cell proliferation rate is diversified among cerebellar regions and decreases over time. - Abstract: Although previous studies implied that cerebellar stem cells exist in some adult mammals, little is known about whether these stem cells can produce new neurons and astrocytes. In this study by bromodeoxyuridine (BrdU) intraperitoneal (i.p.) injection, we found that there are abundant BrdU{sup +} cells in adult mouse cerebellum, and their quantity and density decreasesmore » significantly over time. We also found cell proliferation rate is diversified in different cerebellar regions. Among these BrdU{sup +} cells, very few are mash1{sup +} or nestin{sup +} stem cells, and the vast majority of cerebellar stem cells are quiescent. Data obtained by in vivo retrovirus injection indicate that stem cells do not produce neurons and astrocytes in adult mouse cerebellum. Instead, some cells labeled by retrovirus are Iba1{sup +} microglia. These results indicate that very few stem cells exist in adult mouse cerebellum, and none of these stem cells contribute to neurogenesis and astrogenesis under physiological condition.« less

Reprogrammed mouse astrocytes retain a "memory" of tissue origin and possess more tendencies for neuronal differentiation than reprogrammed mouse embryonic fibroblasts.

PubMed

Tian, Changhai; Wang, Yongxiang; Sun, Lijun; Ma, Kangmu; Zheng, Jialin C

2011-02-01

Direct reprogramming of a variety of somatic cells with the transcription factors Oct4 (also called Pou5f1), Sox2 with either Klf4 and Myc or Lin28 and Nanog generates the induced pluripotent stem cells (iPSCs) with marker similarity to embryonic stem cells. However, the difference between iPSCs derived from different origins is unclear. In this study, we hypothesized that reprogrammed cells retain a "memory" of their origins and possess additional potential of related tissue differentiation. We reprogrammed primary mouse astrocytes via ectopic retroviral expression of OCT3/4, Sox2, Klf4 and Myc and found the iPSCs from mouse astrocytes expressed stem cell markers and formed teratomas in SCID mice containing derivatives of all three germ layers similar to mouse embryonic stem cells besides semblable morphologies. To test our hypothesis, we compared embryonic bodies (EBs) formation and neuronal differentiation between iPSCs from mouse embryonic fibroblasts (MEFsiPSCs) and iPSCs from mouse astrocytes (mAsiPSCs). We found that mAsiPSCs grew slower and possessed more potential for neuronal differentiation compared to MEFsiPSCs. Our results suggest that mAsiPSCs retain a "memory" of the central nervous system, which confers additional potential upon neuronal differentiation.

The role of Cajal cells in chronic prostatitis.

PubMed

Haki Yuksel, Ozgur; Urkmez, Ahmet; Verit, Ayhan

2016-07-04

Types of prostatitis can be defined as groups of syndromes in adult men associated with infectious and noninfectious causes characterized frequently by lower abdominal and perineal signs and diverse clinical symptoms and complications. Etiopathogenesis of chronic prostatitis is not well defined. Moreover, its treatment outcomes are not satisfactory. Presence of c-kit positive interstitial cells in human prostate is already known. It has been demonstrated that these cells can be pacemaker cells which trigger spontaneous slow-wave electrical activity in the prostate and can be responsible for the transport of glandular secretion from acinar cells into major and minor prostatic ducts and finally into urethra. In the light of all these data, when presence of a possible inflammatory pathology is thought to involve prostate that secretes and has a reservoir which drains its secretion (for prostate, prostatic urethra), two points are worth mentioning. Impairment of secretion mechanism and collection of secretion within the organ with reflux of the microbial material from its reservoir back into prostate gland. Both of these potential conditions can be explained by ductal neuromuscular mechanism, which induces secretion. We think that in this neuromuscular mechanism interstitial Cajal cells have an important role in chronic prostatitis. Our hypothesis is that curability of prostatitis is correlated with the number of Cajal cells not subjected to apoptosis.

Anti-tumor effects of peptide analogs targeting neuropeptide hormone receptors on mouse pheochromocytoma cells.

PubMed

Ziegler, C G; Ullrich, M; Schally, A V; Bergmann, R; Pietzsch, J; Gebauer, L; Gondek, K; Qin, N; Pacak, K; Ehrhart-Bornstein, M; Eisenhofer, G; Bornstein, S R

2013-05-22

Pheochromocytoma is a rare but potentially lethal chromaffin cell tumor with currently no effective treatment. Peptide hormone receptors are frequently overexpressed on endocrine tumor cells and can be specifically targeted by various anti-tumor peptide analogs. The present study carried out on mouse pheochromocytoma cells (MPCs) and a more aggressive mouse tumor tissue-derived (MTT) cell line revealed that these cells are characterized by pronounced expression of the somatostatin receptor 2 (sst2), growth hormone-releasing hormone (GHRH) receptor and the luteinizing hormone-releasing hormone (LHRH) receptor. We further demonstrated significant anti-tumor effects mediated by cytotoxic somatostatin analogs, AN-162 and AN-238, by LHRH antagonist, Cetrorelix, by the cytotoxic LHRH analog, AN-152, and by recently developed GHRH antagonist, MIA-602, on MPC and for AN-152 and MIA-602 on MTT cells. Studies of novel anti-tumor compounds on these mouse cell lines serve as an important basis for mouse models of metastatic pheochromocytoma, which we are currently establishing. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Cardiac remodeling in the mouse model of Marfan syndrome develops into two distinctive phenotypes

PubMed Central

Tae, Hyun-Jin; Marshall, Shannon; Krawczyk, Melissa; Talan, Mark

2015-01-01

Marfan syndrome (MFS) is a systemic disorder of connective tissue caused by mutations in fibrillin-1. Cardiac dysfunction in MFS has not been characterized halting the development of therapies of cardiac complication in MFS. We aimed to study the age-dependent cardiac remodeling in the mouse model of MFS FbnC1039G+/− mouse [Marfan heterozygous (HT) mouse] and its association with valvular regurgitation. Marfan HT mice of 2–4 mo demonstrated a mild hypertrophic cardiac remodeling with predominant decline of diastolic function and increased transforming growth factor-β canonical (p-SMAD2/3) and noncanonical (p-ERK1/2 and p-p38 MAPK) signaling and upregulation of hypertrophic markers natriuretic peptides atrium natriuretic peptide and brain natriuretic peptide. Among older HT mice (6–14 mo), cardiac remodeling was associated with two distinct phenotypes, manifesting either dilated or constricted left ventricular chamber. Dilatation of left ventricular chamber was accompanied by biochemical evidence of greater mechanical stress, including elevated ERK1/2 and p38 MAPK phosphorylation and higher brain natriuretic peptide expression. The aortic valve regurgitation was registered in 20% of the constricted group and 60% of the dilated group, whereas mitral insufficiency was observed in 40% of the constricted group and 100% of the dilated group. Cardiac dysfunction was not associated with the increase of interstitial fibrosis and nonmyocyte proliferation. In the mouse model fibrillin-1, haploinsufficiency results in the early onset of nonfibrotic hypertrophic cardiac remodeling and dysfunction, independently from valvular abnormalities. MFS heart is vulnerable to stress-induced cardiac dilatation in the face of valvular regurgitation, and stress-activated MAPK signals represent a potential target for cardiac management in MFS. PMID:26566724

Cardiac remodeling in the mouse model of Marfan syndrome develops into two distinctive phenotypes.

PubMed

Tae, Hyun-Jin; Petrashevskaya, Natalia; Marshall, Shannon; Krawczyk, Melissa; Talan, Mark

2016-01-15

Marfan syndrome (MFS) is a systemic disorder of connective tissue caused by mutations in fibrillin-1. Cardiac dysfunction in MFS has not been characterized halting the development of therapies of cardiac complication in MFS. We aimed to study the age-dependent cardiac remodeling in the mouse model of MFS FbnC1039G+/- mouse [Marfan heterozygous (HT) mouse] and its association with valvular regurgitation. Marfan HT mice of 2-4 mo demonstrated a mild hypertrophic cardiac remodeling with predominant decline of diastolic function and increased transforming growth factor-β canonical (p-SMAD2/3) and noncanonical (p-ERK1/2 and p-p38 MAPK) signaling and upregulation of hypertrophic markers natriuretic peptides atrium natriuretic peptide and brain natriuretic peptide. Among older HT mice (6-14 mo), cardiac remodeling was associated with two distinct phenotypes, manifesting either dilated or constricted left ventricular chamber. Dilatation of left ventricular chamber was accompanied by biochemical evidence of greater mechanical stress, including elevated ERK1/2 and p38 MAPK phosphorylation and higher brain natriuretic peptide expression. The aortic valve regurgitation was registered in 20% of the constricted group and 60% of the dilated group, whereas mitral insufficiency was observed in 40% of the constricted group and 100% of the dilated group. Cardiac dysfunction was not associated with the increase of interstitial fibrosis and nonmyocyte proliferation. In the mouse model fibrillin-1, haploinsufficiency results in the early onset of nonfibrotic hypertrophic cardiac remodeling and dysfunction, independently from valvular abnormalities. MFS heart is vulnerable to stress-induced cardiac dilatation in the face of valvular regurgitation, and stress-activated MAPK signals represent a potential target for cardiac management in MFS.

Unilateral Renal Ischemia as a Model of Acute Kidney Injury and Renal Fibrosis in Cats.

PubMed

Schmiedt, C W; Brainard, B M; Hinson, W; Brown, S A; Brown, C A

2016-01-01

The objectives of this study were to define the acute and chronic effects of 1-hour unilateral in vivo renal ischemia on renal function and histology in cats. Twenty-one adult purpose-bred research cats were anesthetized, and 1 kidney underwent renal artery and vein occlusion for 1 hour. Serum creatinine and urea concentrations, urine protein:creatinine ratio, urine-specific gravity, glomerular filtration rate, hematocrit, platelet concentration and function, and white blood cell count were measured at baseline and variable time points after ischemia. Renal histopathology was evaluated on days 3, 6, 12, 21, 42, and 70 postischemia; changes in smooth muscle actin and interstitial collagen were examined. Following ischemia, whole animal glomerular filtration rate was significantly reduced (57% of baseline on day 6; P < .05). At the early time points, the ischemic kidneys exhibited severe acute epithelial necrosis accompanied by evidence of regeneration of tubules predominantly within the corticomedullary junction. At later periods, postischemic kidneys had evidence of tubular atrophy and interstitial inflammation with significantly more smooth muscle actin and interstitial collagen staining and interstitial fibrosis when compared with the contralateral control kidneys. This study characterizes the course of ischemic acute kidney injury in cats and demonstrates that ischemic acute kidney injury triggers chronic fibrosis, interstitial inflammation, and tubular atrophy in feline kidneys. These late changes are typical of those observed in cats with naturally occurring chronic kidney disease. © The Author(s) 2015.

Fluorescein angiography of the bladder: technique and relevance to bladder cancer and interstitial cystitis patients.

PubMed

Zimmern, P E; Laub, D; Leach, G E

1995-07-01

Fluorescein angiography has been used in the study of bleeding vessels, neovascularity, tumors and ischemic tissues in a variety of disorders. This pilot study was designed to evaluate the feasibility, safety and relevance of this interesting technology for the evaluation of bladder wall vessels in patients with interstitial cystitis and bladder cancer. Five patients with National Institutes of Health defined interstitial cystitis symptoms and 10 with bladder cancer were studied during cytoscopy while they were under general anesthesia. A yellow-green barrier filter (520 nm.) was placed over the cystoscope eyepiece and a blue exciter filter (465 nm.) was attached to the light source. Patients received a 5 ml. bolus of 10% fluorescein intravenously. After hydrodistension, glomerulations in interstitial cystitis patients were more prominent with fluorescein angiography and occurred in the venule phase. Areas of papillary transitional cell tumor and carcinoma in situ developed a brilliant yellow-green fluorescence. Adjacent normal urothelium was nonfluorescent and provided a contrasting dark background facilitating the detection of all lesions. No allergic reaction or other adverse effect related to the fluorescein injection was observed. These unique observations in a limited number of patients suggest that fluorescein angiography of the bladder is a safe and simple procedure. This preliminary report underscores the relevance of fluorescein angiography in the detection of bladder tumor and offers a new approach to the evaluation of bladder wall vessels in interstitial cystitis patients.

Species-specific differences in peroxisome proliferation, catalase, and SOD2 upregulation as well as toxicity in human, mouse, and rat hepatoma cells induced by the explosive and environmental pollutant 2,4,6-trinitrotoluene.

PubMed

Naumenko, Ekaterina Anatolevna; Ahlemeyer, Barbara; Baumgart-Vogt, Eveline

2017-03-01

2,4,6-Trinitrotoluene (TNT) has been widely used as an explosive substance and its toxicity is still of interest as it persisted in polluted areas. TNT is metabolized in hepatocytes which are prone to its toxicity. Since analysis of the human liver or hepatocytes is restricted due to ethical reasons, we investigated the effects of TNT on cell viability, reactive oxygen species (ROS) production, peroxisome proliferation, and antioxidative enzymes in human (HepG2), mouse (Hepa 1-6), and rat (H4IIEC3) hepatoma cell lines. Under control conditions, hepatoma cells of all three species were highly comparable exhibiting identical proliferation rates and distribution of their cell cycle phases. However, we found strong differences in TNT toxicity with the lowest IC 50 values (highest cell death rate) for rat cells, whereas human and mouse cells were three to sevenfold less sensitive. Moreover, a strong decrease in cellular dehydrogenase activity (MTT assay) and increased ROS levels were noted. TNT caused peroxisome proliferation with rat hepatoma cells being most responsive followed by those from mouse and human. Under control conditions, rat cells contained fivefold higher peroxisomal catalase and mitochondrial SOD2 activities and a twofold higher capacity to reduce MTT than human and mouse cells. TNT treatment caused an increase in catalase and SOD2 mRNA and protein levels in human and mouse, but not in rat cells. Similarly, human and mouse cells upregulated SOD2 activity, whereas rat cells failed therein. We conclude that TNT induced oxidative stress, peroxisome proliferation and mitochondrial damage which are highest in rat cells rendering them most susceptible toward TNT. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 989-1006, 2017. © 2016 Wiley Periodicals, Inc.

Characterization of Breast Cancer Interstitial Fluids by TmT Labeling, LTQ-Orbitrap Velos Mass Spectrometry and Pathway Analysis

PubMed Central

Cinzia, Raso; Carlo, Cosentino; Marco, Gaspari; Natalia, Malara; Xuemei, Han; Daniel, McClatchy; Kyu, Park Sung; Maria, Renne; Nuria, Vadalà; Ubaldo, Prati; Giovanni, Cuda; Vincenzo, Mollace; Francesco, Amato; Yates, John R.

