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Sample records for human tissue kallikrein

  1. Crystal structure of recombinant human tissue kallikrein at 2.0 A resolution.

    PubMed Central

    Katz, B. A.; Liu, B.; Barnes, M.; Springman, E. B.

    1998-01-01

    Human tissue kallikrein, a trypsin-like serine protease involved in blood pressure regulation and inflammation processes, was expressed in a deglycosylated form at high levels in Pichia pastoris, purified, and crystallized. The crystal structure at 2.0 A resolution is described and compared with that of porcine kallikrein and of other trypsin-like proteases. The active and S1 sites (nomenclature of Schechter I, Berger A, 1967, Biochem Biophys Res Commun 27:157-162) are similar to those of porcine kallikrein. Compared to trypsin, the S1 site is enlarged owing to the insertion of an additional residue, cis-Pro 219. The replacement Tyr 228 --> Ala further enlarges the S1 pocket. However, the replacement of Gly 226 in trypsin with Ser in human tissue kallikrein restricts accessibility of substrates and inhibitors to Asp 189 at the base of the S1 pocket; there is a hydrogen bond between O delta1Asp189 and O gammaSer226. These changes in the architecture of the S1 site perturb the binding of inhibitors or substrates from the modes determined or inferred for trypsin. The crystal structure gives insight into the structural differences responsible for changes in specificity in human tissue kallikrein compared with other trypsin-like proteases, and into the structural basis for the unusual specificity of human tissue kallikrein in cleaving both an Arg-Ser and a Met-Lys peptide bond in its natural protein substrate, kininogen. A Zn+2-dependent, small-molecule competitive inhibitor of kallikrein (Ki = 3.3 microM) has been identified and the bound structure modeled to guide drug design. PMID:9568894

  2. Autoimmune diabetes is suppressed by treatment with recombinant human tissue Kallikrein-1.

    PubMed

    Maneva-Radicheva, Lilia; Amatya, Christina; Parker, Camille; Ellefson, Jacob; Radichev, Ilian; Raghavan, Arvind; Charles, Matthew L; Williams, Mark S; Robbins, Mark S; Savinov, Alexei Y

    2014-01-01

    The kallikrein-kinin system (KKS) comprises a cascade of proteolytic enzymes and biogenic peptides that regulate several physiological processes. Over-expression of tissue kallikrein-1 and modulation of the KKS shows beneficial effects on insulin sensitivity and other parameters relevant to type 2 diabetes mellitus. However, much less is known about the role of kallikreins, in particular tissue kallikrein-1, in type 1 diabetes mellitus (T1D). We report that chronic administration of recombinant human tissue kallikrein-1 protein (DM199) to non-obese diabetic mice delayed the onset of T1D, attenuated the degree of insulitis, and improved pancreatic beta cell mass in a dose- and treatment frequency-dependent manner. Suppression of the autoimmune reaction against pancreatic beta cells was evidenced by a reduction in the relative numbers of infiltrating cytotoxic lymphocytes and an increase in the relative numbers of regulatory T cells in the pancreas and pancreatic lymph nodes. These effects may be due in part to a DM199 treatment-dependent increase in active TGF-beta1. Treatment with DM199 also resulted in elevated C-peptide levels, elevated glucagon like peptide-1 levels and a reduction in dipeptidyl peptidase-4 activity. Overall, the data suggest that DM199 may have a beneficial effect on T1D by attenuating the autoimmune reaction and improving beta cell health. PMID:25259810

  3. Autoimmune Diabetes Is Suppressed by Treatment with Recombinant Human Tissue Kallikrein-1

    PubMed Central

    Maneva-Radicheva, Lilia; Amatya, Christina; Parker, Camille; Ellefson, Jacob; Radichev, Ilian; Raghavan, Arvind; Charles, Matthew L.; Williams, Mark S.; Robbins, Mark S.; Savinov, Alexei Y.

    2014-01-01

    The kallikrein-kinin system (KKS) comprises a cascade of proteolytic enzymes and biogenic peptides that regulate several physiological processes. Over-expression of tissue kallikrein-1 and modulation of the KKS shows beneficial effects on insulin sensitivity and other parameters relevant to type 2 diabetes mellitus. However, much less is known about the role of kallikreins, in particular tissue kallikrein-1, in type 1 diabetes mellitus (T1D). We report that chronic administration of recombinant human tissue kallikrein-1 protein (DM199) to non-obese diabetic mice delayed the onset of T1D, attenuated the degree of insulitis, and improved pancreatic beta cell mass in a dose- and treatment frequency-dependent manner. Suppression of the autoimmune reaction against pancreatic beta cells was evidenced by a reduction in the relative numbers of infiltrating cytotoxic lymphocytes and an increase in the relative numbers of regulatory T cells in the pancreas and pancreatic lymph nodes. These effects may be due in part to a DM199 treatment-dependent increase in active TGF-beta1. Treatment with DM199 also resulted in elevated C-peptide levels, elevated glucagon like peptide-1 levels and a reduction in dipeptidyl peptidase-4 activity. Overall, the data suggest that DM199 may have a beneficial effect on T1D by attenuating the autoimmune reaction and improving beta cell health. PMID:25259810

  4. Partial genetic deficiency in tissue kallikrein impairs adaptation to high potassium intake in humans.

    PubMed

    Monteiro, Joana S; Blanchard, Anne; Curis, Emmanuel; Chambrey, Régine; Jeunemaitre, Xavier; Azizi, Michel

    2013-12-01

    Inactivation of the tissue kallikrein gene in mice impairs renal handling of potassium due to enhanced H, K-ATPase activity, and induces hyperkalemia. We investigated whether the R53H loss-of-function polymorphism of the human tissue kallikrein gene affects renal potassium handling. In a crossover study, 30 R53R homozygous and 10 R53H heterozygous healthy males were randomly assigned to a low-sodium/high-potassium or a high-sodium/low-potassium diet to modulate tissue kallikrein synthesis. On the seventh day of each diet, participants were studied before and during a 2-h infusion of furosemide to stimulate distal potassium secretion. Urinary kallikrein activity was significantly lower in R53H than in R53R subjects on the low-sodium/high-potassium diet and was similarly reduced in both genotypes on high-sodium/low-potassium. Plasma potassium and renal potassium reabsorption were similar in both genotypes on an ad libitum sodium/potassium diet or after 7 days of a high-sodium/low-potassium diet. However, the median plasma potassium was significantly higher after 7 days of low-sodium/high-potassium diet in R53H than in R53R individuals. Urine potassium excretion and plasma aldosterone concentrations were similar. On the low-sodium/high-potassium diet, furosemide-induced decrease in plasma potassium was significantly larger in R53H than in R53R subjects. Thus, impaired tissue kallikrein stimulation by a low-sodium/high-potassium diet in R53H subjects with partial tissue kallikrein deficiency highlights an inappropriate renal adaptation to potassium load, consistent with experimental data in mice.

  5. Indirect Back-Titration ELISA: A New Format for Estimation of Human Tissue Kallikreins.

    PubMed

    Nasim, Faiz-Ul-Hassan; Ejaz, Samina; Ashraf, Muhammad; Ahmad, Gulzar

    2016-01-01

    Enzyme-linked immunosorbent assay (ELISA) is either based on sandwich, competitive, or inhibition type of format. However, these formats need 2 or 3 monoclonal antibodies (moAB) to estimate 1 antigen. To get a cost-effective, high throughput, ELISA for estimation of human tissue kallikreins we have now developed an indirect, back-titration style, Time Resolved ImmunoFluorometric (TRIF) ELISA that uses only 1 antigen-specific moAB and a general polyclonal antibody. Polystyrene microtiter plate wells coated with a capture antibody, a mouse moAB prepared against a specific human tissue kallikrein are allowed to interact either with the corresponding pure antigen, as the calibrator, or with the corresponding antigen present in a biological fluid or tissue extract. The detection antibody, anti-mouse IgG conjugated with alkaline phosphatase, is added to find the antigen-free immobilized capture moAB. Conjugated enzyme is allowed to hydrolyze diflunisal phosphate to produce a highly fluorescent complex. The fluorescence measured in TRIF mode corresponds to the antigen-free immobilized capture moAB and is used to quantify antigen-bound capture moAB. The detection antibody binds with the antigen-free capture moAB and strength of the signal correlates inversely with the amount of antigen bound to the capture moAB. With a minimum detection level of 20 ng/L the assay has no cross-reactivity with several test molecules. The method is sensitive, specific, applicable to a variety of biological samples, and cost-effective as it uses only 1 moAB and a polyclonal antibody. Using this assay, a single epitope can be estimated without purification. PMID:26180936

  6. Detection of kallikrein-like activity in inflamed synovial tissue.

    PubMed

    Sharma, J N; Zeitlin, I J; Deodhar, S D; Buchanan, W W

    1983-04-01

    The synovial tissue kallikrein of rheumatoid patients, arthritic dogs and normal dogs has been partially characterized and estimated. Both human and dog synovial tissue kinin-forming enzyme appears to be distinct from plasmin, plasma kallikrein and pancreatic kallikrein. In three rheumatoid arthritic patients, the mean synovial tissue kallikrein level was found to be 106 +/- 13.5 ng bradykinin equivalents (BK Equiv) per g per min. In nine arthritic dogs the mean synovial tissue kallikrein level, 60.5 +/- 4.5 ng BK Equiv per g per min, was found to be greater (p less than 0.05) than that of five normal dogs (21.6 +/- 5.3 ng BK Equiv per g per min). The possible significance of this finding is discussed. PMID:6553485

  7. Preclinical characterization of recombinant human tissue kallikrein-1 as a novel treatment for type 2 diabetes mellitus.