2012-01-01

Cancer is currently considered as the end point of numerous genomic and epigenomic mutations and as the result of the interaction of transformed cells within the stromal microenvironment. The present work focuses on breast cancer, one of the most common malignancies affecting the female population in industrialized countries. In this study we perform a proteomic analysis of bioptic samples from human breast cancer, namely interstitial fluids and primary cells, normal vs disease tissues, using Tandem mass Tags (TmT) quantitative mass spectrometry combined with the MudPIT technique. To the best of our knowledge this work, with over 1700 proteins identified, represents the most comprehensive characterization of the breast cancer interstitial fluid proteome to date. Network analysis was used to identify functionally active networks in the breast cancer associated samples. From the list of differentially expressed genes we have retrieved the associated functional interaction networks. Many different signaling pathways were found activated, strongly linked to invasion, metastasis development, proliferation and with a significant cross-talking rate. This pilot study presents evidence that the proposed quantitative proteomic approach can be applied to discriminate between normal and tumoral samples and for the discovery of yet unknown carcinogenesis mechanisms and therapeutic strategies. PMID:22563702

Sphere based fluid systems

NASA Technical Reports Server (NTRS)

Elleman, Daniel D. (Inventor); Wang, Taylor G. (Inventor)

1989-01-01

Systems are described for using multiple closely-packed spheres. In one system for passing fluid, a multiplicity of spheres lie within a container, with all of the spheres having the same outside diameter and with the spheres being closely nested in one another to create multiple interstitial passages of a known size and configuration and smooth walls. The container has an inlet and outlet for passing fluid through the interstitial passages formed between the nested spheres. The small interstitial passages can be used to filter out material, especially biological material such as cells in a fluid, where the cells can be easily destroyed if passed across sharp edges. The outer surface of the spheres can contain a material that absorbs a constitutent in the flowing fluid, such as a particular contamination gas, or can contain a catalyst to chemically react the fluid passing therethrough, the use of multiple small spheres assuring a large area of contact of these surfaces of the spheres with the fluid. In a system for storing and releasing a fluid such as hydrogen as a fuel, the spheres can include a hollow shell containing the fluid to be stored, and located within a compressable container that can be compressed to break the shells and release the stored fluid.

Maturation State and Matrix Microstructure Regulate Interstitial Cell Migration in Dense Connective Tissues.

PubMed

Qu, Feini; Li, Qing; Wang, Xiao; Cao, Xuan; Zgonis, Miltiadis H; Esterhai, John L; Shenoy, Vivek B; Han, Lin; Mauck, Robert L

2018-02-19

Few regenerative approaches exist for the treatment of injuries to adult dense connective tissues. Compared to fetal tissues, adult connective tissues are hypocellular and show limited healing after injury. We hypothesized that robust repair can occur in fetal tissues with an immature extracellular matrix (ECM) that is conducive to cell migration, and that this process fails in adults due to the biophysical barriers imposed by the mature ECM. Using the knee meniscus as a platform, we evaluated the evolving micromechanics and microstructure of fetal and adult tissues, and interrogated the interstitial migratory capacity of adult meniscal cells through fetal and adult tissue microenvironments with or without partial enzymatic digestion. To integrate our findings, a computational model was implemented to determine how changing biophysical parameters impact cell migration through these dense networks. Our results show that the micromechanics and microstructure of the adult meniscus ECM sterically hinder cell mobility, and that modulation of these ECM attributes via an exogenous matrix-degrading enzyme permits migration through this otherwise impenetrable network. By addressing the inherent limitations to repair imposed by the mature ECM, these studies may define new clinical strategies to promote repair of damaged dense connective tissues in adults.

Fluorescence lifetime imaging of induced pluripotent stem cells

NASA Astrophysics Data System (ADS)

Uchugonova, Aisada; Batista, Ana; König, Karsten

2014-02-01

The multiphoton FLIM tomograph MPTflex with its flexible scan head, articulated arm, and the tunable femtosecond laser source was employed to study cell monolayers and 3D cell clusters. FLIM was performed with 250 ps temporal resolution and submicron special resolution using time-correlated single photon counting. The autofluorescence based on NAD(P)H and flavins/flavoproteins has been measured in mouse embryonic fibroblasts, induced pluripotent stem cells (iPS cells) originated from mouse embryonic fibroblasts and non-proliferative mouse embryonic fibroblasts.

Pre-crisis mouse cells show strain-specific covariation in the amount of 54-kilodalton phosphoprotein and in susceptibility to transformation by simian virus 40.

PubMed

Chen, S; Blanck, G; Pollack, R E

1983-09-01

We have used several inbred mouse strains to examine the role of the 54-kilodalton (kDa) cellular phosphoprotein in transformation by the papovavirus simian virus 40. We have measured the endogenous 54-kDa phosphoprotein in cells obtained from these inbred mouse strains. To study the effect of passage, cell cultures were measured for amount of the 54-kDa phosphoprotein at the 2nd and 12th passages. In the absence of any transforming agent, the amount of endogenous 54-kDa phosphoprotein in early pre-crisis mouse cells varied in a strain-specific way. Transformation frequency varied coordinately with endogenous 54-kDa expression. Mouse strains whose cells produced a high level of endogenous 54-kDa phosphoprotein on passage did not further increase its expression after simian virus 40 transformation.

Pre-crisis mouse cells show strain-specific covariation in the amount of 54-kilodalton phosphoprotein and in susceptibility to transformation by simian virus 40.

PubMed Central

Chen, S; Blanck, G; Pollack, R E

1983-01-01

We have used several inbred mouse strains to examine the role of the 54-kilodalton (kDa) cellular phosphoprotein in transformation by the papovavirus simian virus 40. We have measured the endogenous 54-kDa phosphoprotein in cells obtained from these inbred mouse strains. To study the effect of passage, cell cultures were measured for amount of the 54-kDa phosphoprotein at the 2nd and 12th passages. In the absence of any transforming agent, the amount of endogenous 54-kDa phosphoprotein in early pre-crisis mouse cells varied in a strain-specific way. Transformation frequency varied coordinately with endogenous 54-kDa expression. Mouse strains whose cells produced a high level of endogenous 54-kDa phosphoprotein on passage did not further increase its expression after simian virus 40 transformation. Images PMID:6310588

Differences between Mycobacterium-Host Cell Relationships in Latent Tuberculous Infection of Mice Ex Vivo and Mycobacterial Infection of Mouse Cells In Vitro

PubMed Central

Ufimtseva, Elena

2016-01-01

The search for factors that account for the reproduction and survival of mycobacteria, including vaccine strains, in host cells is the priority for studies on tuberculosis. A comparison of BCG-mycobacterial loads in granuloma cells obtained from bone marrow and spleens of mice with latent tuberculous infection and cells from mouse bone marrow and peritoneal macrophage cultures infected with the BCG vaccine in vitro has demonstrated that granuloma macrophages each normally contained a single BCG-Mycobacterium, while those acutely infected in vitro had increased mycobacterial loads and death rates. Mouse granuloma cells were observed to produce the IFNγ, IL-1α, GM-CSF, CD1d, CD25, CD31, СD35, and S100 proteins. None of these activation markers were found in mouse cell cultures infected in vitro or in intact macrophages. Lack of colocalization of lipoarabinomannan-labeled BCG-mycobacteria with the lysosomotropic LysoTracker dye in activated granuloma macrophages suggests that these macrophages were unable to destroy BCG-mycobacteria. However, activated mouse granuloma macrophages could control mycobacterial reproduction in cells both in vivo and in ex vivo culture. By contrast, a considerable increase in the number of BCG-mycobacteria was observed in mouse bone marrow and peritoneal macrophages after BCG infection in vitro, when no expression of the activation-related molecules was detected in these cells. PMID:27066505

The VP40 protein of Marburg virus exhibits impaired budding and increased sensitivity to human tetherin following mouse adaptation.

PubMed

Feagins, Alicia R; Basler, Christopher F

2014-12-01

The Marburg virus VP40 protein is a viral matrix protein that spontaneously buds from cells. It also functions as an interferon (IFN) signaling antagonist by targeting Janus kinase 1 (JAK1). A previous study demonstrated that the VP40 protein of the Ravn strain of Marburg virus (Ravn virus [RAVV]) failed to block IFN signaling in mouse cells, whereas the mouse-adapted RAVV (maRAVV) VP40 acquired the ability to inhibit IFN responses in mouse cells. The increased IFN antagonist function of maRAVV VP40 mapped to residues 57 and 165, which were mutated during the mouse adaptation process. In the present study, we demonstrate that maRAVV VP40 lost the capacity to efficiently bud from human cell lines, despite the fact that both parental and maRAVV VP40s bud efficiently from mouse cell lines. The impaired budding in human cells corresponds with the appearance of protrusions on the surface of maRAVV VP40-expressing Huh7 cells and with an increased sensitivity of maRAVV VP40 to restriction by human tetherin but not mouse tetherin. However, transfer of the human tetherin cytoplasmic tail to mouse tetherin restored restriction of maRAVV VP40. Residues 57 and 165 were demonstrated to contribute to the failure of maRAVV VP40 to bud from human cells, and residue 57 was demonstrated to alter VP40 oligomerization, as assessed by coprecipitation assay, and to determine sensitivity to human tetherin. This suggests that RAVV VP40 acquired, during adaptation to mice, changes in its oligomerization potential that enhanced IFN antagonist function. However, this new capacity impaired RAVV VP40 budding from human cells. Filoviruses, which include Marburg viruses and Ebola viruses, are zoonotic pathogens that cause severe disease in humans and nonhuman primates but do not cause similar disease in wild-type laboratory strains of mice unless first adapted to these animals. Although mouse adaptation has been used as a method to develop small animal models of pathogenesis, the molecular determinants associated with filovirus mouse adaptation are poorly understood. Our study demonstrates how genetic changes that accrued during mouse adaptation of the Ravn strain of Marburg virus have impacted the budding function of the viral VP40 matrix protein. Strikingly, we find impairment of mouse-adapted VP40 budding function in human but not mouse cell lines, and we correlate the impairment with an increased sensitivity of VP40 to restriction by human but not mouse tetherin and with changes in VP40 oligomerization. These data suggest that there are functional costs associated with filovirus adaptation to new hosts and implicate tetherin as a filovirus host restriction factor. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Spleen and Lymphatic System (For Parents)

MedlinePlus

... they have many tiny openings that allow gases, water, and nutrients to pass through to the surrounding cells, nourishing them and taking away waste products. When lymph fluid leaks through in this way it is called interstitial ...

Engineered three-dimensional microfluidic device for interrogating cell-cell interactions in the tumor microenvironment.

PubMed

Hockemeyer, K; Janetopoulos, C; Terekhov, A; Hofmeister, W; Vilgelm, A; Costa, Lino; Wikswo, J P; Richmond, A

2014-07-01

Stromal cells in the tumor microenvironment play a key role in the metastatic properties of a tumor. It is recognized that cancer-associated fibroblasts (CAFs) and endothelial cells secrete factors capable of influencing tumor cell migration into the blood or lymphatic vessels. We developed a microfluidic device that can be used to image the interactions between stromal cells and tumor cell spheroids in a three dimensional (3D) microenvironment while enabling external control of interstitial flow at an interface, which supports endothelial cells. The apparatus couples a 200-μm channel with a semicircular well to mimic the interface of a blood vessel with the stroma, and the design allows for visualization of the interactions of interstitial flow, endothelial cells, leukocytes, and fibroblasts with the tumor cells. We observed that normal tissue-associated fibroblasts (NAFs) contribute to the "single file" pattern of migration of tumor cells from the spheroid in the 3D microenvironment. In contrast, CAFs induce a rapid dispersion of tumor cells out of the spheroid with migration into the 3D matrix. Moreover, treatment of tumor spheroid cultures with the chemokine CXCL12 mimics the effect of the CAFs, resulting in similar patterns of dispersal of the tumor cells from the spheroid. Conversely, addition of CXCL12 to co-cultures of NAFs with tumor spheroids did not mimic the effects observed with CAF co-cultures, suggesting that NAFs produce factors that stabilize the tumor spheroids to reduce their migration in response to CXCL12.

Post-embryonic larval development and metamorphosis of the hydroid Eudendrium racemosum (Cavolini) (Hydrozoa, Cnidaria)

NASA Astrophysics Data System (ADS)

Sommer, C.

1990-09-01

The morphology and histology of the planula larva of Eudendrium racemosum (Cavolini) and its metamorphosis into the primary polyp are described from light microscopic observations. The planula hatches as a differentiated gastrula. During the lecithotrophic larval period, large ectodermal mucous cells, embedded between epitheliomuscular cells, secrete a sticky slime. Two granulated cell types occur in the ectoderm that are interpreted as secretory and sensorynervous cells, but might also be representatives of only one cell type with a multiple function. The entoderm consists of yolk-storing gastrodermal cells, digestive gland cells, interstitial cells, cnidoblasts, and premature cnidocytes. The larva starts metamorphosis by affixing its blunt aboral pole to a substratum. While the planula flattens down, the mucous cells penetrate the mesolamella and migrate through the entoderm into the gastral cavity where they are lysed. Subsequently, interstitial cells, cnidoblasts, and premature cnidocytes migrate in the opposite direction, i.e. from entoderm to ectoderm. Then, the polypoid body organization, comprising head (hydranth), stem and foot, all covered by peridermal secretion, becomes recognisable. An oral constriction divides the hypostomal portion of the gastral cavity from the stomachic portion. Within the hypostomal entoderm, cells containing secretory granules differentiate. Following growth and the multiplication of tentacles, the head periderm disappears. A ring of gland cells differentiates at the hydranth's base. The positioning of cnidae in the tentacle ectoderm, penetration of the mouth opening and the multiplication of digestive gland cells enable the polyp to change from lecithotrophic to planktotrophic nutrition.

Increased Expression of PcG Protein YY1 Negatively Regulates B Cell Development while Allowing Accumulation of Myeloid Cells and LT-HSC Cells

PubMed Central

Pan, Xuan; Jones, Morgan; Jiang, Jie; Zaprazna, Kristina; Yu, Duonan; Pear, Warren; Maillard, Ivan; Atchison, Michael L.

2012-01-01

Ying Yang 1 (YY1) is a multifunctional Polycomb Group (PcG) transcription factor that binds to multiple enhancer binding sites in the immunoglobulin (Ig) loci and plays vital roles in early B cell development. PcG proteins have important functions in hematopoietic stem cell renewal and YY1 is the only mammalian PcG protein with DNA binding specificity. Conditional knock-out of YY1 in the mouse B cell lineage results in arrest at the pro-B cell stage, and dosage effects have been observed at various YY1 expression levels. To investigate the impact of elevated YY1 expression on hematopoetic development, we utilized a mouse in vivo bone marrow reconstitution system. We found that mouse bone marrow cells expressing elevated levels of YY1 exhibited a selective disadvantage as they progressed from hematopoietic stem/progenitor cells to pro-B, pre-B, immature B and re-circulating B cell stages, but no disadvantage of YY1 over-expression was observed in myeloid lineage cells. Furthermore, mouse bone marrow cells expressing elevated levels of YY1 displayed enrichment for cells with surface markers characteristic of long-term hematopoietic stem cells (HSC). YY1 expression induced apoptosis in mouse B cell lines in vitro, and resulted in down-regulated expression of anti-apoptotic genes Bcl-xl and NFκB2, while no impact was observed in a mouse myeloid line. B cell apoptosis and LT-HSC enrichment induced by YY1 suggest that novel strategies to induce YY1 expression could have beneficial effects in the treatment of B lineage malignancies while preserving normal HSCs. PMID:22292011

Human Stem Cells Can Differentiate in Post-implantation Mouse Embryos.

PubMed

Tam, Patrick P L

2016-01-07

The potency of human pluripotent stem cells (hPSCs) to differentiate into germ layer derivatives is conventionally assessed by teratoma induction and in vitro differentiation. In this issue of Cell Stem Cell, Mascetti and Pedersen (2016) demonstrate that the human-mouse post-implantation chimera offers an efficient avenue to test the germ layer differentiation potential of hPSCs in mouse embryos ex vivo. Copyright © 2016 Elsevier Inc. All rights reserved.

Mouse Cytotoxic T Cell-derived Granzyme B Activates the Mitochondrial Cell Death Pathway in a Bim-dependent Fashion*

PubMed Central

Catalán, Elena; Jaime-Sánchez, Paula; Aguiló, Nacho; Simon, Markus M.; Froelich, Christopher J.; Pardo, Julián

2015-01-01

Cytotoxic T cells (Tc) use perforin and granzyme B (gzmB) to kill virus-infected cells and cancer cells. Recent evidence suggests that human gzmB primarily induces apoptosis via the intrinsic mitochondrial pathway by either cleaving Bid or activating Bim leading to the activation of Bak/Bax and subsequent generation of active caspase-3. In contrast, mouse gzmB is thought to predominantly induce apoptosis by directly processing pro-caspase-3. However, in certain mouse cell types gzmB-mediated apoptosis mainly occurs via the mitochondrial pathway. To investigate whether Bim is involved under the latter conditions, we have now employed ex vivo virus-immune mouse Tc that selectively kill by using perforin and gzmB (gzmB+Tc) as effector cells and wild type as well as Bim- or Bak/Bax-deficient spontaneously (3T9) or virus-(SV40) transformed mouse embryonic fibroblast cells as targets. We show that gzmB+Tc-mediated apoptosis (phosphatidylserine translocation, mitochondrial depolarization, cytochrome c release, and caspase-3 activation) was severely reduced in 3T9 cells lacking either Bim or both Bak and Bax. This outcome was related to the ability of Tc cells to induce the degradation of Mcl-1 and Bcl-XL, the anti-apoptotic counterparts of Bim. In contrast, gzmB+Tc-mediated apoptosis was not affected in SV40-transformed mouse embryonic fibroblast cells lacking Bak/Bax. The data provide evidence that Bim participates in mouse gzmB+Tc-mediated apoptosis of certain targets by activating the mitochondrial pathway and suggest that the mode of cell death depends on the target cell. Our results suggest that the various molecular events leading to transformation and/or immortalization of cells have an impact on their relative resistance to the multiple gzmB+Tc-induced death pathways. PMID:25605735

Role of Semaphorin 7a signaling in TGF-β1 induced lung fibrosis and scleroderma-related interstitial lung disease

PubMed Central

Gan, Ye; Reilkoff, Ronald; Peng, Xueyan; Russell, Thomas; Chen, Qingsheng; Mathai, Susan K.; Homer, Robert; Gulati, Mridu; Siner, Jonathan; Elias, Jack; Bucala, Richard; Herzog, Erica

2012-01-01

Objective Semaphorin (Sema) 7a regulates TGF- β1 induced fibrosis. Using a murine model of pulmonary fibrosis in which an inducible, bioactive form of the human TGF- β1 gene is overexpressed in the lung, we tested the hypothesis that Sema-7a exerts its pro-fibrotic effects in part by promoting the tissue accumulation of CD45+ fibrocytes. Methods Fibrosis and fibrocytes were evaluated in TGF- β1 transgenic mice in which the Sema-7a locus had been disrupted. The effect of replacement or deletion of Sema-7a on bone marrow derived cells was ascertained using bone marrow transplantation. The role of the Sema-7a receptor β1 integrin was assessed using neutralizing antibodies. The applicability of these findings to TGF-β1-driven fibrosis in humans was examined in patients with scleroderma-related interstitial lung disease. Results The appearance of fibrocytes in the lungs in TGF- β1 transgenic mice requires Sema-7a. Replacement of Sema-7a in bone marrow derived cells restores lung fibrosis and fibrocytes. Immunoneutralization of β1 integrin reduces pulmonary fibrocytes and fibrosis. Peripheral blood mononuclear cells from patients with scleroderma-related interstitial lung disease show increased mRNA for Sema-7a and the β1 integrin, with Sema-7a located on collagen producing fibrocytes and CD19+ lymphocytes. Peripheral blood fibrocyte outgrowth is enhanced in these patients. Stimulation of normal human peripheral blood mononuclear cells with recombinant Sema-7a enhances fibrocyte differentiation; these effects are attenuated by β1 integrin neutralization. Conclusion Interventions that reduce Sema-7a expression or prevent the Sema-7a - β1 integrin interaction may be ameliorative in TGF- β1-driven or fibrocyte-associated autoimmune fibroses. PMID:21484765

Cytokine expression in patients with bladder pain syndrome/interstitial cystitis ESSIC type 3C.

PubMed

Logadottir, Yr; Delbro, Dick; Fall, Magnus; Gjertsson, Inger; Jirholt, Pernilla; Lindholm, Catharina; Peeker, Ralph

2014-11-01

Bladder wall nitric oxide production in patients with bladder pain syndrome type 3C is increased compared to undetectable nitric oxide in patients with nonHunner bladder pain syndrome and healthy controls. However, the underlying mechanism/s of the increased nitric oxide production is largely unknown. We compared mRNA expression of a select group of cytokines in patients with bladder pain syndrome/interstitial cystitis type 3C and in pain-free controls. Cold cup biopsies from 7 patients with bladder pain syndrome type 3C and 6 healthy subjects were analyzed. mRNA expression of IL-4, 6, 10 and 17A, iNOS, TNF-α, TGF-β and IFN-γ was estimated by real-time polymerase chain reaction. IL-17 protein expression was determined by immunohistochemistry. Mast cells were labeled with tryptase to evaluate cell appearance and count. IL-6, 10 and 17A, and iNOS mRNA levels as well as the number of mast cells infiltrating the bladder mucosa were significantly increased in patients with bladder pain syndrome type 3C compared to healthy controls. TNF-α, TGF-β and IFN-γ mRNA levels were similar in patients and controls. IL-17A expression at the protein level was up-regulated and localized to inflammatory cells and urothelium in patients with bladder pain syndrome type 3C. Patients with bladder pain syndrome/interstitial cystitis had increased mRNA levels of IL-17A, 10 and 6, and iNOS. IL-17A might be important in the inflammatory process. To our knowledge the increase in IL-17A is a novel finding that may have new treatment implications. Copyright © 2014 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

Inflammation and Tissue Remodeling in the Bladder and Urethra in Feline Interstitial Cystitis

PubMed Central

Kullmann, F. Aura; McDonnell, Bronagh M.; Wolf-Johnston, Amanda S.; Lynn, Andrew M.; Getchell, Samuel E.; Ruiz, Wily G.; Zabbarova, Irina V.; Ikeda, Youko; Kanai, Anthony J.; Roppolo, James R.; Bastacky, Sheldon I.; Apodaca, Gerard; Buffington, C. A. Tony; Birder, Lori A.

2018-01-01

Interstitial cystitis/bladder pain syndrome (IC/BPS) is a debilitating chronic disease of unknown etiology. A naturally occurring disease termed feline interstitial cystitis (FIC) reproduces many features of IC/BPS patients. To gain insights into mechanisms underlying IC/BPS, we investigated pathological changes in the lamina propria (LP) of the bladder and proximal urethra in cats with FIC, using histological and molecular methods. Compared to control cat tissue, we found an increased number of de-granulated mast cells, accumulation of leukocytes, increased cyclooxygenase (COX)-1 expression in the bladder LP, and increased COX-2 expression in the urethra LP from cats with FIC. We also found increased suburothelial proliferation, evidenced by mucosal von Brunn’s nests, neovascularization and alterations in elastin content. Scanning electron microscopy revealed normal appearance of the superficial urethral epithelium, including the neuroendocrine cells (termed paraneurons), in FIC urethrae. Together, these histological findings suggest the presence of chronic inflammation of unknown origin leading to tissue remodeling. Since the mucosa functions as part of a “sensory network” and urothelial cells, nerves and other cells in the LP are influenced by the composition of the underlying tissues including the vasculature, the changes observed in the present study may alter the communication of sensory information between different cellular components. This type of mucosal signaling can also extend to the urethra, where recent evidence has revealed that the urethral epithelium is likely to be part of a signaling system involving paraneurons and sensory nerves. Taken together, our data suggest a more prominent role for chronic inflammation and tissue remodeling than previously thought, which may result in alterations in mucosal signaling within the urinary bladder and proximal urethra that may contribute to altered sensations and pain in cats and humans with this syndrome. PMID:29706873

Interstitial fluid flow and drug delivery in vascularized tumors: a computational model.

PubMed

Welter, Michael; Rieger, Heiko

2013-01-01

Interstitial fluid is a solution that bathes and surrounds the human cells and provides them with nutrients and a way of waste removal. It is generally believed that elevated tumor interstitial fluid pressure (IFP) is partly responsible for the poor penetration and distribution of therapeutic agents in solid tumors, but the complex interplay of extravasation, permeabilities, vascular heterogeneities and diffusive and convective drug transport remains poorly understood. Here we consider-with the help of a theoretical model-the tumor IFP, interstitial fluid flow (IFF) and its impact upon drug delivery within tumor depending on biophysical determinants such as vessel network morphology, permeabilities and diffusive vs. convective transport. We developed a vascular tumor growth model, including vessel co-option, regression, and angiogenesis, that we extend here by the interstitium (represented by a porous medium obeying Darcy's law) and sources (vessels) and sinks (lymphatics) for IFF. With it we compute the spatial variation of the IFP and IFF and determine its correlation with the vascular network morphology and physiological parameters like vessel wall permeability, tissue conductivity, distribution of lymphatics etc. We find that an increased vascular wall conductivity together with a reduction of lymph function leads to increased tumor IFP, but also that the latter does not necessarily imply a decreased extravasation rate: Generally the IF flow rate is positively correlated with the various conductivities in the system. The IFF field is then used to determine the drug distribution after an injection via a convection diffusion reaction equation for intra- and extracellular concentrations with parameters guided by experimental data for the drug Doxorubicin. We observe that the interplay of convective and diffusive drug transport can lead to quite unexpected effects in the presence of a heterogeneous, compartmentalized vasculature. Finally we discuss various strategies to increase drug exposure time of tumor cells.

Lentivirus-mediated bifunctional cell labeling for in vivo melanoma study

PubMed Central

Day, Chi-Ping; Carter, John; Bonomi, Carrie; Esposito, Dominic; Crise, Bruce; Ortiz-Conde, Betty; Hollingshead, Melinda; Merlino, Glenn

2009-01-01

SUMMARY Lentiviral vectors (LVs) are capable of labeling a broad spectrum of cell types, achieving stable expression of transgenes. However, for in vivo studies, the duration of marker gene expression has been highly variable. We have developed a series of LVs harboring different promoters for expressing reporter gene in mouse cells. Long-term culture and colony formation of several LV-labeled mouse melanoma cells showed that promoters derived from mammalian house-keeping genes, especially those encoding RNA polymerase II (Pol2) and ferritin (FerH), provided the highest consistency for reporter expression. For in vivo studies, primary B16BL6 mouse melanoma were infected with LVs whose luciferase-GFP fusion gene (Luc/GFP) was driven by either Pol2 or FerH promoters. When transplanted into syngeneic C57BL/6 mice, Luc/GFP-labeled B16BL6 mouse melanoma cells can be monitored by bioluminescence imaging in vivo, and GFP-positive cells can be isolated from the tumors by FACS. Pol2-Luc/GFP labeling, while lower in activity, was more sustainable than FerH-Luc/GFP labeling in B16BL6 over consecutive passages into mice. We conclude that Pol-2-Luc/GFP labeling allows long-term in vivo monitoring and tumor cell isolation in immunocompetent mouse melanoma models. SIGNIFICANCE In this study we have developed and identified lentiviral vectors that allow labeled mouse melanoma cells to maintain long-term and consistent expression of a bifunctional luciferase-GFP marker gene, even in syngeneic mice with an intact immune function. This cell-labeling system can be used to build immunocompetent mouse melanoma models that permit both tumor monitoring and FACS-based tumor cell isolation from tissues, greatly facilitating the in vivo study of melanoma. PMID:19175523

Cloning of ES cells and mice by nuclear transfer.

PubMed

Wakayama, Sayaka; Kishigami, Satoshi; Wakayama, Teruhiko

2009-01-01

We have been able to develop a stable nuclear transfer (NT) method in the mouse, in which donor nuclei are directly injected into the oocyte using a piezo-actuated micromanipulator. Although the piezo unit is a complex tool, once mastered it is of great help not only in NT experiments, but also in almost all other forms of micromanipulation. Using this technique, embryonic stem (ntES) cell lines established from somatic cell nuclei can be generated relatively easily from a variety of mouse genotypes and cell types. Such ntES cells can be used not only for experimental models of human therapeutic cloning but also as a means of preserving mouse genomes instead of preserving germ cells. Here, we describe our most recent protocols for mouse cloning.

Genetic susceptibility to interstitial pulmonary fibrosis in mice induced by vanadium pentoxide (V2O5)

PubMed Central

Walters, Dianne M.; White, Kevin M.; Patel, Ushma; Davis, Martin J.; Veluci-Marlow, Roberta M.; Bhupanapadu Sunkesula, Solomon Raju; Bonner, James C.; Martin, Jessica R.; Gladwell, Wes; Kleeberger, Steven R.