    PubMed

    Kolodka, Tadeusz; Charles, Matthew L; Raghavan, Arvind; Radichev, Ilian A; Amatya, Christina; Ellefson, Jacob; Savinov, Alexei Y; Nag, Abhijeet; Williams, Mark S; Robbins, Mark S

    2014-01-01

    Modulation of the kallikrein-kinin system (KKS) has been shown to have beneficial effects on glucose homeostasis and several other physiological responses relevant to the progression of type 2 diabetes mellitus (T2D). The importance of bradykinin and its receptors in mediating these responses is well documented, but the role of tissue kallikrein-1, the protease that generates bradykinin in situ, is much less understood. We developed and tested DM199, recombinant human tissue kallikrein-1 protein (rhKLK-1), as a potential novel therapeutic for T2D. Hyperinsulinemic-euglycemic clamp studies suggest that DM199 increases whole body glucose disposal in non-diabetic rats. Single-dose administration of DM199 in obese db/db mice and ZDF rats, showed an acute, dose-dependent improvement in whole-body glucose utilization. Sub-acute dosing for a week in ZDF rats improved glucose utilization, with a concomitant rise in fasting insulin levels and HOMA1-%B scores. After cessation of sub-acute dosing, fasting blood glucose levels were significantly lower in ZDF rats during a drug wash-out period. Our studies show for the first time that DM199 administration results in acute anti-hyperglycemic effects in several preclinical models, and demonstrate the potential for further development of DM199 as a novel therapeutic for T2D. PMID:25100328

  8. Crystal structures of human tissue kallikrein 4: activity modulation by a specific zinc binding site.

    PubMed

    Debela, Mekdes; Magdolen, Viktor; Grimminger, Valerie; Sommerhoff, Christian; Messerschmidt, Albrecht; Huber, Robert; Friedrich, Rainer; Bode, Wolfram; Goettig, Peter

    2006-10-01

    Human tissue kallikrein 4 (hK4) belongs to a 15-member family of closely related serine proteinases. hK4 is predominantly expressed in prostate, activates hK3/PSA, and is up-regulated in prostate and ovarian cancer. We have identified active monomers of recombinant hK4 besides inactive oligomers in solution. hK4 crystallised in the presence of zinc, nickel, and cobalt ions in three crystal forms containing cyclic tetramers and octamers. These structures display a novel metal site between His25 and Glu77 that links the 70-80 loop with the N-terminal segment. Micromolar zinc as present in prostatic fluid inhibits the enzymatic activity of hK4 against fluorogenic substrates. In our measurements, wild-type hK4 exhibited a zinc inhibition constant (IC50) of 16 microM including a permanent residual activity, in contrast to the zinc-independent mutants H25A and E77A. Since the Ile16 N terminus of wild-type hK4 becomes more accessible for acetylating agents in the presence of zinc, we propose that zinc affects the hK4 active site via the salt-bridge formed between the N terminus and Asp194 required for a functional active site. hK4 possesses an unusual 99-loop that creates a groove-like acidic S2 subsite. These findings explain the observed specificity of hK4 for the P1 to P4 substrate residues. Moreover, hK4 shows a negatively charged surface patch, which may represent an exosite for prime-side substrate recognition. PMID:16950394

  9. Human Tissue Kallikrein Activity in Angiographically Documented Chronic Stable Coronary Artery Disease

    PubMed Central

    Figueiredo, Estêvão Lanna; Magalhães, Carolina Antunes; Belli, Karlyse Claudino; Mandil, Ari; Garcia, José Carlos Faria; Araújo, Rosanã Aparecida; Figueiredo, Amintas Fabiano de Souza; Pellanda, Lucia Campos

    2015-01-01

    Background Human tissue kallikrein (hK1) is a key enzyme in the kallikrein–kinin system (KKS). hK1-specific amidase activity is reduced in urine samples from hypertensive and heart failure (HF) patients. The pathophysiologic role of hK1 in coronary artery disease (CAD) remains unclear. Objective To evaluate hK1-specific amidase activity in the urine of CAD patients Methods Sixty-five individuals (18–75 years) who underwent cardiac catheterism (CATH) were included. Random midstream urine samples were collected immediately before CATH. Patients were classified in two groups according to the presence of coronary lesions: CAD (43 patients) and non-CAD (22 patients). hK1 amidase activity was estimated using the chromogenic substrate D-Val-Leu-Arg-Nan. Creatinine was determined using Jaffé’s method. Urinary hK1-specific amidase activity was expressed as µM/(min · mg creatinine) to correct for differences in urine flow rates. Results Urinary hK1-specific amidase activity levels were similar between CAD [0.146 µM/(min ·mg creatinine)] and non-CAD [0.189 µM/(min . mg creatinine)] patients (p = 0.803) and remained similar to values previously reported for hypertensive patients [0.210 µM/(min . mg creatinine)] and HF patients [0.104 µM/(min . mg creatinine)]. CAD severity and hypertension were not observed to significantly affect urinary hK1-specific amidase activity. Conclusion CAD patients had low levels of urinary hK1-specific amidase activity, suggesting that renal KKS activity may be reduced in patients with this disease. PMID:26351984

  10. Pharmacological effects of recombinant human tissue kallikrein on bradykinin B2 receptors

    PubMed Central

    Charest-Morin, Xavier; Raghavan, Arvind; Charles, Matthew L; Kolodka, Tadeusz; Bouthillier, Johanne; Jean, Mélissa; Robbins, Mark S; Marceau, François

    2015-01-01

    Tissue kallikrein (KLK-1), a serine protease, initiates the release of bradykinin (BK)-related peptides from low-molecular weight kininogen. KLK-1 and the BK B2 receptor (B2R) mediate beneficial effects on the progression of type 2 diabetes and renal disease, but the precise role of KLK-1 independent of its kinin-forming activity remains unclear. We used DM199, a recombinant form of human KLK-1, along with the isolated human umbilical vein, a robust bioassay of the B2R, to address the previous claims that KLK-1 directly binds to and activates the human B2R, with possible receptor cleavage. DM199 (1–10 nmol/L) contracted the isolated vein via the B2R, but in a tachyphylactic, kinin-dependent manner, without desensitization of the tissue to exogenously added BK. In binding experiments with recombinant N-terminally tagged myc-B2Rs expressed in HEK 293a cells, DM199 displaced [3H]BK binding from the rabbit myc-B2R, but not from the human or rat myc-B2Rs. No evidence of myc-B2R degradation by immunoblot analysis was apparent following treatment of these 3 myc-B2R constructs with DM199 (30 min, ≤10 nmol/L). In HEK 293 cells stably expressing rabbit B2R-GFP, DM199 (11–108 pmol/L) elicited signaling-dependent endocytosis and reexpression, while a higher concentration (1.1 nmol/L) induced a partially irreversible endocytosis of the construct (microscopy), paralleled by the appearance of free GFP in cells (immunoblotting, indicative of incomplete receptor down-regulation). The pharmacology of DM199 at relevant concentrations (<10 nmol/L) is essentially based on the activity of locally generated kinins. Binding to and mild down-regulation of the B2R is possibly a species-dependent idiosyncratic response to DM199. PMID:26038695

  11. Pharmacological effects of recombinant human tissue kallikrein on bradykinin B2 receptors.

    PubMed

    Charest-Morin, Xavier; Raghavan, Arvind; Charles, Matthew L; Kolodka, Tadeusz; Bouthillier, Johanne; Jean, Mélissa; Robbins, Mark S; Marceau, François

    2015-03-01

    Tissue kallikrein (KLK-1), a serine protease, initiates the release of bradykinin (BK)-related peptides from low-molecular weight kininogen. KLK-1 and the BK B2 receptor (B2R) mediate beneficial effects on the progression of type 2 diabetes and renal disease, but the precise role of KLK-1 independent of its kinin-forming activity remains unclear. We used DM199, a recombinant form of human KLK-1, along with the isolated human umbilical vein, a robust bioassay of the B2R, to address the previous claims that KLK-1 directly binds to and activates the human B2R, with possible receptor cleavage. DM199 (1-10 nmol/L) contracted the isolated vein via the B2R, but in a tachyphylactic, kinin-dependent manner, without desensitization of the tissue to exogenously added BK. In binding experiments with recombinant N-terminally tagged myc-B2Rs expressed in HEK 293a cells, DM199 displaced [(3)H]BK binding from the rabbit myc-B2R, but not from the human or rat myc-B2Rs. No evidence of myc-B2R degradation by immunoblot analysis was apparent following treatment of these 3 myc-B2R constructs with DM199 (30 min, ≤10 nmol/L). In HEK 293 cells stably expressing rabbit B2R-GFP, DM199 (11-108 pmol/L) elicited signaling-dependent endocytosis and reexpression, while a higher concentration (1.1 nmol/L) induced a partially irreversible endocytosis of the construct (microscopy), paralleled by the appearance of free GFP in cells (immunoblotting, indicative of incomplete receptor down-regulation). The pharmacology of DM199 at relevant concentrations (<10 nmol/L) is essentially based on the activity of locally generated kinins. Binding to and mild down-regulation of the B2R is possibly a species-dependent idiosyncratic response to DM199.

  12. Evolutionary History of Tissue Kallikreins

    PubMed Central

    Pavlopoulou, Athanasia; Pampalakis, Georgios; Michalopoulos, Ioannis; Sotiropoulou, Georgia

    2010-01-01

    The gene family of human kallikrein-related peptidases (KLKs) encodes proteins with diverse and pleiotropic functions in normal physiology as well as in disease states. Currently, the most widely known KLK is KLK3 or prostate-specific antigen (PSA) that has applications in clinical diagnosis and monitoring of prostate cancer. The KLK gene family encompasses the largest contiguous cluster of serine proteases in humans which is not interrupted by non-KLK genes. This exceptional and unique characteristic of KLKs makes them ideal for evolutionary studies aiming to infer the direction and timing of gene duplication events. Previous studies on the evolution of KLKs were restricted to mammals and the emergence of KLKs was suggested about 150 million years ago (mya). In order to elucidate the evolutionary history of KLKs, we performed comprehensive phylogenetic analyses of KLK homologous proteins in multiple genomes including those that have been completed recently. Interestingly, we were able to identify novel reptilian, avian and amphibian KLK members which allowed us to trace the emergence of KLKs 330 mya. We suggest that a series of duplication and mutation events gave rise to the KLK gene family. The prominent feature of the KLK family is that it consists of tandemly and uninterruptedly arrayed genes in all species under investigation. The chromosomal co-localization in a single cluster distinguishes KLKs from trypsin and other trypsin-like proteases which are spread in different genetic loci. All the defining features of the KLKs were further found to be conserved in the novel KLK protein sequences. The study of this unique family will further assist in selecting new model organisms for functional studies of proteolytic pathways involving KLKs. PMID:21072173

  13. Inhibitors of Kallikrein in Human Plasma

    PubMed Central

    McConnell, David J.

    1972-01-01

    Human plasma was fractionated by ammonium sulfate precipitation, DEAE-cellulose chromatography, and Sephadex G-200 gel filtration to determine which method would give the greatest number of clearly separable kallikrein inhibitory peaks. With G-200 gel filtration three peaks could be separated which were demonstrated to contain α2-macroglobulin, C1̄ inactivator, and α1-antitrypsin. No other kallikrein inhibitors could be identified. The fractions containing C1̄ inactivator and α2-macroglobulin appeared to be more effective against kallikrein than that containing α1-antitrypsin. A patient with hereditary angioneurotic edema was shown to have an abnormal C1̄ inactivator protein capable of interfering with kallikrein's biologic, but not its esterolytic activity. Heat-treated human plasma, a commonly used source of kininogen for experiments with kallikrein, was shown to have kallikrein inhibitory activity. PMID:4113391

  14. Tissue kallikrein in cardiovascular, cerebrovascular and renal diseases and skin wound healing.

    PubMed

    Chao, Julie; Shen, Bo; Gao, Lin; Xia, Chun-Fang; Bledsoe, Grant; Chao, Lee

    2010-04-01

    Tissue kallikrein (KLK1) processes low-molecular weight kininogen to produce vasoactive kinins, which exert biological functions via kinin receptor signaling. Using various delivery approaches, we have demonstrated that tissue kallikrein through kinin B2 receptor signaling exhibits a wide spectrum of beneficial effects by reducing cardiac and renal injuries, restenosis and ischemic stroke, and by promoting angiogenesis and skin wound healing, independent of blood pressure reduction. Protection by tissue kallikrein in oxidative organ damage is attributed to the inhibition of apoptosis, inflammation, hypertrophy and fibrosis. Tissue kallikrein also enhances neovascularization in ischemic heart and limb. Moreover, tissue kallikrein/kinin infusion not only prevents but also reverses kidney injury, inflammation and fibrosis in salt-induced hypertensive rats. Furthermore, there is a wide time window for kallikrein administration in protection against ischemic brain infarction, as delayed kallikrein infusion for 24 h after cerebral ischemia in rats is effective in reducing neurological deficits, infarct size, apoptosis and inflammation. Importantly, in the clinical setting, human tissue kallikrein has been proven to be effective in the treatment of patients with acute brain infarction when injected within 48 h after stroke onset. Finally, kallikrein promotes skin wound healing and keratinocyte migration by direct activation of protease-activated receptor 1. PMID:20180644

  15. Angiogenesis in cervical cancer is mediated by HeLa metabolites through endothelial cell tissue kallikrein.

    PubMed

    Naidoo, Strinivasen; Raidoo, Deshandra Munsamy

    2009-08-01

    High vascularity correlates with poor clinical outcome in cancer of the uterine cervix. We investigated whether human cervical cancer cell (HeLa) metabolites influenced endothelial cell proliferation through the serine protease, tissue kallikrein. The angiogenic potential of tissue kallikrein is proposed due to its proteolytic, mitogenic and invasive properties. Under pre-defined conditions, we examined the regulation of tissue kallikrein simultaneously in both endothelial and HeLa cells using immunochemistry, ELISA, cell proliferation assays and in situ RT-PCR. In an endothelial-cervical carcinoma conditioned-medium model, HeLa metabolites caused a dramatic decrease in endothelial cellular tissue kallikrein and a concomitant proliferation of endothelial cells. ELISA on the conditioned media showed a dose-dependent increase of tissue kallikrein, while in situ RT-PCR demonstrated no change in tissue kallikrein mRNA in both endothelial and HeLa cells when challenged with each other's metabolites. This demonstration of the ability of cervical cancer to simultaneously manipulate both tissue kallikrein processing within endothelial cells and angiogenesis is novel. Should this occur in vivo, the tissue kallikrein released from the endothelial cells into the microenvironment may simultaneously degrade the matrix and elicit a mitogenic effect by promoting angiogenesis. Pre-treatment with TK inhibitors and/or anti-angiogenic therapies may prove to benefit future cervical cancer patients. PMID:19578768

  16. Structure of plasma and tissue kallikreins.

    PubMed

    Pathak, M; Wong, S S; Dreveny, I; Emsley, J

    2013-09-01

    The kallikrein kinin system (KKS) consists of serine proteases involved in the production of peptides called kinins, principally bradykinin and Lys-bradykinin (kallidin). The KKS contributes to a variety of physiological processes including inflammation, blood pressure control and coagulation. Here we review the protein structural data available for these serine proteases and examine the molecular mechanisms of zymogen activation and substrate recognition focusing on plasma kallikrein (PK) and tissue kallikrein (KLK1) cleavage of kininogens. PK circulates as a zymogen bound to high-molecular-weight kininogen (HK). PK is activated by coagulation factor XIIa and then cleaves HK to generate bradykinin and factor XII to generate further XIIa.A structure has been described for the activated PK protease domain in complex with the inhibitor benzamidine. Kallikrein-related peptidases (KLKs) have a distinct domain structure and exist as a family of 15 genes which are differentially expressed in many tissues and the central nervous system.They cleave a wide variety of substrates including low-molecular-weight kininogen (LK) and matrix proteins. Crystal structures are available for KLK1, 3, 4, 5, 6 and 7 activated protease domains typically in complex with S1 pocket inhibitors. A substrate mimetic complex is described for KLK3 which provides insight into substrate recognition. A zymogen crystal structure determined for KLK6 reveals a closed S1 pocket and a novel mechanism of zymogen activation. Overall these structures have proved highly informative in understanding the molecular mechanisms of the KKS and provide templates to design inhibitors for treatment of a variety of diseases. PMID:23494059

  17. Tissue kallikrein-kinin therapy in hypertension and organ damage.

    PubMed

    Chao, Julie; Bledsoe, Grant; Chao, Lee

    2014-01-01

    Tissue kallikrein is a serine proteinase that cleaves low molecular weight kininogen to produce kinin peptides, which in turn activate kinin receptors to trigger multiple biological functions. In addition to its kinin-releasing activity, tissue kallikrein directly interacts with the kinin B2 receptor, protease-activated receptor-1, and gamma-epithelial Na channel. The tissue kallikrein-kinin system (KKS) elicits a wide spectrum of biological activities, including reducing hypertension, cardiac and renal damage, restenosis, ischemic stroke, and skin wound injury. Both loss-of-function and gain-of-function studies have shown that the KKS plays an important endogenous role in the protection against health pathologies. Tissue kallikrein/kinin treatment attenuates cardiovascular, renal, and brain injury by inhibiting oxidative stress, apoptosis, inflammation, hypertrophy, and fibrosis and promoting angiogenesis and neurogenesis. Approaches that augment tissue kallikrein-kinin activity might provide an effective strategy for the treatment of hypertension and associated organ damage. PMID:25130039

  18. Unleashing the therapeutic potential of human kallikrein-related serine proteases.

    PubMed

    Prassas, Ioannis; Eissa, Azza; Poda, Gennadiy; Diamandis, Eleftherios P

    2015-03-01

    Tissue kallikreins are a family of fifteen secreted serine proteases encoded by the largest protease gene cluster in the human genome. In the past decade, substantial progress has been made in characterizing the natural substrates, endogenous inhibitors and in vivo functions of kallikreins, and studies have delineated important pathophysiological roles for these proteases in a variety of tissues. Thus, kallikreins are now considered attractive targets for the development of novel therapeutics for airway, cardiovascular, tooth, brain, skin and neoplastic diseases. In this Review, we discuss recent advances in our understanding of the physiological functions and pathological implications of kallikrein proteases, and highlight progress in the identification of kallikrein inhibitors, which together are bringing us closer to therapeutically targeting kallikreins in selected disease settings.

  19. Human kallikrein 3 (prostate specific antigen) and human kallikrein 5 expression in salivary gland tumors.

    PubMed

    Darling, M R; Tsai, S; Jackson-Boeters, L; Daley, T D; Diamandis, E P

    2006-01-01

    The human kallikrein 5 protein (hK5) is expressed in many normal tissues, most notably in skin, breast, salivary gland and esophagus. It has also been shown to be a potential biomarker for breast, ovarian and testicular cancer. Human kallikrein 3 (hK3; prostate-specific antigen) is the most useful marker for adenocarcinoma of the prostate gland. The aim of this study was to determine whether hK3 and hK5 are expressed in salivary gland tissues and salivary gland tumors (both benign and malignant), in order to compare normal with tumor tissues. Pleomorphic adenomas, adenoid cystic carcinomas, polymorphous low-grade adenocarcinomas, acinic cell carcinomas, mucoepidermoid carcinomas and adenocarcinomas not otherwise specified of both minor and major salivary glands were examined. The results of this study indicate that most salivary gland tumors do not show high levels of expression of hK5. Staining was most prominent in keratinizing epithelia in pleomorphic adenomas. hK3 is not expressed in salivary gland tumors.