2014-01-01

Interstitial lung diseases (ILDs) are characterized by injury, inflammation, and scarring of alveoli, leading to impaired function. The etiology of idiopathic forms of ILD is not understood, making them particularly difficult to study due to the lack of appropriate animal models. Consequently, few effective therapies have emerged. We developed an inbred mouse model of ILD using vanadium pentoxide (V2O5), the most common form of a transition metal found in cigarette smoke, fuel ash, mineral ores, and steel alloys. Pulmonary responses to V2O5, including dose-dependent increases in lung permeability, inflammation, collagen content, and dysfunction, were significantly greater in DBA/2J mice compared to C57BL/6J mice. Inflammatory and fibrotic responses persisted for 4 mo in DBA/2J mice, while limited responses in C57BL/6J mice resolved. We investigated the genetic basis for differential responses through genetic mapping of V2O5-induced lung collagen content in BXD recombinant inbred (RI) strains and identified significant linkage on chromosome 4 with candidate genes that associate with V2O5-induced collagen content across the RI strains. Results suggest that V2O5 may induce pulmonary fibrosis through mechanisms distinct from those in other models of pulmonary fibrosis. These findings should further advance our understanding of mechanisms involved in ILD and thereby aid in identification of new therapeutic targets.—Walters, D. M., White, K. M., Patel, U., Davis, M. J., Veluci-Marlow, R. M., Bhupanapadu Sunkesula, S. R., Bonner, J. C., Martin, J. R., Gladwell, W., Kleeberger, S. R. Genetic susceptibility to interstitial pulmonary fibrosis in mice induced by vanadium pentoxide (V2O5). PMID:24285090

Dynamics of circulating gamma delta T cell activity in an immunocompetent mouse model of high-grade glioma

USDA-ARS?s Scientific Manuscript database

Human gamma delta T cells are potent effectors against glioma cell lines in vitro and in human/mouse xenograft models of glioblastoma, however, this effect has not been investigated in an immunocompetent mouse model. In this report, we established GL261 intracranial gliomas in syngeneic WT C57BL/6 m...

The Protective Effect of Baicalin against UVB Irradiation Induced Photoaging: An In Vitro and In Vivo Study

PubMed Central

Yin, Zhi-qiang; Hu, Yan-yan; Xu, Yang; Wu, Di; Permatasari, Felicia; Luo, Dan; Zhou, Bing-rong

2014-01-01

Objective This study was aimed to evaluate the anti-photoaging effects of baicalin on Ultraviolet B (UVB)-induced photoaging in the dorsal skin of hairless mice and premature senescence in human dermal fibroblasts. Methods We established in vivo and in vitro photoaging models by repeated exposures to UVB irradiation. By HE staining, masson staining, immunohistostaing and real-time RT-PCR, we analyzed epidermal thickness, collagen expression and the mRNA and protein levels of type I collagen, type III collagen, interstitial collagenase (MMP-1 and MMP-3) in UVB exposed dorsal mice skin. The aging condition in human dermal fibroblasts was determined by senescence-associated β-galactosidase (SA-β-gal) staining. Cell viability was determined using the Cell Counting Kit-8 (CCK-8). The G1 phase cell growth arrest was analyzed by flow cytometry. The senescence-related protein levels of p16INK-4a, p21WAF-1, and p53 and protein levels of phosphorylated histone H2AX were estimated by Western blotting. Results Topically application of baicalin treatment reduced UVB-induced epidermal thickening of mouse skin and also result in an increase in the production of collagen I and III, and a decrease in the expression of MMP-1 and MMP-3. Compared with the UVB-irradiated group, we found that the irradiated fibroblasts additionally treated with baicalin demonstrated a decrease in the expression of SA-β-gal, a increase in the cell viability, a decrease in the G1 phase cell proportion, a downregulation in the level of senescence-associated and γ-H2AX proteins. However, Baicalin had no difference in the normal fibroblasts without UVB irradiation and long-term Baicalin incubation of UVB-SIPS fibroblasts gave no effects on the cell proliferation. Conclusions Taken together, these results suggest that baicalin significantly antagonizes photoaging induced by UVB in vivo and in vitro, indicating the potential of baicalin application for anti-photoaging treatment. PMID:24949843

The protective effect of baicalin against UVB irradiation induced photoaging: an in vitro and in vivo study.

PubMed

Zhang, Jia-an; Yin, Zhi; Ma, Li-wen; Yin, Zhi-qiang; Hu, Yan-yan; Xu, Yang; Wu, Di; Permatasari, Felicia; Luo, Dan; Zhou, Bing-rong

2014-01-01

This study was aimed to evaluate the anti-photoaging effects of baicalin on Ultraviolet B (UVB)-induced photoaging in the dorsal skin of hairless mice and premature senescence in human dermal fibroblasts. We established in vivo and in vitro photoaging models by repeated exposures to UVB irradiation. By HE staining, masson staining, immunohistostaing and real-time RT-PCR, we analyzed epidermal thickness, collagen expression and the mRNA and protein levels of type I collagen, type III collagen, interstitial collagenase (MMP-1 and MMP-3) in UVB exposed dorsal mice skin. The aging condition in human dermal fibroblasts was determined by senescence-associated β-galactosidase (SA-β-gal) staining. Cell viability was determined using the Cell Counting Kit-8 (CCK-8). The G1 phase cell growth arrest was analyzed by flow cytometry. The senescence-related protein levels of p16INK-4a, p21WAF-1, and p53 and protein levels of phosphorylated histone H2AX were estimated by Western blotting. Topically application of baicalin treatment reduced UVB-induced epidermal thickening of mouse skin and also result in an increase in the production of collagen I and III, and a decrease in the expression of MMP-1 and MMP-3. Compared with the UVB-irradiated group, we found that the irradiated fibroblasts additionally treated with baicalin demonstrated a decrease in the expression of SA-β-gal, a increase in the cell viability, a decrease in the G1 phase cell proportion, a downregulation in the level of senescence-associated and γ-H2AX proteins. However, Baicalin had no difference in the normal fibroblasts without UVB irradiation and long-term Baicalin incubation of UVB-SIPS fibroblasts gave no effects on the cell proliferation. Taken together, these results suggest that baicalin significantly antagonizes photoaging induced by UVB in vivo and in vitro, indicating the potential of baicalin application for anti-photoaging treatment.

In vivo visualization and ex vivo quantification of murine breast cancer cells in the mouse brain using MRI cell tracking and electron paramagnetic resonance.

PubMed

Danhier, Pierre; Magat, Julie; Levêque, Philippe; De Preter, Géraldine; Porporato, Paolo E; Bouzin, Caroline; Jordan, Bénédicte F; Demeur, Gladys; Haufroid, Vincent; Feron, Olivier; Sonveaux, Pierre; Gallez, Bernard

2015-03-01

Cell tracking could be useful to elucidate fundamental processes of cancer biology such as metastasis. The aim of this study was to visualize, using MRI, and to quantify, using electron paramagnetic resonance (EPR), the entrapment of murine breast cancer cells labeled with superparamagnetic iron oxide particles (SPIOs) in the mouse brain after intracardiac injection. For this purpose, luciferase-expressing murine 4 T1-luc breast cancer cells were labeled with fluorescent Molday ION Rhodamine B SPIOs. Following intracardiac injection, SPIO-labeled 4 T1-luc cells were imaged using multiple gradient-echo sequences. Ex vivo iron oxide quantification in the mouse brain was performed using EPR (9 GHz). The long-term fate of 4 T1-luc cells after injection was characterized using bioluminescence imaging (BLI), brain MRI and immunofluorescence. We observed hypointense spots due to SPIO-labeled cells in the mouse brain 4 h after injection on T2 *-weighted images. Histology studies showed that SPIO-labeled cancer cells were localized within blood vessels shortly after delivery. Ex vivo quantification of SPIOs showed that less than 1% of the injected cells were taken up by the mouse brain after injection. MRI experiments did not reveal the development of macrometastases in the mouse brain several days after injection, but immunofluorescence studies demonstrated that these cells found in the brain established micrometastases. Concerning the metastatic patterns of 4 T1-luc cells, an EPR biodistribution study demonstrated that SPIO-labeled 4 T1-luc cells were also entrapped in the lungs of mice after intracardiac injection. BLI performed 6 days after injection of 4 T1-luc cells showed that this cell line formed macrometastases in the lungs and in the bones. Conclusively, EPR and MRI were found to be complementary for cell tracking applications. MRI cell tracking at 11.7 T allowed sensitive detection of isolated SPIO-labeled cells in the mouse brain, whereas EPR allowed the assessment of the number of SPIO-labeled cells in organs shortly after injection. Copyright © 2015 John Wiley & Sons, Ltd.

Derivation of novel human ground state naive pluripotent stem cells.

PubMed

Gafni, Ohad; Weinberger, Leehee; Mansour, Abed AlFatah; Manor, Yair S; Chomsky, Elad; Ben-Yosef, Dalit; Kalma, Yael; Viukov, Sergey; Maza, Itay; Zviran, Asaf; Rais, Yoach; Shipony, Zohar; Mukamel, Zohar; Krupalnik, Vladislav; Zerbib, Mirie; Geula, Shay; Caspi, Inbal; Schneir, Dan; Shwartz, Tamar; Gilad, Shlomit; Amann-Zalcenstein, Daniela; Benjamin, Sima; Amit, Ido; Tanay, Amos; Massarwa, Rada; Novershtern, Noa; Hanna, Jacob H

2013-12-12

Mouse embryonic stem (ES) cells are isolated from the inner cell mass of blastocysts, and can be preserved in vitro in a naive inner-cell-mass-like configuration by providing exogenous stimulation with leukaemia inhibitory factor (LIF) and small molecule inhibition of ERK1/ERK2 and GSK3β signalling (termed 2i/LIF conditions). Hallmarks of naive pluripotency include driving Oct4 (also known as Pou5f1) transcription by its distal enhancer, retaining a pre-inactivation X chromosome state, and global reduction in DNA methylation and in H3K27me3 repressive chromatin mark deposition on developmental regulatory gene promoters. Upon withdrawal of 2i/LIF, naive mouse ES cells can drift towards a primed pluripotent state resembling that of the post-implantation epiblast. Although human ES cells share several molecular features with naive mouse ES cells, they also share a variety of epigenetic properties with primed murine epiblast stem cells (EpiSCs). These include predominant use of the proximal enhancer element to maintain OCT4 expression, pronounced tendency for X chromosome inactivation in most female human ES cells, increase in DNA methylation and prominent deposition of H3K27me3 and bivalent domain acquisition on lineage regulatory genes. The feasibility of establishing human ground state naive pluripotency in vitro with equivalent molecular and functional features to those characterized in mouse ES cells remains to be defined. Here we establish defined conditions that facilitate the derivation of genetically unmodified human naive pluripotent stem cells from already established primed human ES cells, from somatic cells through induced pluripotent stem (iPS) cell reprogramming or directly from blastocysts. The novel naive pluripotent cells validated herein retain molecular characteristics and functional properties that are highly similar to mouse naive ES cells, and distinct from conventional primed human pluripotent cells. This includes competence in the generation of cross-species chimaeric mouse embryos that underwent organogenesis following microinjection of human naive iPS cells into mouse morulas. Collectively, our findings establish new avenues for regenerative medicine, patient-specific iPS cell disease modelling and the study of early human development in vitro and in vivo.

Anti-huCD20 Antibody Therapy for Antibody-Mediated Rejection of Renal Allografts in a Mouse Model

PubMed Central

Abe, Toyofumi; Ishii, Daisuke; Gorbacheva, Victoria; Kohei, Naoki; Tsuda, Hidetoshi; Tanaka, Toshiaki; Dvorina, Nina; Nonomura, Norio; Takahara, Shiro; Valujskikh, Anna; Baldwin, William M.; Fairchild, Robert L.

2016-01-01

We have reported that B6.CCR5−/− mice reject renal allografts with high serum donor-specific antibody (DSA) titers and marked C4d deposition in grafts, features consistent with AMR. B6.huCD20/CCR5−/− mice, where human CD20 expression is restricted to B cells, rejected A/J renal allografts by day 26 post-transplant with DSA first detected in serum on day 5 post-transplant and increased thereafter. Recipient treatment with anti-huCD20 mAb prior to the transplant and weekly up to 7 weeks post-transplant promoted long-term allograft survival (> 100 days) with low DSA titers. To investigate the effect of B cell depletion at the time serum DSA was first detected, recipients were treated with anti-huCD20 mAb on days 5, 8 and 12 post-transplant. This regimen significantly reduced DSA titers and graft inflammation on day 15 post-transplant and prolonged allograft survival > 60 days. However, DSA returned to the titers observed in control treated recipients by day 30 post-transplant and histological analyses on day 60 post-transplant indicated severe interstitial fibrosis. These results indicate that anti-huCD20 mAb had the greatest effect as a prophylactic treatment and that the distinct kinetics of DSA responses accounts for acute renal allograft failure versus the development of fibrosis. PMID:25731734

EGF receptor tyrosine kinase inhibitors diminish transforming growth factor-alpha-induced pulmonary fibrosis.

PubMed

Hardie, William D; Davidson, Cynthia; Ikegami, Machiko; Leikauf, George D; Le Cras, Timothy D; Prestridge, Adrienne; Whitsett, Jeffrey A; Korfhagen, Thomas R

2008-06-01

Transforming growth factor-alpha (TGF-alpha) is a ligand for the EGF receptor (EGFR). EGFR activation is associated with fibroproliferative processes in human lung disease and animal models of pulmonary fibrosis. We determined the effects of EGFR tyrosine kinase inhibitors gefitinib (Iressa) and erlotinib (Tarceva) on the development and progression of TGF-alpha-induced pulmonary fibrosis. Using a doxycycline-regulatable transgenic mouse model of lung-specific TGF-alpha expression, we determined effects of treatment with gefitinib and erlotinib on changes in lung histology, total lung collagen, pulmonary mechanics, pulmonary hypertension, and expression of genes associated with synthesis of ECM and vascular remodeling. Induction in the lung of TGF-alpha caused progressive pulmonary fibrosis over an 8-wk period. Daily administration of gefitinib or erlotinib prevented development of fibrosis, reduced accumulation of total lung collagen, prevented weight loss, and prevented changes in pulmonary mechanics. Treatment of mice with gefitinib 4 wk after the induction of TGF-alpha prevented further increases in and partially reversed total collagen levels and changes in pulmonary mechanics and pulmonary hypertension. Increases in expression of genes associated with synthesis of ECM as well as decreases of genes associated with vascular remodeling were also prevented or partially reversed. Administration of gefitinib or erlotinib did not cause interstitial fibrosis or increases in lavage cell counts. Administration of small molecule EGFR tyrosine kinase inhibitors prevented further increases in and partially reversed pulmonary fibrosis induced directly by EGFR activation without inducing inflammatory cell influx or additional lung injury.

XRCC4 suppresses medulloblastomas with recurrent translocations in p53-deficient mice

PubMed Central

Yan, Catherine T.; Kaushal, Dhruv; Murphy, Michael; Zhang, Yu; Datta, Abhishek; Chen, Changzhong; Monroe, Brianna; Mostoslavsky, Gustavo; Coakley, Kristen; Gao, Yijie; Mills, Kevin D.; Fazeli, Alex P.; Tepsuporn, Suprawee; Hall, Giles; Mulligan, Richard; Fox, Edward; Bronson, Roderick; De Girolami, Umberto; Lee, Charles; Alt, Frederick W.

2006-01-01

Inactivation of the XRCC4 nonhomologous end-joining factor in the mouse germ line leads to embryonic lethality, in association with apoptosis of newly generated, postmitotic neurons. We now show that conditional inactivation of the XRCC4 in nestin-expressing neuronal progenitor cells, although leading to no obvious phenotype in a WT background, leads to early onset of neuronally differentiated medulloblastomas (MBs) in a p53-deficient background. A substantial proportion of the XRCC4/p53-deficient MBs have high-level N-myc gene amplification, often intrachromosomally in the context of complex translocations or other alterations of chromosome 12, on which N-myc resides, or extrachromosomally within double minutes. In addition, most XRCC4/p53-deficient MBs harbor clonal translocations of chromosome 13, which frequently involve chromosome 6 as a partner. One copy of the patched gene (Ptc), which lies on chromosome 13, was deleted in all tested XRCC4/p53-deficient MBs in the context of translocations or interstitial deletions. In addition, Cyclin D2, a chromosome 6 gene, was amplified in a subset of tumors. Notably, amplification of Myc-family or Cyclin D2 genes and deletion of Ptc also have been observed in human MBs. We therefore conclude that, in neuronal cells of mice, the nonhomologous end-joining pathway plays a critical role in suppressing genomic instability that, in a p53-deficient background, routinely contributes to genesis of MBs with recurrent chromosomal alterations. PMID:16670198

TGF-β induces surface LAP expression on murine CD4 T cells independent of Foxp3 induction.

PubMed

Oida, Takatoku; Weiner, Howard L

2010-11-24

It has been reported that human FOXP3(+) CD4 Tregs express GARP-anchored surface latency-associated peptide (LAP) after activation, based on the use of an anti-human LAP mAb. Murine CD4 Foxp3(+) Tregs have also been reported to express surface LAP, but these studies have been hampered by the lack of suitable anti-mouse LAP mAbs. We generated anti-mouse LAP mAbs by immunizing TGF-β(-/-) animals with a mouse Tgfb1-transduced P3U1 cell line. Using these antibodies, we demonstrated that murine Foxp3(+) CD4 Tregs express LAP on their surface. In addition, retroviral transduction of Foxp3 into mouse CD4(+)CD25(-) T cells induced surface LAP expression. We then examined surface LAP expression after treating CD4(+)CD25(-) T cells with TGF-β and found that TGF-β induced surface LAP not only on T cells that became Foxp3(+) but also on T cells that remained Foxp3(-) after TGF-β treatment. GARP expression correlated with the surface LAP expression, suggesting that surface LAP is GARP-anchored also in murine T cells. Unlike human CD4 T cells, surface LAP expression on mouse CD4 T cells is controlled by Foxp3 and TGF-β. Our newly described anti-mouse LAP mAbs will provide a useful tool for the investigation and functional analysis of T cells that express LAP on their surface.

Comparison of exendin-4 on beta-cell replication in mouse and human islet grafts.

PubMed

Tian, Lei; Gao, Jie; Weng, Guangbin; Yi, Huimin; Tian, Bole; O'Brien, Timothy D; Guo, Zhiguang

2011-08-01

Exendin-4 can stimulate β-cell replication in mice. Whether it can stimulate β-cell replication in human islet grafts remains unknown. Therefore, we compared the effects of exendin-4 on β-cell replication in mouse and human islet grafts. Islets, isolated from mouse and human donors at different ages, were transplanted into diabetic mice and/or diabetic nude mice that were given bromodeoxyuridine (BrdU) with or without exendin-4. At 4 weeks post-transplantation, islet grafts were removed for insulin and BrdU staining and quantification of insulin(+)/BrdU(+) cells. Although diabetes was reversed in all mice transplanting syngeneic mouse islets from young or old donors, normoglycemia was achieved significantly faster in exendin-4 treated mice. Mouse islet grafts in exendin-4 treated mice had significantly more insulin(+)/BrdU(+) β cells than in untreated mice (P < 0.01). Human islet grafts from ≤22-year-old donors had more insulin(+)/BrdU(+) β cells in exendin-4 treated mice than that in untreated mice (P < 0.01). However, human islet grafts from ≥35-year-old donors contained few insulin(+)/BrdU(+) β cells in exendin-4 treated or untreated mice. Our data demonstrated that the capacity for β-cell replication in mouse and human islet grafts is different with and without exendin-4 treatment and indicated that GLP-1 agonists can stimulate β-cell replication in human islets from young donors. © 2011 The Authors. Transplant International © 2011 European Society for Organ Transplantation.

Comparative stereology of the mouse and finch left ventricle.

PubMed

Bossen, E H; Sommer, J R; Waugh, R A

1978-01-01

The volume fractions and surface per unit cell volume of some subcellular components of the left ventricles of the finch and mouse were quantitated by stereologic techniques. These species were chosen for study because they have similar heart rates but differ morphologically in some respects: fiber diameter is larger in the mouse; the mouse has transverse tubules while the finch does not; and the finch has a form of junctional sarcoplasmic reticulum (JSR), extended JSR (EJSR), located in the cell interior with no direct plasmalemmal contact, while the mouse interior JSR (IJSR) abuts on transverse tubules. Our data show that the volume fraction (Vv) and surface area per unit cell volume (Sv) of total SR, and free SR (FSR) are similar. The volume fractions of mitochondria, myofibrils, and total junctional SR were also similar. The Sv of the cell surface of the finch was similar to the Sv of the cell surface of the mouse (Sv-plasmalemma plus Sv of the transverse tubules). The principal difference was in the distribution of JSR; the mouse peripheral JSR (PJSR) represents only 9% of the total JSR, while the finch PJSR accounts for 24% of the bird's JSR. The similar volume fractions of total junctional SR (PJSR + EJSR in the finch; PJSR + IJSR in the mouse) suggest that the EJSR is not an embryologic remnant, and raises the possibility that some function of JSR is independent of plasmalemmal contact.

Primary Tumor and MEF Cell Isolation to Study Lung Metastasis.

PubMed

Dong, Shengli; Maziveyi, Mazvita; Alahari, Suresh K

2015-05-20

In breast tumorigenesis, the metastatic stage of the disease poses the greatest threat to the affected individual. Normal breast cells with altered genotypes now possess the ability to invade and survive in other tissues. In this protocol, mouse mammary tumors are removed and primary cells are prepared from tumors. The cells isolated from this procedure are then available for gene profiling experiments. For successful metastasis, these cells must be able to intravasate, survive in circulation, extravasate to distant organs, and survive in that new organ system. The lungs are the typical target of breast cancer metastasis. A set of genes have been discovered that mediates the selectivity of metastasis to the lung. Here we describe a method of studying lung metastasis from a genetically engineered mouse model.. Furthermore, another protocol for analyzing mouse embryonic fibroblasts (MEFs) from the mouse embryo is included. MEF cells from the same animal type provide a clue of non-cancer cell gene expression. Together, these techniques are useful in studying mouse mammary tumorigenesis, its associated signaling mechanisms and pathways of the abnormalities in embryos.

Quantitative Analyses of Synergistic Responses between Cannabidiol and DNA-Damaging Agents on the Proliferation and Viability of Glioblastoma and Neural Progenitor Cells in Culture

PubMed Central

Deng, Liting; Ng, Lindsay; Ozawa, Tatsuya

2017-01-01

Evidence suggests that the nonpsychotropic cannabis-derived compound, cannabidiol (CBD), has antineoplastic activity in multiple types of cancers, including glioblastoma multiforme (GBM). DNA-damaging agents remain the main standard of care treatment available for patients diagnosed with GBM. Here we studied the antiproliferative and cell-killing activity of CBD alone and in combination with DNA-damaging agents (temozolomide, carmustine, or cisplatin) in several human GBM cell lines and in mouse primary GBM cells in cultures. This activity was also studied in mouse neural progenitor cells (NPCs) in culture to assess for potential central nervous system toxicity. We found that CBD induced a dose-dependent reduction of both proliferation and viability of all cells with similar potencies, suggesting no preferential activity for cancer cells. Hill plot analysis indicates an allosteric mechanism of action triggered by CBD in all cells. Cotreatment regimens combining CBD and DNA-damaging agents produced synergistic antiproliferating and cell-killing responses over a limited range of concentrations in all human GBM cell lines and mouse GBM cells as well as in mouse NPCs. Remarkably, antagonistic responses occurred at low concentrations in select human GBM cell lines and in mouse GBM cells. Our study suggests limited synergistic activity when combining CBD and DNA-damaging agents in treating GBM cells, along with little to no therapeutic window when considering NPCs. PMID:27821713

Ligand activation of peroxisome proliferator-activated receptor-β/δ (PPAR β/δ) inhibits cell growth in a mouse mammary gland cancer cell line

PubMed Central

Foreman, Jennifer E.; Sharma, Arun K.; Amin, Shantu; Gonzalez, Frank J.; Peters, Jeffrey M.

2009-01-01

The effects of ligand activation of PPARβ/δ were examined in the mouse mammary tumor cell line (C20). Expression of PPARβ/δ was markedly lower in C20 cells as compared to the human non-tumorigenic mammary gland derived cell line (MCF10A) and mouse keratinocytes. Ligand activation of PPARβ/δ in C20 cells caused upregulation of the PPARβ/δ target gene angiopoietin-like 4 (Angptl4). Inhibition of C20 cell proliferation and clonogenicity was observed following treatment with GW0742 or GW501516, two highly specific PPARβ/δ ligands. In addition, an increase in apoptosis was observed in C20 cells cultured with 10 µM GW501516 that preceded the observed inhibition of cell proliferation. Results from this study show that proliferation of the C20 mouse mammary gland cancer cell line is inhibited by ligand activation of PPARβ/δ due in part to increased apoptosis. PMID:19660859

Selective Neurodegeneration, Without Neurofibrillary Tangles, in a Mouse Model of Niemann-Pick C Disease

PubMed Central

German, Dwight C.; Quintero, E. Matthew; Liang, Chang-Lin; Ng, Benton; Punia, Surender; Xie, Chonglun; Dietschy, John M.

2012-01-01

The BALB/c mouse model of Niemann-Pick type C (NPC) disease exhibits neuropathological similarities to the human condition. There is an age-related cerebral atrophy, demyelination of the corpus callosum, and degeneration of cerebellar Purkinje cells in the NPC mouse. In human NPC, many cortical and subcortical neurons contain neurofibrillary tangles, which are thought by some investigators to play an important role in the neurodegenerative process. The purpose of the present study was to determine whether neurodegeneration occurs in the NPC mouse, in brain regions other than the cerebellum and whether the degeneration is related to the presence of neurofibrillary tangles. Using light microscopic methods with immunohistochemistry, electron microscopy, and cell counting methods, 11-week-old NPC+/+ and NPC−/− animals were examined. In the NPC−/− mice, there were 96% fewer Purkinje cells, 28% fewer neurons in the prefrontal cortex, 20% fewer neurons in the thalamus, and 63% fewer glial cells in the corpus callosum. On the other hand, previous studies indicate normal numbers of neurons and glial cells in these same neuroanatomical regions in young NPC−/− mice. There were normal numbers of cholinergic neurons in sections assessed in the striatum and basal forebrain in the 11-week-old animals and no evidence of neurofibrillary tangles within cells. The present data indicate that both neurons and glial cells die in the NPC mouse but that all cells are not equally vulnerable. There was no evidence for neurofibrillary tangles in the NPC mouse, and therefore the degenerative process in the mouse is unrelated to the neurofibrillary tangle. PMID:11298365

Mast cells in airway diseases and interstitial lung disease.

PubMed

Cruse, Glenn; Bradding, Peter

2016-05-05

Mast cells are major effector cells of inflammation and there is strong evidence that mast cells play a significant role in asthma pathophysiology. There is also a growing body of evidence that mast cells contribute to other inflammatory and fibrotic lung diseases such as chronic obstructive pulmonary disease and idiopathic pulmonary fibrosis. This review discusses the role that mast cells play in airway diseases and highlights how mast cell microlocalisation within specific lung compartments and their cellular interactions are likely to be critical for their effector function in disease. Published by Elsevier B.V.

Serosurvey for the Prevalence of Brucella Canis Antibodies in Dogs in Central Ohio

DTIC Science & Technology

1983-01-01

negative urease production ...................... positive indol production ....................... negative1 0 ,58 ,6 2 citrate utilization...retained its spermatogenic ability. The epididymides had ac- cumulations of lymphocytes in the interstitial cell layers ranging from a few cells to large...There was diffuse in- filtration of plasma cells between the inner plexiform layer and the nerve fiber layer of the retina. Some serous exudate and a few

[Cellular mechanism of the generation of spontaneous activity in gastric muscle].

PubMed

Nakamura, Eri; Kito, Yoshihiko; Fukuta, Hiroyasu; Yanai, Yoshimasa; Hashitani, Hikaru; Yamamoto, Yoshimichi; Suzuki, Hikaru

2004-03-01

In gastric smooth muscles, interstitial cells of Cajal (ICC) might be the pacemaker cells of spontaneous activities since ICC are rich in mitochondria and are connected with smooth muscle cells via gap junctions. Several types of ICC are distributed widely in the stomach wall. A group of ICC distributed in the myenteric layer (ICC-MY) were the pacemaker cells of gastrointestinal smooth muscles. Pacemaker potentials were generated in ICC-MY, and the potentials were conducted to circular smooth muscles to trigger slow waves and also conducted to longitudinal muscles to form follower potentials. In circular muscle preparations, interstitial cells distributed within muscle bundles (ICC-IM) produced unitary potentials, which were conducted to circular muscles to form slow potentials by summation. In mutant mice lacking inositol trisphosphate (IP(3)) receptor, slow waves were absent in gastric smooth muscles. The generation of spontaneous activity was impaired by the inhibition of Ca(2+)-release from internal stores through IP(3) receptors, inhibition of mitochondrial Ca(2+)-handling with proton pump inhibitors, and inhibition of ATP-sensitive K(+)-channels at the mitochondrial inner membrane. These results suggested that mitochondrial Ca(2+)-handling causes the generation of spontaneous activity in pacemaker cells. Possible involvement of protein kinase C (PKC) in the Ca(2+) signaling system was also suggested.