  20. Purified human plasma kallikrein aggregates human blood neutrophils.

    PubMed Central

    Schapira, M; Despland, E; Scott, C F; Boxer, L A; Colman, R W

    1982-01-01

    Exposure of human blood polymorphonuclear leukocytes (PMN) to purified active plasma kallikrein resulted in PMN aggregation when kallikrein was present at concentrations ranging from 0.4 to 0.6 U/ml (0.18-0.27 microM). Kallikrein-induced PMN aggregation was not mediated through C5-derived peptides, because identical responses were observed whether or not kallikrein had been preincubated with an antibody to C5. Moreover, kallikrein was specific for aggregating PMN, because no aggregation was observed with Factor XII active fragments (23 nM), Factor XIa (0.6 U/ml or 15nM), thrombin (1.6 microM), plasmin (2 microM), porcine pancreatic elastase (2 microM), bovine pancreatic chymotrypsin (2 microM), or bradykinin (1 microM). Bovine pancreatic trypsin (2 microM) aggregated PMN, but to a lesser extent than kallikrein (0.18 microM). Kallikrein was a potent aggregant agent for PMN because similar responses were observed with kallikrein (0.5 U/ml or 0.23 microM) and an optimal dose (0.2 microM) of N-formyl-methionyl-leucyl-phenylalanine. In addition, PMN incubation with kallikrein resulted in stimulation of their oxidative metabolism as assessed by an increased oxygen uptake. Neutropenia and leukostasis observed in diseases associated with activation of the contact phase system may be the result of PMN aggregation by plasma kallikrein. PMID:6917855

  1. Cardiovascular abnormalities with normal blood pressure in tissue kallikrein-deficient mice

    NASA Astrophysics Data System (ADS)

    Meneton, Pierre; Bloch-Faure, May; Hagege, Albert A.; Ruetten, Hartmut; Huang, Wei; Bergaya, Sonia; Ceiler, Debbie; Gehring, Doris; Martins, Isabelle; Salmon, Georges; Boulanger, Chantal M.; Nussberger, Jürg; Crozatier, Bertrand; Gasc, Jean-Marie; Heudes, Didier; Bruneval, Patrick; Doetschman, Tom; Ménard, Joël; Alhenc-Gelas, François

    2001-02-01

    Tissue kallikrein is a serine protease thought to be involved in the generation of bioactive peptide kinins in many organs like the kidneys, colon, salivary glands, pancreas, and blood vessels. Low renal synthesis and urinary excretion of tissue kallikrein have been repeatedly linked to hypertension in animals and humans, but the exact role of the protease in cardiovascular function has not been established largely because of the lack of specific inhibitors. This study demonstrates that mice lacking tissue kallikrein are unable to generate significant levels of kinins in most tissues and develop cardiovascular abnormalities early in adulthood despite normal blood pressure. The heart exhibits septum and posterior wall thinning and a tendency to dilatation resulting in reduced left ventricular mass. Cardiac function estimated in vivo and in vitro is decreased both under basal conditions and in response to βadrenergic stimulation. Furthermore, flow-induced vasodilatation is impaired in isolated perfused carotid arteries, which express, like the heart, low levels of the protease. These data show that tissue kallikrein is the main kinin-generating enzyme in vivo and that a functional kallikrein-kinin system is necessary for normal cardiac and arterial function in the mouse. They suggest that the kallikrein-kinin system could be involved in the development or progression of cardiovascular diseases.

  2. A synovial amidase acting on tissue kallikrein-selective substrate in clinical and experimental arthritis.

    PubMed

    Al-Haboubi, H A; Bennett, D; Sharma, J N; Thomas, G R; Zeitlin, I J

    1986-01-01

    Increased levels of amidase acting on a tissue-kallikrein selective substrate, Val.Leu.Arg.pNA, with an activity optimum at pH9, were detected in blood-free inflamed tissues from adjuvant arthritic rats (p less than 0.01). The component of this activity resistant to inhibition by soybean trypsin inhibitor (SBTI) also greatly increased (p less than 0.05). Both the SBTI-sensitive and SBTI-resistant components were inhibited by aprotinin (93% and 72% respectively). Kallikrein-like amidase also increased in inflamed synovia from seropositive rheumatoid, and osteoarthritic dogs when compared with healthy canine synovia. This increase was parallelled by an increase in kinin-forming enzyme which was also measured in rheumatoid and healthy animals and this activity was inhibited 72% by aprotinin. Total kallikrein-like amidase also increased 989% (p less than 0.05) in synovia from seropositive rheumatoid human patients, compared with healthy synovial tissue. Evidence is presented indicating that the origin of this enzymic activity may be plasma kallikrein. PMID:3643734

  3. Human kallikrein 8 expression in salivary gland tumors.

    PubMed

    Darling, Mark R; Tsai, Sam; Jackson-Boeters, Linda; Daley, Thomas D; Diamandis, Eleftherios P

    2008-09-01

    The human kallikrein 8 protein (KLK8) is expressed in many normal tissues including esophagus, skin, testis, tonsil, kidney, breast, and salivary gland, and is found in biological fluids including breast milk, amniotic fluid, seminal fluid and serum. It has also been shown to be a biomarker and prognostic factor for breast cancer. The aim of this study was to determine whether KLK8 is expressed in salivary gland tissues and salivary gland tumors (both benign and malignant), in order to compare normal with tumor tissues. Pleomorphic adenomas, adenoid cystic carcinomas, polymorphous low grade adenocarcinomas, acinic cell carcinomas, mucoepidermoid carcinomas, and adenocarcinomas NOS of both minor and major salivary glands were examined. The results of this study indicate that most salivary gland tumors show high levels of expression of KLK8.

  4. Linear competitive inhibition of human tissue kallikrein by 4-aminobenzamidine and benzamidine and linear mixed inhibition by 4-nitroaniline and aniline.

    PubMed

    Sousa, M O; Miranda, T L; Costa, E B; Bittar, E R; Santoro, M M; Figueiredo, A F

    2001-01-01

    Hydrolysis of D-valyl-L-leucyl-L-arginine p-nitroanilide (7.5-90.0 microM) by human tissue kallikrein (hK1) (4.58-5.27 nM) at pH 9.0 and 37 degrees C was studied in the absence and in the presence of increasing concentrations of 4-aminobenzamidine (96-576 microM), benzamidine (1.27-7.62 mM), 4-nitroaniline (16.5-66 microM) and aniline (20-50 mM). The kinetic parameters determined in the absence of inhibitors were: Km = 12.0 +/- 0.8 microM and k cat = 48.4 +/- 1.0 min(-1). The data indicate that the inhibition of hK1 by 4-aminobenzamidine and benzamidine is linear competitive, while the inhibition by 4-nitroaniline and aniline is linear mixed, with the inhibitor being able to bind both to the free enzyme with a dissociation constant Ki yielding an EI complex, and to the ES complex with a dissociation constant Ki', yielding an ESI complex. The calculated Ki values for 4-aminobenzamidine, benzamidine, 4-nitroaniline and aniline were 146 +/- 10, 1,098 +/- 91, 38.6 +/- 5.2 and 37,340 +/- 5,400 microM, respectively. The calculated Ki' values for 4-nitroaniline and aniline were 289.3 +/- 92.8 and 310,500 +/- 38,600 microM, respectively. The fact that Ki'>Ki indicates that 4-nitroaniline and aniline bind to a second binding site in the enzyme with lower affinity than they bind to the active site. The data about the inhibition of hK1 by 4-aminobenzamidine and benzamidine help to explain previous observations that esters, anilides or chloromethyl ketone derivatives of Nalpha-substituted arginine are more sensitive substrates or inhibitors of hK1 than the corresponding lysine compounds. PMID:11151026

  5. Human plasma kallikrein releases neutrophil elastase during blood coagulation.

    PubMed Central

    Wachtfogel, Y T; Kucich, U; James, H L; Scott, C F; Schapira, M; Zimmerman, M; Cohen, A B; Colman, R W

    1983-01-01

    Elastase is released from human neutrophils during the early events of blood coagulation. Human plasma kallikrein has been shown to stimulate neutrophil chemotaxis, aggregation, and oxygen consumption. Therefore, the ability of kallikrein to release neutrophil elastase was investigated. Neutrophils were isolated by dextran sedimentation, and elastase release was measured by both an enzyme-linked immunosorbent assay, and an enzymatic assay using t-butoxy-carbonyl-Ala-Ala-Pro-Val-amino methyl coumarin as the substrate. Kallikrein, 0.1-1.0 U/ml, (0.045-0.45 microM), was incubated with neutrophils that were preincubated with cytochalasin B (5 micrograms/ml). The release of elastase was found to be proportional to the kallikrein concentration. Kallikrein released a maximum of 34% of the total elastase content, as measured by solubilizing the neutrophils in the nonionic detergent Triton X-100. A series of experiments was carried out to determine if kallikrein was a major enzyme involved in neutrophil elastase release during blood coagulation. When 10 million neutrophils were incubated in 1 ml of normal plasma in the presence of 30 mM CaCl2 for 90 min, 2.75 micrograms of elastase was released. In contrast, neutrophils incubated in prekallikrein-deficient or Factor XII-deficient plasma released less than half of the elastase, as compared with normal plasma. The addition of purified prekallikrein to prekallikrein-deficient plasma restored neutrophil elastase release to normal levels. Moreover, release of elastase was enhanced in plasma deficient in C1-inhibitor, the major plasma inhibitor of kallikrein. This release was not dependent upon further steps in the coagulation pathway, or on C5a, since levels of elastase, released in Factor XI- or C5-deficient plasma, were similar to that in normal plasma, and an antibody to C5 failed to inhibit elastase release. These data suggest that kallikrein may be a major enzyme responsible for the release of elastase during blood

  6. Human kallikrein 13 expression in salivary gland tumors.

    PubMed

    Darling, M R; Jackson-Boeters, L; Daley, T D; Diamandis, E P

    2006-01-01

    The human kallikrein 13 protein (hK13) is expressed in many normal tissues. Petraki et al have previously described presence of hK13 in salivary gland tissue, localized to duct epithelia and some acinar cells. The aim of this study was to determine whether hK13 is expressed in salivary gland tissues and salivary gland tumors (both benign and malignant), in order to compare normal with tumor tissues. Pleomorphic adenomas (PA), adenoid cystic carcinomas (ACC), polymorphous low grade adenocarcinomas (PLGA), acinic cell carcinomas (ACI), mucoepidermoid carcinomas (MEC) and adenocarcinomas not otherwise specified (ANOS) of both minor and major salivary glands were examined. The results of this study indicate that most salivary gland tumors show high levels of expression of hK13. Overall, staining in PA was significantly less than that seen in normal salivary gland tissue. PLGA, ACC and ANOS each stained significantly more than normal salivary gland tissue while MEC and ACI did not. Ductal cells and cells lining duct-like structures showed a higher intensity of staining than non-ductal cells in most tumors. Tumors which exhibited only non-ductal cells also exhibited cytoplasmic staining. In conclusion, we demonstrate the high expression of hK13 in several common salivary gland tumors.