FoxO and Stress Responses in the Cnidarian Hydra vulgaris

PubMed Central

Bridge, Diane; Theofiles, Alexander G.; Holler, Rebecca L.; Marcinkevicius, Emily; Steele, Robert E.; Martínez, Daniel E.

2010-01-01

Background In the face of changing environmental conditions, the mechanisms underlying stress responses in diverse organisms are of increasing interest. In vertebrates, Drosophila, and Caenorhabditis elegans, FoxO transcription factors mediate cellular responses to stress, including oxidative stress and dietary restriction. Although FoxO genes have been identified in early-arising animal lineages including sponges and cnidarians, little is known about their roles in these organisms. Methods/Principal Findings We have examined the regulation of FoxO activity in members of the well-studied cnidarian genus Hydra. We find that Hydra FoxO is expressed at high levels in cells of the interstitial lineage, a cell lineage that includes multipotent stem cells that give rise to neurons, stinging cells, secretory cells and gametes. Using transgenic Hydra that express a FoxO-GFP fusion protein in cells of the interstitial lineage, we have determined that heat shock causes localization of the fusion protein to the nucleus. Our results also provide evidence that, as in bilaterian animals, Hydra FoxO activity is regulated by both Akt and JNK kinases. Conclusions These findings imply that basic mechanisms of FoxO regulation arose before the evolution of bilaterians and raise the possibility that FoxO is involved in stress responses of other cnidarian species, including corals. PMID:20657733

FoxO and stress responses in the cnidarian Hydra vulgaris.

PubMed

Bridge, Diane; Theofiles, Alexander G; Holler, Rebecca L; Marcinkevicius, Emily; Steele, Robert E; Martínez, Daniel E

2010-07-21

In the face of changing environmental conditions, the mechanisms underlying stress responses in diverse organisms are of increasing interest. In vertebrates, Drosophila, and Caenorhabditis elegans, FoxO transcription factors mediate cellular responses to stress, including oxidative stress and dietary restriction. Although FoxO genes have been identified in early-arising animal lineages including sponges and cnidarians, little is known about their roles in these organisms. We have examined the regulation of FoxO activity in members of the well-studied cnidarian genus Hydra. We find that Hydra FoxO is expressed at high levels in cells of the interstitial lineage, a cell lineage that includes multipotent stem cells that give rise to neurons, stinging cells, secretory cells and gametes. Using transgenic Hydra that express a FoxO-GFP fusion protein in cells of the interstitial lineage, we have determined that heat shock causes localization of the fusion protein to the nucleus. Our results also provide evidence that, as in bilaterian animals, Hydra FoxO activity is regulated by both Akt and JNK kinases. These findings imply that basic mechanisms of FoxO regulation arose before the evolution of bilaterians and raise the possibility that FoxO is involved in stress responses of other cnidarian species, including corals.

A comparison of the in vitro cytotoxicity of conventional and resin modified glass ionomer cements

PubMed Central

Selimović-Dragaš, Mediha; Huseinbegović, Amina; Kobašlija, Sedin; Hatibović-Kofman, Šahza

2012-01-01

To evaluate cytotoxicity of experimental conventional and resin modified glass-ionomer cements on UMR-106 osteoblast cell cultures and cell cultures of NIH3T3 mouse fibroblasts specimens were prepared for every experimental material and divided into: group 1. Conventional glass-ionomer cements: GC Fuji IX GP Fast, GC Fuji Triage and Ketac Silver; group 2. Resin modified glass-ionomer cements: GC Fuji II LC, GC Fuji Plus and Vitrebond; group 3. Positive control was presented by specimens of composite Vit-l-ecence® and negative control-group 4. was presented by α-minimum essential medium for UMR-106 – osteoblast-like cells and Dulbecco’s Modified Eagle’s Medium for NIH3T3 mouse fibroblast cells. Both cell cultures were exposed to 10% of eluate of each single specimen of each experimental material. Experimental dishes were incubated for 24 h. Cell metabolism was evaluated using methyltetrazolium assay. Kruskal-Wallis test and Tukey-Kramer post hoc test for the materials evaluated on NIH3T3 mouse fibroblast cells, as well as UMR-106 osteoblast-like cells showed significantly more cytotoxicity of RMGICs, predominantly Vitrebond to both GICs and composite-Vit-l-ecence®. The lowest influence on cell’s metabolism on UMR-106 osteoblas-like cells was shown by Ketac Silver and the lowest influence on cell’s metabolism on NIH3T3 mouse fibroblast cells was shown by Fuji IX GP Fast. Statistical evaluation of sensitivity of cell lines UMR-106 osteoblast-like cells and NIH3T3 mouse fibroblast cells, using Mann-Whitney test, showed that NIH3T3 mouse fibroblast cells were more sensitive for the evaluation of cytotoxicity of dental materials. PMID:23198945

Progression of Mouse Skin Carcinogenesis Is Associated with Increased Erα Levels and Is Repressed by a Dominant Negative Form of Erα

PubMed Central

Michalopoulos, Ioannis; Sideridou, Maria; Tsimaratou, Katerina; Christodoulou, Ioannis; Pyrillou, Katerina; Gorgoulis, Vassilis; Vlahopoulos, Spiros; Zoumpourlis, Vassilis

2012-01-01

Estrogen receptors (ER), namely ERα and ERβ, are hormone-activated transcription factors with an important role in carcinogenesis. In the present study, we aimed at elucidating the implication of ERα in skin cancer, using chemically-induced mouse skin tumours, as well as cell lines representing distinct stages of mouse skin oncogenesis. First, using immunohistochemical staining we showed that ERα is markedly increased in aggressive mouse skin tumours in vivo as compared to the papilloma tumours, whereas ERβ levels are low and become even lower in the aggressive spindle tumours of carcinogen-treated mice. Then, using the multistage mouse skin carcinogenesis model, we showed that ERα gradually increases during promotion and progression stages of mouse skin carcinogenesis, peaking at the most aggressive stage, whereas ERβ levels only slightly change throughout skin carcinogenesis. Stable transfection of the aggressive, spindle CarB cells with a dominant negative form of ERα (dnERα) resulted in reduced ERα levels and reduced binding to estrogen responsive elements (ERE)-containing sequences. We characterized two highly conserved EREs on the mouse ERα promoter through which dnERα decreased endogenous ERα levels. The dnERα-transfected CarB cells presented altered protein levels of cytoskeletal and cell adhesion molecules, slower growth rate and impaired anchorage-independent growth in vitro, whereas they gave smaller tumours with extended latency period of tumour onset in vivo. Our findings suggest an implication of ERα in the aggressiveness of spindle mouse skin cancer cells, possibly through regulation of genes affecting cell shape and adhesion, and they also provide hints for the effective targeting of spindle cancer cells by dnERα. PMID:22870269

Recombinant mouse periostin ameliorates coronal sutures fusion in Twist1+/- mice.

PubMed

Bai, Shanshan; Li, Dong; Xu, Liang; Duan, Huichuan; Yuan, Jie; Wei, Min

2018-04-17

Saethre-Chotzen syndrome is an autosomal dominantly inherited disorder caused by mutations in the twist family basic helix-loop-helix transcription factor 1 (TWIST1) gene. Surgical procedures are frequently required to reduce morphological and functional defects in patients with Saethre-Chotzen syndrome. Therefore, the development of noninvasive procedures to treat Saethre-Chotzen syndrome is critical. We identified that periostin, which is an extracellular matrix protein that plays an important role in both bone and connective tissues, is downregulated in craniosynostosis patients. We aimed to verify the effects of different concentrations (0, 50, 100, and 200 μg/l) of recombinant mouse periostin in Twist1 +/- mice (a mouse model of Saethre-Chotzen syndrome) coronal suture cells in vitro and in vivo. Cell proliferation, migration, and osteogenic differentiation were observed and detected. Twist1 +/- mice were also injected with recombinant mouse periostin to verify the treatment effects. Cell Counting Kit-8 results showed that recombinant mouse periostin inhibited the proliferation of suture-derived cells in a time- and concentration-dependent manner. Cell migration was also suppressed when treated with recombinant mouse periostin. Real-time quantitative PCR and Western blotting results suggested that messenger ribonucleic acid and protein expression of alkaline phosphatase, bone sialoprotein, collagen type I, and osteocalcin were all downregulated after treatment with recombinant mouse periostin. However, the expression of Wnt-3a, Wnt-1, and β-catenin were upregulated. The in vivo results demonstrated that periostin-treated Twist1 +/- mice showed patent coronal sutures in comparison with non-treated Twist1 +/- mice which have coronal craniosynostosis. Our results suggest that recombinant mouse periostin can inhibit coronal suture cell proliferation and migration and suppress osteogenic differentiation of suture-derived cells via Wnt canonical signaling, as well as ameliorate coronal suture fusion in Twist1 +/- mice.

Photocrosslinking of gelatin macromers to synthesize porous hydrogels that promote valvular interstitial cell function.

PubMed

Benton, Julie A; DeForest, Cole A; Vivekanandan, Vani; Anseth, Kristi S

2009-11-01

The development of novel three-dimensional cell culture platforms for the culture of aortic valvular interstitial cells (VICs) has been fraught with many challenges. Although the most tunable, purely synthetic systems have not been successful at promoting cell survivability or function. On the other hand, entirely natural materials lack mechanical integrity. Here we explore a novel hybrid system consisting of gelatin macromers synthetically modified with methacrylate functionalities allowing for photoencapsulation of cells. Scanning electron microscopy observations show a microporous structure induced during polymerization within the hydrogel. This porous structure was tunable with polymerization rate and did not appear to have interconnected pores. Treatment with collagenase caused bulk erosion indicating enzymatic degradation controls the matrix remodeling. VICs, an important cell line for heart valve tissue engineering, were photoencapsulated and examined for cell-directed migration and differentiation. VICs were able to achieve their native morphology within 2 weeks of culture. The addition of the pro-fibrotic growth factor, transforming growth factor-beta1, accelerated this process and also was capable of inducing enhanced alpha-smooth muscle actin and collagen-1 expression, indicating a differentiation from quiescent fibroblasts to active myofibroblasts as demonstrated by quantitative real-time polymerase chain reaction and immunohistochemistry. Although these studies were limited to VICs, this novel hydrogel system may also be useful for studying other fibroblastic cell types.

Fluid Mechanics, Arterial Disease, and Gene Expression.

PubMed

Tarbell, John M; Shi, Zhong-Dong; Dunn, Jessilyn; Jo, Hanjoong

2014-01-01

This review places modern research developments in vascular mechanobiology in the context of hemodynamic phenomena in the cardiovascular system and the discrete localization of vascular disease. The modern origins of this field are traced, beginning in the 1960s when associations between flow characteristics, particularly blood flow-induced wall shear stress, and the localization of atherosclerotic plaques were uncovered, and continuing to fluid shear stress effects on the vascular lining endothelial) cells (ECs), including their effects on EC morphology, biochemical production, and gene expression. The earliest single-gene studies and genome-wide analyses are considered. The final section moves from the ECs lining the vessel wall to the smooth muscle cells and fibroblasts within the wall that are fluid me chanically activated by interstitial flow that imposes shear stresses on their surfaces comparable with those of flowing blood on EC surfaces. Interstitial flow stimulates biochemical production and gene expression, much like blood flow on ECs.

Axial elongation in mouse embryos involves mediolateral cell intercalation behavior in the paraxial mesoderm

NASA Astrophysics Data System (ADS)

Yen, WeiWei; Burdsal, Carol; Periasamy, Ammasi; Sutherland, Ann E.

2006-02-01

The cell mechanical and signaling pathways involved in gastrulation have been studied extensively in invertebrates and amphibians, such as Xenopus, and more recently in non-mammalian vertebrates such as zebrafish and chick. However, because culturing mouse embryos extra-utero is very difficult, this fundamental process has been least characterized in the mouse. As the primary mammalian model for genetics, biochemistry, and the study of human disease and birth defects, it is important to investigate how gastrulation proceeds in murine embryos. We have developed a method of using 4D multiphoton excitation microscopy and extra-utero culture to visualize and characterize the morphogenetic movements in mouse embryos dissected at 8.5 days of gestation. Cells are labeled by expression of an X chromosome-linked enhanced green fluorescent protein (EGFP) transgene. This method has provided a unique approach, where, for the first time, patterns of cell behavior in the notochord and surrounding paraxial mesoderm can be visualized and traced quantitatively. Our observations of mouse embryos reveal both distinct differences as well as striking similarities in patterned cell motility relative to other vertebrate models such as Xenopus, where axial extension is driven primarily by mediolateral oriented cell behaviors in the notochord and paraxial somitic mesoderm. Unlike Xenopus, the width of the mouse notochord remains the same between 4-somite stage and 8-somite stage embryos. This implies the mouse notochord plays a lesser role in driving axial extension compared to Xenopus, although intercalation may occur where the anterior region of the node becomes notochordal plate. In contrast, the width of mouse paraxial mesoderm narrows significantly during this period and cells within the paraxial mesoderm are both elongated and aligned perpendicular to the midline. In addition, these cells are observed to intercalate, consistent with a role for paraxial mesoderm in driving convergence and extension. These cell behaviors are similar to those characterized in the axial mesoderm of frog embryos during convergence and extension[1], and suggests that tissues may play different roles in axial elongation between the frog and the mouse.

Zinc stimulates glucose oxidation and glycemic control by modulating the insulin signaling pathway in human and mouse skeletal muscle cell lines.