  7. Crystallization and preliminary crystallographic studies of human kallikrein 7, a serine protease of the multigene kallikrein family

    SciTech Connect

    Fernández, Israel S.; Ständker, Ludger; Forssmann, Wolf-Georg; Giménez-Gallego, Guillermo; Romero, Antonio

    2007-08-01

    The cloning, expression, purification and crystallization of recombinant human kallikrein 7, directly synthesized in the active form in E. coli, is described. Diffraction data were collected to 2.8 Å resolution from native crystals. Human kallikreins are a group of serine proteases of high sequence homology whose genes are grouped as a single cluster at chromosome 19. Although the physiological roles of kallikreins are generally still unknown, members of the kallikrein family have been clearly implicated in pathological situations such as cancer and psoriasis. Human kallikrein 7 (hK7) has been shown to be involved in pathological keratinization, psoriasis and ovarian cancer. In order to gain insight into the molecular structure of this protein, hK7 was crystallized after recombinant production in its folded and active form using a periplasmic secretion vector in Escherichia coli. The crystals belonged to the rhombohedral space group H32 and diffracted to 2.8 Å. The phase problem was solved by molecular replacement using the mouse kallikrein-related protein neuropsin. Completion of the model and structure refinement are under way.

  8. Human kallikrein 6 expression in salivary gland tumors.

    PubMed

    Darling, M R; Jackson-Boeters, L; Daley, T D; Diamandis, E P

    2006-03-01

    Human kallikrein 6 (hK6), also known as zyme/protease M/neurosin), is expressed in many normal glandular tissues. The aim of this study was to determine whether hK6 is expressed in salivary gland tissues and salivary gland tumors (both benign and malignant), using an immunohistochemical method. Pleomorphic adenomas (PA), adenoid cystic carcinomas, polymorphous low-grade adenocarcinomas, acinic cell carcinomas, mucoepidermoid carcinomas, and adenocarcinomas not otherwise specified of both minor and major salivary glands were examined. Cells lining duct-like structures and non-duct-like cells were scored. Only in PA of minor salivary gland origin was overall staining higher in duct-like than in non-duct-like cells. In all other tumors exhibiting both types of cells, hK6 staining was similar in both duct-like and non-duct-like cells. Tumors that exhibited non-duct-like cells only also exhibited cytoplasmic staining. Results of this study show that salivary gland tumors express hK6, apparently downregulated in comparison with normal salivary gland tissue, and that this expression is not specific for any of the tumors studied.

  9. Specificity of S'1 and S'2 subsites of human tissue kallikrein using the reactive-centre loop of kallistatin: the importance of P'1 and P'2 positions in design of inhibitors.

    PubMed

    Pimenta, Daniel C; Fogaça, Sandro E; Melo, Robson L; Juliano, Luiz; Juliano, Maria A

    2003-05-01

    We have demonstrated that the S'(1) and S'(2) subsites of human tissue kallikrein (hK1) play determinant roles in the recognition and hydrolysis of substrates. The presence of serine at position P'(1) and arginine at P'(2) resulted in the best substrate, Abz-Ala-Ile-Lys-Phe-Phe-Ser-Arg-Gln-EDDnp, which was derived from the kallistatin reactive-centre loop sequence and quencher groups o-aminobenzoic acid (Abz) and N-(2,4-dinitrophenyl)ethylenediamine (EDDnp). Serine and arginine are also the residues at positions P'(1) and P'(2) in human kininogen, from which hK1 releases Lys-bradykinin. Several peptide analogues of Abz-Ala-Ile-Lys-Phe-Phe-Ser-Arg-Gln-EDDnp, in which the Ser and Arg residues were substituted with various other amino acids, were synthesized and tested as substrates. Most of them were hydrolysed slowly, although they showed significant binding to hK1, as demonstrated by their competitive inhibition constants (K(i)). Using this information, six peptides were designed, synthesized and assayed as inhibitors of hK1. Abz-Lys-Phe-Phe-Pro-Arg-Gln-EDDnp, Abz-Lys-Phe-Arg-Pro-Arg-Gln-EDDnp and acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH(2) inhibited hK1 in the range 20-30 nM (letters in italics denote the D-form of the amino acid). The peptide acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH(2) was a weak inhibitor for other serine proteases, as indicated by the higher K (i) values compared with hK1, but this peptide was a potent inhibitor of human plasma kallikrein, which has a K (i) value of 8 nM. This result was surprising, since this enzyme is known to be a restricted arginyl-hydrolase. In conclusion, acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH(2) can be used as a leader compound to design specific inhibitors for hK1, plasma kallikrein, or for both at same time, if the inhibition of kinin release is the main goal. PMID:12578561

  10. [Isolation and properties of 2 kallikreins from human saliva].

    PubMed

    Vovchuk, S V; Barabash, R D; Levitsky, A P

    1975-01-01

    Two kallikreins (K-I and K-II) were purified from mixed, parotid and submandibular human saliva. The kallikreins were separated by chromatography on CM-cellulose. The pH optima of activity were at pH 9.3 and pH 9.6-9.8; Km for BAEE was 1.10-3M and 4.10-3M, respectively. The esterase activity of K-I and K-II was inhibited by trasylol-like inhibitors, while the plant and synthetic inhibitors of trypsin were uneffective. In dog, rabbit and rat the hypotensive effect of K-II and its action upon the permeability of rabbit skin was 4-5 fold higher than the effects of K-I. Ki and K-II did not alter the arterial blood pressure in guinea pig. PMID:3025

  11. Tissue kallikrein permits early renal adaptation to potassium load.

    PubMed

    El Moghrabi, Soumaya; Houillier, Pascal; Picard, Nicolas; Sohet, Fabien; Wootla, Bharath; Bloch-Faure, May; Leviel, Françoise; Cheval, Lydie; Frische, Sebastian; Meneton, Pierre; Eladari, Dominique; Chambrey, Régine

    2010-07-27

    Tissue kallikrein (TK) is a serine protease synthetized in renal tubular cells located upstream from the collecting duct where renal potassium balance is regulated. Because secretion of TK is promoted by K+ intake, we hypothesized that this enzyme might regulate plasma K+ concentration ([K+]). We showed in wild-type mice that renal K+ and TK excretion increase in parallel after a single meal, representing an acute K+ load, whereas aldosterone secretion is not modified. Using aldosterone synthase-deficient mice, we confirmed that the control of TK secretion is aldosterone-independent. Mice with TK gene disruption (TK-/-) were used to assess the impact of the enzyme on plasma [K+]. A single large feeding did not lead to any significant change in plasma [K+] in TK+/+, whereas TK-/- mice became hyperkalemic. We next examined the impact of TK disruption on K+ transport in isolated cortical collecting ducts (CCDs) microperfused in vitro. We found that CCDs isolated from TK-/- mice exhibit net transepithelial K+ absorption because of abnormal activation of the colonic H+,K+-ATPase in the intercalated cells. Finally, in CCDs isolated from TK-/- mice and microperfused in vitro, the addition of TK to the perfusate but not to the peritubular bath caused a 70% inhibition of H+,K+-ATPase activity. In conclusion, we have identified the serine protease TK as a unique kalliuretic factor that protects against hyperkalemia after a dietary K+ load.

  12. Tissue kallikrein-binding protein reduces blood pressure in transgenic mice.

    PubMed

    Chen, L M; Ma, J x; Liang, Y M; Chao, L; Chao, J

    1996-11-01

    The kallikrein-kinin system participates in blood pressure regulation. One of the kallikrein-kinin system components, kallikrein-binding protein, binds to tissue kallikrein and inhibits its activity in vitro. To investigate potential roles of rat kallikrein-binding protein (RKBP) in vivo, we have developed transgenic mice that express an RKBP gene under the control of the mouse metallothionein metal-responsive promoter. Expression of the transgene, RKBP, was detected in the liver, kidney, lung, heart, pancreas, salivary glands, spleen, brain, testis, and adrenal gland at the mRNA and protein levels. Systolic blood pressures of homozygous transgenic mice were 88.5 +/- 0.8 mm Hg (mean +/- S.E., n = 19, P < 0.001) for one line and 88.8 +/- 1.6 mm Hg (mean +/- S.E., n = 19, P < 0.001) for another, as compared with 100.5 +/- 0.8 mm Hg (mean +/- S.E., n = 18) for control mice. Direct blood pressure measurements of these transgenic mice through an arterial cannula showed similar reductions of blood pressure. Intravenous injection of purified RKBP into mice via a catheter produced a dose-dependent reduction of the mean arterial blood pressure. Our findings suggest that RKBP may function as a vasodilator in vivo, independent of regulating the activity of tissue kallikrein. PMID:8910346

  13. Purification and characterisation of blarinasin, a new tissue kallikrein-like protease from the short-tailed shrew Blarina brevicauda: comparative studies with blarina toxin.