PubMed

Norouzi, Shaghayegh; Adulcikas, John; Sohal, Sukhwinder Singh; Myers, Stephen

2018-01-01

Zinc is a metal ion that is an essential cell signaling molecule. Highlighting this, zinc is an insulin mimetic, activating cellular pathways that regulate cellular homeostasis and physiological responses. Previous studies have linked dysfunctional zinc signaling with several disease states including cancer, obesity, cardiovascular disease and type 2 diabetes. The present study evaluated the insulin-like effects of zinc on cell signaling molecules including tyrosine, PRSA40, Akt, ERK1/2, SHP-2, GSK-3β and p38, and glucose oxidation in human and mouse skeletal muscle cells. Insulin and zinc independently led to the phosphorylation of these proteins over a 60-minute time course in both mouse and human skeletal muscle cells. Similarly, utilizing a protein array we identified that zinc could active the phosphorylation of p38, ERK1/2 and GSK-3B in human and ERK1/2 and GSK-3B in mouse skeletal muscle cells. Glucose oxidation assays were performed on skeletal muscle cells treated with insulin, zinc, or a combination of both and resulted in a significant induction of glucose consumption in mouse (p<0.01) and human (p<0.05) skeletal muscle cells when treated with zinc alone. Insulin, as expected, increased glucose oxidation in mouse (p<0.001) and human (0.001) skeletal muscle cells, however the combination of zinc and insulin did not augment glucose consumption in these cells. Zinc acts as an insulin mimetic, activating key molecules implicated in cell signaling to maintain glucose homeostasis in mouse and human skeletal muscle cells. Zinc is an important metal ion implicated in several biological processes. The role of zinc as an insulin memetic in activating key signaling molecules involved in glucose homeostasis could provide opportunities to utilize this ion therapeutically in treating disorders associated with dysfunctional zinc signaling.

N-ethylmaleimide activates a Cl−-independent component of K+ flux in mouse erythrocytes

PubMed Central

Shmukler, Boris E.; Hsu, Ann; Alves, Jessica; Trudel, Marie; Rust, Marco B.; Hubner, Christian A.; Rivera, Alicia; Alper, Seth L.

2013-01-01

The K-Cl cotransporters (KCCs) of mouse erythrocytes exhibit higher basal activity than those of human erythrocytes, but are similarly activated by cell swelling, by hypertonic urea, and by staurosporine. However, the dramatic stimulation of human erythroid KCCs by N-ethylmaleimide (NEM) is obscured in mouse erythrocytes by a prominent NEM-stimulated K+ efflux that lacks Cl−-dependence. The NEM-sensitivity of Cl−-independent K+ efflux of mouse erythrocytes is lower than that of KCC. The genetically engineered absence of the K-Cl cotransporters KCC3 and KCC1 from mouse erythrocytes does not modify Cl−-independent K+ efflux. Mouse erythrocytes genetically devoid of the Gardos channel KCNN4 show increased NEM-sensitivity of both Cl−-independent K+ efflux and K-Cl cotransport. The increased NEM-sensitivity and stimulation magnitude of Cl−-independent K+ efflux in mouse erythrocytes expressing transgenic hypersickling human hemoglobin SAD (HbSAD) is independent of the presence of KCC3 and KCC1, but absence of KCNN4 reduces the stimulatory effect of HbSAD. NEM-stimulated Cl−-independent K+ efflux of mouse red cells is insensitive to ouabain and bumetanide, but partially inhibited by chloroquine, barium, and amiloride. The NEM-stimulated activity is modestly reduced at pH 6.0, but not significantly altered at pH 8.0, and abolished at 0°C. Although the molecular identity of this little-studied K+ efflux pathway of mouse erythrocytes remains unknown, it’s potential role in the pathophysiology of sickle red cell dehydration will be important for extrapolation of studies in mouse models of sickle cell disease to our understanding of humans with sickle cell anemia. PMID:23481459

N-ethylmaleimide activates a Cl(-)-independent component of K(+) flux in mouse erythrocytes.

PubMed

Shmukler, Boris E; Hsu, Ann; Alves, Jessica; Trudel, Marie; Rust, Marco B; Hubner, Christian A; Rivera, Alicia; Alper, Seth L

2013-06-01

The K-Cl cotransporters (KCCs) of mouse erythrocytes exhibit higher basal activity than those of human erythrocytes, but are similarly activated by cell swelling, by hypertonic urea, and by staurosporine. However, the dramatic stimulation of human erythroid KCCs by N-ethylmaleimide (NEM) is obscured in mouse erythrocytes by a prominent NEM-stimulated K(+) efflux that lacks Cl(-)-dependence. The NEM-sensitivity of Cl(-)-independent K(+) efflux of mouse erythrocytes is lower than that of KCC. The genetically engineered absence of the K-Cl cotransporters KCC3 and KCC1 from mouse erythrocytes does not modify Cl(-)-independent K(+) efflux. Mouse erythrocytes genetically devoid of the Gardos channel KCNN4 show increased NEM-sensitivity of both Cl(-)-independent K(+) efflux and K-Cl cotransport. The increased NEM-sensitivity and stimulation magnitude of Cl(-)-independent K(+) efflux in mouse erythrocytes expressing transgenic hypersickling human hemoglobin SAD (HbSAD) are independent of the presence of KCC3 and KCC1, but absence of KCNN4 reduces the stimulatory effect of HbSAD. NEM-stimulated Cl(-)-independent K(+) efflux of mouse red cells is insensitive to ouabain and bumetanide, but partially inhibited by chloroquine, barium, and amiloride. The NEM-stimulated activity is modestly reduced at pH6.0 but not significantly altered at pH8.0, and is abolished at 0°C. Although the molecular identity of this little-studied K(+) efflux pathway of mouse erythrocytes remains unknown, its potential role in the pathophysiology of sickle red cell dehydration will be important for the extrapolation of studies in mouse models of sickle cell disease to our understanding of humans with sickle cell anemia. Copyright © 2013 Elsevier Inc. All rights reserved.

AMPK/p53 Axis Is Essential for α-Lipoic Acid-Regulated Metastasis in Human and Mouse Colon Cancer Cells.

PubMed

Park, Sunmi; Choi, Seung Kug; Choi, Yura; Moon, Hyun-Seuk

2015-10-01

α-Lipoic acid (ALA) has an anticancer property of lung, cervix, and prostate cancer cells. However, direct evidence that ALA contributes to the development of colon cancer has not been fully elucidated. In addition, no previous studies have evaluated whether ALA may regulate malignant potential, such as adhesion, invasion, and colony formation of colon cancer cells. To address the aforementioned questions, we conducted in vitro ALA signaling studies using human (HT29) and mouse (MCA38) colon cancer cell lines. We observed that cell proliferation is reduced by ALA administration in a dose-dependent manner in human and mouse colon cancer cell lines. Specifically, 0.5 to 1 mM concentration of ALA significantly decreased cell proliferation when compared with control. Similarly, we found that ALA downregulates adhesion, invasion, and colony formation. Finally, we observed that ALA activates p53 and AMPK signaling pathways in human and mouse colon cancer cells. We found for the first time that ALA suppresses cell proliferation and malignant potential via p53 and AMPK signaling pathways in human and mouse colon cancer cells. These new and early mechanistic studies provide a causal role of ALA in colon cancer, suggesting that ALA might be a useful agent in the management or chemoprevention of colon cancer.

Potassium dependent rescue of a myopathy with core-like structures in mouse

PubMed Central

Hanson, M Gartz; Wilde, Jonathan J; Moreno, Rosa L; Minic, Angela D; Niswander, Lee

2015-01-01

Myopathies decrease muscle functionality. Mutations in ryanodine receptor 1 (RyR1) are often associated with myopathies with microscopic core-like structures in the muscle fiber. In this study, we identify a mouse RyR1 model in which heterozygous animals display clinical and pathological hallmarks of myopathy with core-like structures. The RyR1 mutation decreases sensitivity to activated calcium release and myoplasmic calcium levels, subsequently affecting mitochondrial calcium and ATP production. Mutant muscle shows a persistent potassium leak and disrupted expression of regulators of potassium homeostasis. Inhibition of KATP channels or increasing interstitial potassium by diet or FDA-approved drugs can reverse the muscle weakness, fatigue-like physiology and pathology. We identify regulators of potassium homeostasis as biomarkers of disease that may reveal therapeutic targets in human patients with myopathy of central core disease (CCD). Altogether, our results suggest that amelioration of potassium leaks through potassium homeostasis mechanisms may minimize muscle damage of myopathies due to certain RyR1 mutations. DOI: http://dx.doi.org/10.7554/eLife.02923.001 PMID:25564733

CRISPR-Mediated Knockout of Cybb in NSG Mice Establishes a Model of Chronic Granulomatous Disease for Human Stem-Cell Gene Therapy Transplants.

PubMed

Sweeney, Colin L; Choi, Uimook; Liu, Chengyu; Koontz, Sherry; Ha, Seung-Kwon; Malech, Harry L

2017-07-01

Chronic granulomatous disease (CGD) is characterized by defects in the production of microbicidal reactive oxygen species (ROS) by phagocytes. Testing of gene and cell therapies for the treatment of CGD in human hematopoietic cells requires preclinical transplant models. The use of the lymphocyte-deficient NOD.Cg-Prkdc scid Il2rg tm1Wjl/ SzJ (NSG) mouse strain for human hematopoietic cell xenografts to test CGD therapies is complicated by the presence of functional mouse granulocytes capable of producing ROS for subsequent bacterial and fungal killing. To establish a phagocyte-defective mouse model of X-linked CGD (X-CGD) in NSG mice, clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 was utilized for targeted knockout of mouse Cybb on the X-chromosome by microinjection of NSG mouse zygotes with Cas9 mRNA and CRISPR single-guide RNA targeting Cybb exon 1 or exon 3. This resulted in a high incidence of indel formation at the CRISPR target site, with all mice exhibiting deletions in at least one Cybb allele based on sequence analysis of tail snip DNA. A female mouse heterozygous for a 235-bp deletion in Cybb exon 1 was bred to an NSG male to establish the X-CGD NSG mouse strain, NSG.Cybb[KO]. Resulting male offspring with the 235 bp deletion were found to be defective for production of ROS by neutrophils and other phagocytes, and demonstrated increased susceptibility to spontaneous bacterial and fungal infections with granulomatous inflammation. The establishment of the phagocyte-defective NSG.Cybb[KO] mouse model enables the in vivo assessment of gene and cell therapy strategies for treating CGD in human hematopoietic cell transplants without obfuscation by functional mouse phagocytes, and may also be useful for modeling other phagocyte disorders in humanized NSG mouse xenografts.

Framework 'interstitial' oxygen in La(10)(GeO(4))(5-)(GeO(5))O(2) apatite electrolyte.

PubMed

Pramana, Stevin S; Klooster, Wim T; White, T J

2007-08-01

Oxygen conduction at low temperatures in apatites make these materials potentially useful as electrolytes in solid-oxide fuel cells, but our understanding of the defect structures enabling ion migration is incomplete. While conduction along [001] channels is dominant, considerable inter-tunnel mobility has been recognized. Using neutron powder diffraction of stoichiometric 'La(10)(GeO(4))(6)O(3)', it has been shown that this compound is more correctly described as an La(10)(GeO(4))(5-)(GeO(5))O(2) apatite, in which high concentrations of interstitial oxygen reside within the channel walls. It is suggested that these framework interstitial O atoms provide a reservoir of ions that can migrate into the conducting channels of apatite, via a mechanism of inter-tunnel oxygen diffusion that transiently converts GeO(4) tetrahedra to GeO(5) distorted trigonal bipyramids. This structural modification is consistent with known crystal chemistry and may occur generally in oxide apatites.

Advances in the treatment of rheumatic interstitial lung disease.

PubMed

Vassallo, Robert; Thomas, Charles F

2004-05-01

Interstitial lung disease frequently complicates the rheumatic diseases. The purpose of this review is to outline recent advances and current concepts regarding the management of these interstitial lung diseases. Several histologic lesions cause interstitial lung disease in rheumatic diseases, including nonspecific interstitial pneumonia, usual interstitial pneumonia, organizing pneumonia, lymphocytic interstitial pneumonia, desquamative interstitial pneumonia, and acute interstitial pneumonia. Although the relative frequency of occurrence of these histopathologic lesions is not definitively established, it seems that nonspecific interstitial pneumonia accounts for a large proportion of rheumatic disease-associated interstitial lung diseases. Although usual interstitial pneumonia generally responds poorly to corticosteroid therapy, other forms of interstitial pneumonia are often steroid responsive and have a more favorable long-term prognosis. Pulmonary hypertension is increasingly recognized as a complication of these interstitial lung diseases. Treatment of pulmonary hypertension in these patients provides clinical benefit and may suppress pulmonary inflammation and fibrosis. Lung transplantation is a treatment option for selected patients with severe pulmonary involvement and limited life expectancy. Interstitial lung disease is common in the rheumatic diseases, may be caused by a variety of lesions that respond differently to treatment, and may lead to the development of pulmonary hypertension. Whether the prognosis of interstitial lung disease associated with rheumatic disease is similar to that associated with the idiopathic interstitial pneumonias is not known. Treatment of these interstitial lung diseases should take into account the specific histologic lesion, the activity of the underlying rheumatic disease, and associated pulmonary hypertension, if present. The diagnosis of a rheumatic disease is no longer an absolute contraindication to lung transplantation.

Subregional localization and characterization of Ly6aGFP-expressing hematopoietic cells in the mouse embryonic head.

PubMed

Li, Zhuan; Vink, Chris S; Mariani, Samanta A; Dzierzak, Elaine

2016-08-01

Hematopoietic cell generation in the midgestation mouse embryo occurs through the natural transdifferentiation of temporally and spatially restricted set of hemogenic endothelial cells. These cells take on hematopoietic fate in the aorta, vitelline and umbilical arteries and appear as hematopoietic cell clusters that emerge from the vascular wall. Genetic and live imaging data have supported this. Recently, the embryonic head has been shown to contain fully functional hematopoietic stem cells (HSC). By lineage tracing, cerebrovascular specific endothelial cells were shown to contribute to the postnatal mouse hematopoietic system. Since Ly6aGFP is a marker of all HSCs, some hematopoietic cluster cells and hemogenic endothelial cells in the midgestation mouse aorta, we examine here whether embryonic head HSCs and vascular endothelial cells are positive for this marker. Whereas some head vasculature, single hematopoietic cells and all HSCs are Ly6aGFP expressing, we do not find clusters of hematopoietic cells emerging from the cerebrovasculature that are characteristic of endothelial-to-hematopoietic transition. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Cinemicrographic study of the cell movement in the primitive-streak-stage mouse embryo.

PubMed

Nakatsuji, N; Snow, M H; Wylie, C C

1986-07-01

Migration of the mesoderm cells in the primitive-streak-stage mouse embryo was directly studied by cinemicrography using whole embryo culture and Nomarski differential interference contrast optics. Relative transparency and small size of the early mouse embryos enabled direct observation of the individual cells and their cell processes. Seven-day-old mouse embryos were isolated and cultured in a small chamber in a medium consisting of 50% rat serum and 50% Dulbecco's modified minimum essential medium. The mesoderm cells move away from the primitive streak in both anterior and antimesometrial (distal) directions at a mean velocity of 46 micron h-1. They extend cell processes and constantly change cell shape. They do not translocate extensively as isolated single cells, but usually maintain attachment to other mesoderm cells. They show frequent cell division preceded by rounding up of the cell bodies, and accompanied by vigorous blebbing before and after cytokinesis. This study shows that it is possible to examine the motility of embryonic cells inside the mammalian embryo by direct observation if the embryo is small and transparent enough for the use of the Nomarski optics.