    PubMed

    Kita, Masaki; Okumura, Yuushi; Ohdachi, Satoshi D; Oba, Yuichi; Yoshikuni, Michiyasu; Nakamura, Yasuo; Kido, Hiroshi; Uemura, Daisuke

    2005-02-01

    A new tissue kallikrein-like protease, blarinasin, has been purified from the salivary glands of the short-tailed shrew Blarina brevicauda. Blarinasin is a 32-kDa N-glycosylated protease with isoelectric values ranging between 5.3 and 5.7, and an optimum pH of 8.5 for enzyme activity. The cloned blarinasin cDNA coded for a pre-pro-sequence and a mature peptide of 252 amino acids with a catalytic triad typical for serine proteases and 43.7-54.0% identity to other mammalian tissue kallikreins. Blarinasin preferentially hydrolysed Pro-Phe-Arg-4-methylcoumaryl-7-amide (MCA) and N-tert-butyloxycarbonyl-Val-Leu-Lys-MCA, and preferentially converted human high-molecular-weight kininogen (HK) to bradykinin. The activity of blarinasin was prominently inhibited by aprotinin (K(i) =3.4 nM). A similar kallikrein-like protease, the lethal venom blarina toxin, has previously been purified from the salivary glands of the shrew Blarina and shows 67.9% identity to blarinasin. However, blarinasin was not toxic in mice. Blarinasin is a very abundant kallikrein-like protease and represents 70-75% of kallikrein-like enzymes in the salivary gland of B. brevicauda. PMID:15843162

  14. Putative functions of tissue kallikrein-related peptidases in vaginal fluid.

    PubMed

    Muytjens, Carla M J; Vasiliou, Stella K; Oikonomopoulou, Katerina; Prassas, Ioannis; Diamandis, Eleftherios P

    2016-10-01

    Cervical-vaginal fluid (CVF) is a complex biological fluid that hydrates the mucosa of the lower female reproductive system. In-depth proteomic and biochemical studies on CVF have revealed that it contains large amounts of endogenous proteases and protease inhibitors, including an abundance of several members of the tissue kallikrein-related peptidase (KLK) family. Despite their ubiquitous presence in human tissues and fluids, KLK expression levels vary considerably, with maximum expression observed in reproduction-related tissues and fluids. The roles of KLKs in the lower female reproductive system are not fully understood. The activation of KLKs in CVF is dependent on pH and various modes of KLK regulation in the vagina exist. KLKs have been postulated to have roles in physiological functions related to antimicrobial processes, vaginal and cervical epithelial desquamation, sperm transport, and the processing of fetal membranes as observed in preterm premature rupture of membranes. Increased understanding of the functional roles of KLKs in the lower female reproductive system could lead to new diagnostic and therapeutic modalities for conditions such as vaginal infections and vaginal atrophy.

  15. Putative functions of tissue kallikrein-related peptidases in vaginal fluid.

    PubMed

    Muytjens, Carla M J; Vasiliou, Stella K; Oikonomopoulou, Katerina; Prassas, Ioannis; Diamandis, Eleftherios P

    2016-10-01

    Cervical-vaginal fluid (CVF) is a complex biological fluid that hydrates the mucosa of the lower female reproductive system. In-depth proteomic and biochemical studies on CVF have revealed that it contains large amounts of endogenous proteases and protease inhibitors, including an abundance of several members of the tissue kallikrein-related peptidase (KLK) family. Despite their ubiquitous presence in human tissues and fluids, KLK expression levels vary considerably, with maximum expression observed in reproduction-related tissues and fluids. The roles of KLKs in the lower female reproductive system are not fully understood. The activation of KLKs in CVF is dependent on pH and various modes of KLK regulation in the vagina exist. KLKs have been postulated to have roles in physiological functions related to antimicrobial processes, vaginal and cervical epithelial desquamation, sperm transport, and the processing of fetal membranes as observed in preterm premature rupture of membranes. Increased understanding of the functional roles of KLKs in the lower female reproductive system could lead to new diagnostic and therapeutic modalities for conditions such as vaginal infections and vaginal atrophy. PMID:27603220

  16. Tissue kallikrein mediates neurite outgrowth through epidermal growth factor receptor and flotillin-2 pathway in vitro.

    PubMed

    Lu, Zhengyu; Cui, Mei; Zhao, Hong; Wang, Tao; Shen, Yan; Dong, Qiang

    2014-02-01

    Tissue kallikrein (TK) was previously shown to take most of its biological effects through bradykinin receptors. In this study, we assumed that TK mediated neurite outgrowth was independent of bradykinin receptors. To test the hypothesis, we investigated TK-induced neurite outgrowth and its signaling mechanisms in cultured primary neurons and human SH-SY5Y cells. We found that TK stimulation could increase the number of processes and mean process length of primary neurons, which were blocked by epidermal growth factor receptor (EGFR) inhibitor or down-regulation, small interfering RNA for flotillin-2 and extracellular signal-regulated kinase (ERK) 1/2 inhibitor. Moreover, TK-induced neurite outgrowth was associated with EGFR and ERK1/2 activation, which were inhibited by EGFR antagonist or RNA interference and flotillin-2 knockdown. Interestingly, inhibition of bradykinin receptors had no significant effects on EGFR and ERK1/2 phosphorylation. In the present research, our data also suggested that EGFR and flotillin-2 formed constitutive complex that translocated to around the nuclei in the TK stimulation. In sum, our findings provided evidence that TK could promote neurite outgrowth via EGFR, flotillin-2 and ERK1/2 signaling pathway in vitro. PMID:24211626

  17. Tissue kallikrein mediates pro-inflammatory pathways and activation of protease-activated receptor-4 in proximal tubular epithelial cells.

    PubMed

    Yiu, Wai Han; Wong, Dickson W L; Chan, Loretta Y Y; Leung, Joseph C K; Chan, Kwok Wah; Lan, Hui Yao; Lai, Kar Neng; Tang, Sydney C W

    2014-01-01

    Tissue kallikrein (KLK1) expression is up-regulated in human diabetic kidney tissue and induced by high glucose (HG) in human proximal tubular epithelial cells (PTEC). Since the kallikrein-kinin system (KKS) has been linked to cellular inflammatory process in many diseases, it is likely that KLK1 expression may mediate the inflammatory process during the development of diabetic nephropathy. In this study, we explored the role of KLK1 in tubular pro-inflammatory responses under the diabetic milieu. Recombinant KLK1 stimulated the production of inflammatory cytokines in PTEC via the activation of p42/44 and p38 MAPK signaling pathways. Molecular knockdown of endogenous KLK1 expression by siRNA transfection in PTEC attenuated advanced glycation end-products (AGE)-induced IL-8 and ICAM-1 productions in vitro. Interestingly, exposure of PTEC to KLK1 induced the expression of protease-activated receptors (PARs). There was a 2.9-fold increase in PAR-4, 1.4-fold increase in PAR-1 and 1.2-fold increase in PAR-2 mRNA levels. Activation of PAR-4 by a selective agonist was found to elicit the pro-inflammatory and pro-fibrotic phenotypes in PTEC while blockade of the receptor by specific antagonist attenuated high glucose-induced IL-6, CCL-2, CTGF and collagen IV expression. Calcium mobilization by the PAR-4 agonist in PTEC was desensitized by pretreatment with KLK1. Consistent with these in vitro findings, there was a markedly up-regulation of tubular PAR-4 expression in human diabetic renal cortical tissues. Together, these results suggest that up-regulation of KLK1 in tubular epithelial cells may mediate pro-inflammatory pathway and PAR activation during diabetic nephropathy and provide a new therapeutic target for further investigation. PMID:24586431

  18. Tissue Kallikrein Mediates Pro-Inflammatory Pathways and Activation of Protease-Activated Receptor-4 in Proximal Tubular Epithelial Cells

    PubMed Central

    Yiu, Wai Han; Wong, Dickson W. L.; Chan, Loretta Y. Y.; Leung, Joseph C. K.; Chan, Kwok Wah; Lan, Hui Yao; Lai, Kar Neng; Tang, Sydney C. W.

    2014-01-01

    Tissue kallikrein (KLK1) expression is up-regulated in human diabetic kidney tissue and induced by high glucose (HG) in human proximal tubular epithelial cells (PTEC). Since the kallikrein-kinin system (KKS) has been linked to cellular inflammatory process in many diseases, it is likely that KLK1 expression may mediate the inflammatory process during the development of diabetic nephropathy. In this study, we explored the role of KLK1 in tubular pro-inflammatory responses under the diabetic milieu. Recombinant KLK1 stimulated the production of inflammatory cytokines in PTEC via the activation of p42/44 and p38 MAPK signaling pathways. Molecular knockdown of endogenous KLK1 expression by siRNA transfection in PTEC attenuated advanced glycation end-products (AGE)-induced IL-8 and ICAM-1 productions in vitro. Interestingly, exposure of PTEC to KLK1 induced the expression of protease-activated receptors (PARs). There was a 2.9-fold increase in PAR-4, 1.4-fold increase in PAR-1 and 1.2-fold increase in PAR-2 mRNA levels. Activation of PAR-4 by a selective agonist was found to elicit the pro-inflammatory and pro-fibrotic phenotypes in PTEC while blockade of the receptor by specific antagonist attenuated high glucose-induced IL-6, CCL-2, CTGF and collagen IV expression. Calcium mobilization by the PAR-4 agonist in PTEC was desensitized by pretreatment with KLK1. Consistent with these in vitro findings, there was a markedly up-regulation of tubular PAR-4 expression in human diabetic renal cortical tissues. Together, these results suggest that up-regulation of KLK1 in tubular epithelial cells may mediate pro-inflammatory pathway and PAR activation during diabetic nephropathy and provide a new therapeutic target for further investigation. PMID:24586431

  19. Human kallikrein 14: a new potential biomarker for ovarian and breast cancer.

    PubMed

    Borgoño, Carla A; Grass, Linda; Soosaipillai, Antoninus; Yousef, George M; Petraki, Constantina D; Howarth, David H C; Fracchioli, Stefano; Katsaros, Dionyssios; Diamandis, Eleftherios P

    2003-12-15

    Human kallikrein gene 14 (KLK14) is a recently discovered member of the tissue kallikrein family of secreted serine proteases, which includes hK3/prostate-specific antigen, the best cancer biomarker to date. Given that KLK14 is hormonally regulated, differentially expressed in endocrine-related cancers, and a prognostic marker for breast and ovarian cancer at the mRNA level, we hypothesize that its encoded protein, hK14, like hK3/prostate-specific antigen, may constitute a new biomarker for endocrine-related malignancies. The objective of this study was to generate immunological reagents for hK14, to develop an ELISA and immunohistochemical techniques to study its expression in normal and cancerous tissues and biological fluids. Recombinant hK14 was produced in Pichia pastoris, purified by affinity chromatography, and injected into mice and rabbits for polyclonal antibody generation. Using the mouse and rabbit antisera, a sandwich-type immunofluorometric ELISA and immunohistochemical methodologies were developed for hK14. The ELISA was sensitive (detection limit of 0.1 micro g/liter), specific for hK14, linear from 0 to 20 micro g/liter with between-run and within-run coefficients of variation of <10%. hK14 was quantified in human tissue extracts and biological fluids. Highest levels were observed in the breast, skin, prostate, seminal plasma, and amniotic fluid, with almost undetectable levels in normal serum. hK14 concentration was higher in 40% of ovarian cancer tissues compared with normal ovarian tissues. Serum hK14 levels were elevated in a proportion of patients with ovarian (65%) and breast (40%) cancers. Immunohistochemical analyses indicated strong cytoplasmic staining of hK14 by the epithelial cells of normal and malignant skin, ovary, breast, and testis. In conclusion, we report the first ELISA and immunohistochemical assays for hK14 and describe its distribution in tissues and biological fluids. Our preliminary data indicate that hK14 is a potential

  20. Selective kallikrein inhibitors alter human neutrophil elastase release during extracorporeal circulation.

    PubMed

    Wachtfogel, Y T; Hack, C E; Nuijens, J H; Kettner, C; Reilly, T M; Knabb, R M; Bischoff, R; Tschesche, H; Wenzel, H; Kucich, U

    1995-03-01

    Cardiopulmonary bypass causes hemorrhagic complications and initiates a biochemical and cellular "whole body inflammatory response." This study investigates whether a variety of selective inhibitors of the contact pathway of intrinsic coagulation modulate complement and neutrophil activation during simulated extracorporeal circulation. After 60 min of recirculation in the presence of the slow tight-binding boronic acid inhibitor, Bz-Pro-Phe-boroArg-OH (10.7 microM), complete inhibition of kallikrein-C1-inhibitor complex formation and marked inhibition of C1-C1-inhibitor complex formation and the release of human neutrophil elastase were observed. Arg15-aprotinin (3.1 microM), Ala357,Arg358 alpha 1-antitrypsin (2.6 microM), and soybean trypsin inhibitor (48.0 microM) either completely or partially inhibited the generation of kallikrein-C1-inhibitor complexes but were less effective inhibitors of human neutrophil elastase release. The second-order rate constants for the inhibition of kallikrein in purified systems are consistent with the order of effectiveness of the inhibitors in blocking human neutrophil elastase release in heparinized blood. Our results suggest that low-molecular-weight selective inhibitors of kallikrein may be effective agents in the attenuation of the contact-mediated inflammatory response in cardiopulmonary bypass.

  1. Expression and bioregulation of the kallikrein-related peptidases family in the human neutrophil.

    PubMed

    Lizama, Alejandro J; Andrade, Yessica; Colivoro, Patricio; Sarmiento, Jose; Matus, Carola E; Gonzalez, Carlos B; Bhoola, Kanti D; Ehrenfeld, Pamela; Figueroa, Carlos D

    2015-08-01

    The family of kallikrein-related peptidases (KLKs) has been identified in a variety of immunolabeled human tissue sections, but no previous study has experimentally confirmed their presence in the human neutrophil. We have investigated the expression and bioregulation of particular KLKs in the human neutrophil and, in addition, examined whether stimulation by a kinin B(1) receptor (B1R) agonist or fMet-Leu-Phe (fMLP) induces their secretion. Western blot analysis of neutrophil homogenates indicated that the MM of the KLKs ranged from 27 to 50 kDa. RT-PCR showed that blood neutrophils expressed only KLK1, KLK4, KLK10, KLK13, KLK14 and KLK15 mRNAs, whereas the non-differentiated HL-60 cells expressed most of them, with exception of KLK3 and KLK7. Nevertheless, mRNAs for KLK2, KLK5, KLK6 and KLK9 that were previously undetectable appeared after challenging with a mixture of cytokines. Both kinin B(1)R agonist and fMLP induced secretion of KLK1, KLK6, KLK10, KLK13 and KLK14 into the culture medium in similar amounts, whereas the B(1)R agonist caused the release of lower amounts of KLK2, KLK4 and KLK5. When secreted, the differing proteolytic activity of KLKs provides the human neutrophil with a multifunctional enzymatic capacity supporting a new dimension for its role in human disorders of diverse etiology.

  2. Temporal variations of glandular kallikrein, protein and amylase in mixed human saliva.

    PubMed

    Jenzano, J W; Brown, C K; Mauriello, S M

    1987-01-01

    Variations in the level of glandular kallikrein in human saliva may reflect physiological changes. Diurnal or circadian variations of many salivary components are important in relating changes in such components to oral or systemic conditions especially as most clinical studies are conducted between 0800 and 1700 h. Whole saliva was collected from 14 healthy young subjects at 0800, 1100, 1400 and 1700 h on two Fridays. Samples were centrifuged at 10,000 g for 10 min at 4 degrees C and the supernatant fractions stored at -20 degrees C. The enzymic activity of kallikrein was measured with D-valylleucylarginine-p-nitro-anilide as substrate. The activity of alpha-amylase and the total protein concentration (biuret) were also determined. Results were analysed in a repeated-measures design: there were no significant differences in kallikrein levels either within days or across days. There were significant differences for total protein and alpha-amylase levels within days but, in general, not across days. Minimal individual levels for protein and alpha-amylase were mostly at 0800 h; maxima were generally at 1400 or 1700 h. Kallikrein levels had no marked pattern of maximal or minimal distribution.

  3. Cleavage Activation of Human-adapted Influenza Virus Subtypes by Kallikrein-related Peptidases 5 and 12*

    PubMed Central

    Hamilton, Brian S.; Whittaker, Gary R.

    2013-01-01

    A critical step in the influenza virus replication cycle is the cleavage activation of the HA precursor. Cleavage activation of influenza HA enables fusion with the host endosome, allowing for release of the viral genome into the host cell. To date, studies have determined that HA activation is driven by trypsin-like host cell proteases, as well as yet to be identified bacterial proteases. Although the number of host proteases that can activate HA is growing, there is still uncertainty regarding which secreted proteases are able to support multicycle replication of influenza. In this study, we have determined that the kallikrein-related peptidases 5 and 12 are secreted from the human respiratory tract and have the ability to cleave and activate HA from the H1, H2, and H3 subtypes. Each peptidase appears to have a preference for particular influenza subtypes, with kallikrein 5 cleaving the H1 and H3 subtypes most efficiently and kallikrein 12 cleaving the H1 and H2 subtypes most efficiently. Cleavage analysis using HA cleavage site peptide mimics revealed that the amino acids neighboring the arginine cleavage site affect cleavage efficiency. Additionally, the thrombolytic zymogens plasminogen, urokinase, and plasma kallikrein have all been shown to cleave and activate influenza but are found circulating mainly as inactive precursors. Kallikrein 5 and kallikrein 12 were examined for their ability to activate the thrombolytic zymogens, and both resulted in activation of each zymogen, with kallikrein 12 being a more potent activator. Activation of the thrombolytic zymogens may therefore allow for both direct and indirect activation of the HA of human-adapted influenza viruses by kallikrein 5 and kallikrein 12. PMID:23612974

  4. Expression of kallikrein 4 (Klk4) in dental and non-dental tissues

    PubMed Central

    Simmer, James P.; Richardson, Amelia S.; Smith, Charles E.; Hu, Yuanyuan; Hu, Jan C-C.

    2011-01-01

    Kallikrein 4 (Klk4) is critical for proper dental enamel formation. Klk4 null mice and persons with two defective KLK4 alleles have obvious enamel defects, with no other apparent phenotype. Klk4 mRNA or protein is reported to be in tissues besides teeth, including prostate, ovary, kidney, liver, and salivary gland. In this study we used the Klk4 knockout/NLS-lacZ knockin mouse to assay Klk4 expression using ß-galactosidase histochemistry. Incubations for 5 h were used to detect Klk4 expression with minimal endogenous background, while overnight incubations susceptible to false positives were used to look for trace Klk4 expression. Developing maxillary molars at postnatal days 5, 6, 7, 8, and 14, developing mandibular incisors at postnatal day 14, and selected non-dental tissues from adult wild-type and Klk4lacZ/lacZ mice were examined by X-gal histochemistry. After 5 h incubation, Xgal staining was observed specifically in the nuclei of maturation stage ameloblasts in molars and incisors from Klk4lacZ/lacZ mice and was detected weakly in nuclei of salivary gland ducts and patches of prostate epithelia. We conclude that Klk4 is predominantly a tooth-specific protease with low expression in submandibular salivary gland and prostate, and with no detectable expression in liver, kidney, testis, ovary, oviduct, epididymis, and vas deferens. PMID:22243250

  5. Kallikrein genes are associated with lupus and glomerular basement membrane–specific antibody–induced nephritis in mice and humans

    PubMed Central

    Liu, Kui; Li, Quan-Zhen; Delgado-Vega, Angelica M.; Abelson, Anna-Karin; Sánchez, Elena; Kelly, Jennifer A.; Li, Li; Liu, Yang; Zhou, Jinchun; Yan, Mei; Ye, Qiu; Liu, Shenxi; Xie, Chun; Zhou, Xin J.; Chung, Sharon A.; Pons-Estel, Bernardo; Witte, Torsten; de Ramón, Enrique; Bae, Sang-Cheol; Barizzone, Nadia; Sebastiani, Gian Domenico; Merrill, Joan T.; Gregersen, Peter K.; Gilkeson, Gary G.; Kimberly, Robert P.; Vyse, Timothy J.; Kim, Il; D’Alfonso, Sandra; Martin, Javier; Harley, John B.; Criswell, Lindsey A.; Wakeland, Edward K.; Alarcón-Riquelme, Marta E.; Mohan, Chandra

    2009-01-01

    Immune-mediated nephritis contributes to disease in systemic lupus erythematosus, Goodpasture syndrome (caused by antibodies specific for glomerular basement membrane [anti-GBM antibodies]), and spontaneous lupus nephritis. Inbred mouse strains differ in susceptibility to anti-GBM antibody–induced and spontaneous lupus nephritis. This study sought to clarify the genetic and molecular factors that may be responsible for enhanced immune-mediated renal disease in these models. When the kidneys of 3 mouse strains sensitive to anti-GBM antibody–induced nephritis were compared with those of 2 control strains using microarray analysis, one-fifth of the underexpressed genes belonged to the kallikrein gene family, which encodes serine esterases. Mouse strains that upregulated renal and urinary kallikreins exhibited less evidence of disease. Antagonizing the kallikrein pathway augmented disease, while agonists dampened the severity of anti-GBM antibody–induced nephritis. In addition, nephritis-sensitive mouse strains had kallikrein haplotypes that were distinct from those of control strains, including several regulatory polymorphisms, some of which were associated with functional consequences. Indeed, increased susceptibility to anti-GBM antibody–induced nephritis and spontaneous lupus nephritis was achieved by breeding mice with a genetic interval harboring the kallikrein genes onto a disease-resistant background. Finally, both human SLE and spontaneous lupus nephritis were found to be associated with kallikrein genes, particularly KLK1 and the KLK3 promoter, when DNA SNPs from independent cohorts of SLE patients and controls were compared. Collectively, these studies suggest that kallikreins are protective disease-associated genes in anti-GBM antibody–induced nephritis and lupus. PMID:19307730