Data from SILAC-based quantitative analysis of lysates from mouse microglial cells treated with Withaferin A (WA).

PubMed

Narayan, Malathi; Seeley, Kent W; Jinwal, Umesh K

2016-06-01

Mass spectrometry data collected in a study analyzing the effect of withaferin A (WA) on a mouse microglial (N9) cell line is presented in this article. Data was collected from SILAC-based quantitative analysis of lysates from mouse microglial cells treated with either WA or DMSO vehicle control. This article reports all the proteins that were identified in this analysis. The data presented here is related to the published research article on the effect of WA on the differential regulation of proteins in mouse microglial cells [1]. Mass spectrometry data has also been deposited in the ProteomeXchange with the identifier PXD003032.

Bardoxolone Methyl Evaluation in Patients With Pulmonary Hypertension (PH) - LARIAT

ClinicalTrials.gov

2018-06-08

Pulmonary Arterial Hypertension; Pulmonary Hypertension; Interstitial Lung Disease; Idiopathic Interstitial Pneumonia; Idiopathic Pulmonary Fibrosis; Sarcoidosis; Respiratory Bronchiolitis Associated Interstitial Lung Disease; Desquamative Interstitial Pneumonia; Cryptogenic Organizing Pneumonia; Acute Interstitial Pneumonitis; Idiopathic Lymphoid Interstitial Pneumonia; Idiopathic Pleuroparenchymal Fibroelastosis

Mammary Cancer and Activation of Transposable Elements

DTIC Science & Technology

2013-09-01

formal monthly electronic lab meeting between Peaston lab and Edwards lab. And regularly hold meetings. • An informal schedule was set up with a plan...cytes and ADS-derived induced pluripotent stem cells (ADS-iPSCs) (19) and primary mouse ES cells to isolated sperm and oocytes (20). We selected an...051 59 5 92% H9-IMR90 5875 7 669 782 605 58 91% oocyte - ES cell (mouse) 4727 1 204 883 334 25 93% sperm - ES cell (mouse) 4580 4 364 748 1027 104 91

Mouse Embryonic Stem Cell Adherent Cell Differentiation and Cytotoxicity (ACDC) assay

EPA Science Inventory

The Embryonic Stem Cell Test (EST) is an assay which evaluates xenobiotic-induced effects using three endpoints: mouse embryonic stem cell (mESC) differentiation, mESC viability, and 3T3-cell viability. Our research goal was to develop an improved high-throughput assay by establi...

Do we understand any more about bladder interstitial cells?-ICI-RS 2013.

PubMed

Kanai, Anthony; Fry, Christopher; Hanna-Mitchell, Ann; Birder, Lori; Zabbarova, Irina; Bijos, Dominika; Ikeda, Youko

2014-06-01

To present a brief review on discussions from "Do we understand any more about lower urinary tract interstitial cells?" session at the 2013 International Consultation on Incontinence-Research Society (ICI-RS) meeting in Bristol, UK. Discussion focused on bladder interstitial cell (IC) subtypes, their localization and characterization, and communication between themselves, the urothelium, and detrusor smooth muscle. The role of ICs in bladder pathologies and new methods for studying ICs were also addressed. ICs have been studied extensively in the lower urinary tract and have been characterized based on comparisons with ICs of Cajal in the gastro-intestinal tract. In fetal bladders it is believed that ICs drive intrinsic contractions to expel urine through the urachus. These contractions diminish postpartum as bladder innervation develops. Voiding in human neonates occurs when filling triggers a spinal cord reflex that contracts the detrusor; in rodents, maternal stimulation of the perineum triggers voiding. Following spinal cord injury, intrinsic contractions, and spinal micturition reflexes develop, similar to those seen during neonatal development. These enhanced contractions may stimulate nociceptive and mechanosensitive afferents contributing to neurogenic detrusor overactivity and incontinence. The IC-mediated activity is believed to be initiated in the lamina propria by responding to urothelial factors. These IC may act syncytially through gap junction coupling and modulate detrusor activity through unknown mechanisms. There has been a great deal of information discovered regarding bladder ICs, however, many of their (patho)physiological functions and mechanisms are still unclear and necessitates further research. Neurourol. Urodynam. 33:573-576, 2014. © 2014 Wiley Periodicals, Inc. © 2014 Wiley Periodicals, Inc.

A Contractile Network of Interstitial Cells of Cajal in the Supratarsal Mueller's Smooth Muscle Fibers With Sparse Sympathetic Innervation

PubMed Central

Yuzuriha, Shunsuke; Matsuo, Kiyoshi; Ban, Ryokuya; Yano, Shiharu; Moriizumi, Tetsuji

2012-01-01

Background: We previously reported that the supratarsal Mueller's muscle is innervated by both sympathetic efferent fibers and trigeminal proprioceptive afferent fibers, which function as mechanoreceptors-inducing reflexive contractions of both the levator and frontalis muscles. Controversy still persists regarding the role of the mechanoreceptors in Mueller's muscle; therefore, we clinically and histologically investigated Mueller's muscle. Methods: We evaluated the role of phenylephrine administration into the upper fornix in contraction of Mueller's smooth muscle fibers and how intraoperative stretching of Mueller's muscle alters the degree of eyelid retraction in 20 patients with aponeurotic blepharoptosis. In addition, we stained Mueller's muscle in 7 cadavers with antibodies against α-smooth muscle actin, S100, tyrosine hydroxylase, c-kit, and connexin 43. Results: Maximal eyelid retraction occurred approximately 3.8 minutes after administration of phenylephrine and prolonged eyelid retraction for at least 20 minutes after administration. Intraoperative stretching of Mueller's muscle increased eyelid retraction due to its reflexive contraction. The tyrosine hydroxylase antibody sparsely stained postganglionic sympathetic nerve fibers, whereas the S100 and c-kit antibodies densely stained the interstitial cells of Cajal (ICCs) among Mueller's smooth muscle fibers. A connexin 43 antibody failed to stain Mueller's muscle. Conclusions: A contractile network of ICCs may mediate neurotransmission within Mueller's multiunit smooth muscle fibers that are sparsely innervated by postganglionic sympathetic fibers. Interstitial cells of Cajal may also serve as mechanoreceptors that reflexively contract Mueller's smooth muscle fibers, forming intimate associations with intramuscular trigeminal proprioceptive fibers to induce reflexive contraction of the levator and frontalis muscles. PMID:22359687

Mapping bone interstitial fluid movement: Displacement of ferritin tracer during histological processing

PubMed Central

Ciani, Cesare; Doty, Stephen B.; Fritton, Susannah P.

2014-01-01

Bone interstitial fluid flow is thought to play a fundamental role in the mechanical stimulation of bone cells, either via shear stresses or cytoskeletal deformations. Recent evidence indicates that osteocytes are surrounded by a fiber matrix that may be involved in the mechanotransduction of external stimuli as well as in nutrient exchange. In our previous tracer studies designed to map how different-sized molecules travel through the bone porosities, we found that injected ferritin was confined to blood vessels and did not pass into the mineralized matrix. However, other investigators have shown that ferritin forms halo-shaped labeling that enters the mineralized matrix around blood vessels. This labeling is widely used to explain normal interstitial fluid movement in bone; in particular, it is said to demonstrate bulk centrifugal interstitial fluid movement away from a highly pressurized vascular porosity. In addition, appositional ferritin fronts are said to demonstrate centrifugal interstitial fluid movement from the medullary canal to the periosteal surface. The purpose of this study was to investigate the conflicting ferritin labeling results by evaluating the role of different histological processes in the formation of ferritin “halos.” Ferritin was injected into the rat vasculature and allowed to circulate for 5 min. Samples obtained from tibiae were reacted for different times with Perl's reagent and then were either paraffin-embedded or sectioned with a cryostat. Halo-like labeling surrounding vascular pores was found in all groups, ranging from 1.2–3.9% for the samples treated with the shortest histological processes (unembedded, frozen sections) to 5.6–15% for the samples treated with the longest histological processes (paraffin-embedded sections). These results indicate that different histological processing methods are able to create ferritin “halos,” with some processing methods allowing more redistribution of the ferritin tracer than others. Based on these results and the fact that “halo” labeling has not been found with any other tracer, as we seek to further delineate the movement of interstitial fluid and the role it plays in bone mechanotransduction, we believe that ferritin “halo” labeling should not be used to demonstrate physiological bone interstitial fluid flow. PMID:15964255

Mapping bone interstitial fluid movement: displacement of ferritin tracer during histological processing.

PubMed

Ciani, Cesare; Doty, Stephen B; Fritton, Susannah P

2005-09-01

Bone interstitial fluid flow is thought to play a fundamental role in the mechanical stimulation of bone cells, either via shear stresses or cytoskeletal deformations. Recent evidence indicates that osteocytes are surrounded by a fiber matrix that may be involved in the mechanotransduction of external stimuli as well as in nutrient exchange. In our previous tracer studies designed to map how different-sized molecules travel through the bone porosities, we found that injected ferritin was confined to blood vessels and did not pass into the mineralized matrix. However, other investigators have shown that ferritin forms halo-shaped labeling that enters the mineralized matrix around blood vessels. This labeling is widely used to explain normal interstitial fluid movement in bone; in particular, it is said to demonstrate bulk centrifugal interstitial fluid movement away from a highly pressurized vascular porosity. In addition, appositional ferritin fronts are said to demonstrate centrifugal interstitial fluid movement from the medullary canal to the periosteal surface. The purpose of this study was to investigate the conflicting ferritin labeling results by evaluating the role of different histological processes in the formation of ferritin "halos." Ferritin was injected into the rat vasculature and allowed to circulate for 5 min. Samples obtained from tibiae were reacted for different times with Perl's reagent and then were either paraffin-embedded or sectioned with a cryostat. Halo-like labeling surrounding vascular pores was found in all groups, ranging from 1.2-3.9% for the samples treated with the shortest histological processes (unembedded, frozen sections) to 5.6-15% for the samples treated with the longest histological processes (paraffin-embedded sections). These results indicate that different histological processing methods are able to create ferritin "halos," with some processing methods allowing more redistribution of the ferritin tracer than others. Based on these results and the fact that "halo" labeling has not been found with any other tracer, as we seek to further delineate the movement of interstitial fluid and the role it plays in bone mechanotransduction, we believe that ferritin "halo" labeling should not be used to demonstrate physiological bone interstitial fluid flow.

Infrared spectroscopic analysis of human interstitial fluid in vitro and in vivo using FT-IR spectroscopy and pulsed quantum cascade lasers (QCL): Establishing a new approach to non invasive glucose measurement.

PubMed

Pleitez, Miguel; von Lilienfeld-Toal, Hermann; Mäntele, Werner

2012-01-01

Interstitial fluid, i.e. the liquid present in the outermost layer of living cells of the skin between the Stratum corneum and the Stratum spinosum, was analyzed by Fourier transform infrared spectroscopy and by infrared spectroscopy using pulsed quantum cascade infrared lasers with photoacoustic detection. IR spectra of simulated interstitial fluid samples and of real samples from volunteers in the 850-1800cm(-1) range revealed that the major components of interstitial fluid are albumin and glucose within the physiological range, with only traces of sodium lactate if at all. The IR absorbance of glucose in interstitial fluid in vivo was probed in healthy volunteers using a setup with quantum cascade lasers and photoacoustic detection previously described. A variation of blood glucose between approx. 80mg/dl and 250mg/dl in the volunteers was obtained using the standard oral glucose tolerance test (OGT). At two IR wavelengths, 1054cm(-1) and 1084cm(-1), a reasonable correlation between the photoacoustic signal from the skin and the blood glucose value as determined by conventional glucose test sticks using blood from the finger tip was obtained. The infrared photoacoustic glucose signal (PAGS) may serve as the key for a non-invasive glucose measurement, since the glucose content in interstitial fluid closely follows blood glucose in the time course and in the level (a delay of some minutes and a level of approx. 80-90% of the glucose level in blood). Interstitial fluid is present in skin layers at a depth of only 15-50μm and is thus within the reach of mid-IR energy in an absorbance measurement. A non-invasive glucose measurement for diabetes patients based on mid-infrared quantum cascade lasers and photoacoustic detection could replace the conventional measurement using enzymatic test stripes and a drop of blood from the finger tip, thus reducing pain and being a cost-efficient alternative for millions of diabetes patients. Copyright © 2011 Elsevier B.V. All rights reserved.

Infrared spectroscopic analysis of human interstitial fluid in vitro and in vivo using FT-IR spectroscopy and pulsed quantum cascade lasers (QCL): Establishing a new approach to non invasive glucose measurement

NASA Astrophysics Data System (ADS)

Pleitez, Miguel; von Lilienfeld-Toal, Hermann; Mäntele, Werner

2012-01-01

Interstitial fluid, i.e. the liquid present in the outermost layer of living cells of the skin between the Stratum corneum and the Stratum spinosum, was analyzed by Fourier transform infrared spectroscopy and by infrared spectroscopy using pulsed quantum cascade infrared lasers with photoacoustic detection. IR spectra of simulated interstitial fluid samples and of real samples from volunteers in the 850-1800 cm -1 range revealed that the major components of interstitial fluid are albumin and glucose within the physiological range, with only traces of sodium lactate if at all. The IR absorbance of glucose in interstitial fluid in vivo was probed in healthy volunteers using a setup with quantum cascade lasers and photoacoustic detection previously described [11]. A variation of blood glucose between approx. 80 mg/dl and 250 mg/dl in the volunteers was obtained using the standard oral glucose tolerance test (OGT). At two IR wavelengths, 1054 cm -1 and 1084 cm -1, a reasonable correlation between the photoacoustic signal from the skin and the blood glucose value as determined by conventional glucose test sticks using blood from the finger tip was obtained. The infrared photoacoustic glucose signal (PAGS) may serve as the key for a non-invasive glucose measurement, since the glucose content in interstitial fluid closely follows blood glucose in the time course and in the level (a delay of some minutes and a level of approx. 80-90% of the glucose level in blood). Interstitial fluid is present in skin layers at a depth of only 15-50 μm and is thus within the reach of mid-IR energy in an absorbance measurement. A non-invasive glucose measurement for diabetes patients based on mid-infrared quantum cascade lasers and photoacoustic detection could replace the conventional measurement using enzymatic test stripes and a drop of blood from the finger tip, thus reducing pain and being a cost-efficient alternative for millions of diabetes patients.