  6. Renal (tissue) kallikrein-kinin system in the kidney and novel potential drugs for salt-sensitive hypertension.

    PubMed

    Katori, Makoto; Majima, Masataka

    2014-01-01

    A large variety of antihypertensive drugs, such as angiotensin converting enzyme inhibitors, diuretics, and others, are prescribed to hypertensive patients, with good control of the condition. In addition, all individuals are generally believed to be salt sensitive and, thus, severe restriction of salt intake is recommended to all. Nevertheless, the physiological defense mechanisms in the kidney against excess salt intake have not been well clarified. The present review article demonstrated that the renal (tissue) kallikrein-kinin system (KKS) is ideally situated within the nephrons of the kidney, where it functions to inhibit the reabsorption of NaCl through the activation of bradykinin (BK)-B2 receptors localized along the epithelial cells of the collecting ducts (CD). Kinins generated in the CD are immediately inactivated by two kidney-specific kinin-inactivating enzymes (kininases), carboxypeptidase Y-like exopeptidase (CPY), and neutral endopeptidase (NEP). Our work demonstrated that ebelactone B and poststatin are selective inhibitors of these kininases. The reduced secretion of the urinary kallikrein is linked to the development of salt-sensitive hypertension, whereas potassium ions and ATP-sensitive potassium channel blockers ameliorate salt-sensitive hypertension by accelerating the release of renal kallikrein. On the other hand, ebelactone B and poststatin prolong the life of kinins in the CD after excess salt intake, thereby leading to the augmentation of natriuresis and diuresis, and the ensuing suppression of salt-sensitive hypertension. In conclusion, accelerators of the renal kallikrein release and selective renal kininase inhibitors are both novel types of antihypertensive agents that may be useful for treatment of salt-sensitive hypertension. PMID:25130040

  7. Kallikrein-8 Proteolytically Processes Human Papillomaviruses in the Extracellular Space To Facilitate Entry into Host Cells

    PubMed Central

    Cerqueira, Carla; Samperio Ventayol, Pilar; Vogeley, Christian

    2015-01-01

    ABSTRACT The entry of human papillomaviruses into host cells is a complex process. It involves conformational changes at the cell surface, receptor switching, internalization by a novel endocytic mechanism, uncoating in endosomes, trafficking of a subviral complex to the Golgi complex, and nuclear entry during mitosis. Here, we addressed how the stabilizing contacts in the capsid of human papillomavirus 16 (HPV16) may be reversed to allow uncoating of the viral genome. Using biochemical and cell-biological analyses, we determined that the major capsid protein L1 underwent proteolytic cleavage during entry. In addition to a dispensable cathepsin-mediated proteolysis that occurred likely after removal of capsomers from the subviral complex in endosomes, at least two further proteolytic cleavages of L1 were observed, one of which was independent of the low-pH environment of endosomes. This cleavage occurred extracellularly. Further analysis showed that the responsible protease was the secreted trypsin-like serine protease kallikrein-8 (KLK8) involved in epidermal homeostasis and wound healing. Required for infection, the cleavage was facilitated by prior interaction of viral particles with heparan sulfate proteoglycans. KLK8-mediated cleavage was crucial for further conformational changes exposing an important epitope of the minor capsid protein L2. Occurring independently of cyclophilins and of furin that mediate L2 exposure, KLK8-mediated cleavage of L1 likely facilitated access to L2, located in the capsid lumen, and potentially uncoating. Since HPV6 and HPV18 also required KLK8 for entry, we propose that the KLK8-dependent entry step is conserved. IMPORTANCE Our analysis of the proteolytic processing of incoming HPV16, an etiological agent of cervical cancer, demonstrated that the capsid is cleaved extracellularly by a serine protease active during wound healing and that this cleavage was crucial for infection. The cleavage of L1 is one of at least four structural

  8. Thrombin and human plasma kallikrein inhibition during simulated extracorporeal circulation block platelet and neutrophil activation.

    PubMed

    Wachtfogel, Y T; Kettner, C; Hack, C E; Nuijens, J H; Reilly, T M; Knabb, R M; Kucich, U; Niewiarowski, S; Edmunds, L H; Colman, R W

    1998-10-01

    Cardiopulmonary bypass causes hemorrhagic complications, and initiates a chemical and cellular inflammatory response. Contact of blood with synthetic surfaces leads to qualitative and quantitative alterations in platelets, neutrophils, complement, and contact systems. Despite the fact that cardiopulmonary bypass is carried out in the presence of high doses of heparin, there is significant activation of both platelets and neutrophils. Thrombin is protected on cell and fibrin surfaces from antithrombin, even in the presence of high doses of heparin (approximately 5 U/ml). We therefore studied the effect of a small (Mr = 497), highly effective (Ki = 41 pM), reversible tripeptide inhibitor of thrombin, DUP 714 (1 microM), in a well characterized model of simulated extracorporeal circulation. In the absence of DUP 714, platelet counts decreased by 75% 5 min after the start of extracorporeal bypass and increased to 48% at 120 min of recirculation. DUP 714 significantly preserved platelet counts, decreased plasma levels of platelet beta-thromboglobulin levels, but did not prevent a decrease in sensitivity of platelets to adenosine diphosphate. Kallikrein-C1-inhibitor and C1-C1-inhibitor complexes increased progressively from 0.32 U/ml to 0.67 U/ml and from 4.45 U/ml to 7.25 U/ml, respectively, during 120 min of recirculation without DUP 714. Addition of DUP 714 significantly inhibited kallikrein-C1-inhibitor complex formation but did not affect C1-C1-inhibitor complexes. In the absence of DUP 714, human neutrophil elastase levels rose from a baseline of 0.01 +/- 0.00 microg/ml to 1.18 +/- 0.21 microg/ml during 120 min of recirculation. Human neutrophil elastase release at 120 min was significantly inhibited in the presence of DUP 714 to 37% of the value with heparin alone. These results indicated that addition of this novel thrombin (and kallikrein) inhibitor to heparin preserved platelet counts, decreased platelet secretion, and provided the additional benefit of

  9. SPINK5 knockdown in organotypic human skin culture as a model system for Netherton syndrome: effect of genetic inhibition of serine proteases kallikrein 5 and kallikrein 7.

    PubMed

    Wang, Shirley; Olt, Sabine; Schoefmann, Nicole; Stuetz, Anton; Winiski, Anthony; Wolff-Winiski, Barbara

    2014-07-01

    Netherton syndrome (NS; OMIM 256500) is a genetic skin disease resulting from defects in the serine protease inhibitor Kazal-type 5 (SPINK5) gene, which encodes the protease inhibitor lympho-epithelial Kazal type inhibitor (LEKTI). We established a SPINK5 knockdown skin model by transfecting SPINK5 small interfering RNA (siRNA) into normal human epidermal keratinocytes, which were used together with fibroblast-populated collagen gels to generate organotypic skin cultures. This model recapitulates some of the NS skin morphology: thicker, parakeratotic stratum corneum frequently detached from the underlying epidermis and loss of corneodesmosomes. As enhanced serine protease activity has been implicated in the disease pathogenesis, we investigated the impact of the kallikreins KLK5 [stratum corneum trypsin-like enzyme (SCTE)] and KLK7 [stratum corneum chymotrypsin-like enzyme (SCCE)] on the SPINK5 knockdown phenotype by generating double knockdowns in the organotypic model. Knockdown of KLK5 or KLK7 partially ameliorated the epidermal architecture: increased epidermal thickness and expression of desmocollin 1 (DSC1), desmoglein 1 (DSG1) and (pro)filaggrin. Thus, inhibition of serine proteases KLK5 and KLK7 could be therapeutically beneficial in NS.

  10. Quantification of Human Kallikrein-Related Peptidases in Biological Fluids by Multiplatform Targeted Mass Spectrometry Assays.

    PubMed

    Karakosta, Theano D; Soosaipillai, Antoninus; Diamandis, Eleftherios P; Batruch, Ihor; Drabovich, Andrei P

    2016-09-01

    Human kallikrein-related peptidases (KLKs) are a group of 15 secreted serine proteases encoded by the largest contiguous cluster of protease genes in the human genome. KLKs are involved in coordination of numerous physiological functions including regulation of blood pressure, neuronal plasticity, skin desquamation, and semen liquefaction, and thus represent promising diagnostic and therapeutic targets. Until now, quantification of KLKs in biological and clinical samples was accomplished by enzyme-linked immunosorbent assays (ELISA). Here, we developed multiplex targeted mass spectrometry assays for the simultaneous quantification of all 15 KLKs. Proteotypic peptides for each KLK were carefully selected based on experimental data and multiplexed in single assays. Performance of assays was evaluated using three different mass spectrometry platforms including triple quadrupole, quadrupole-ion trap, and quadrupole-orbitrap instruments. Heavy isotope-labeled synthetic peptides with a quantifying tag were used for absolute quantification of KLKs in sweat, cervico-vaginal fluid, seminal plasma, and blood serum, with limits of detection ranging from 5 to 500 ng/ml. Analytical performance of assays was evaluated by measuring endogenous KLKs in relevant biological fluids, and results were compared with selected ELISAs. The multiplex targeted proteomic assays were demonstrated to be accurate, reproducible, sensitive, and specific alternatives to antibody-based assays. Finally, KLK4, a highly prostate-specific protein and a speculated biomarker of prostate cancer, was unambiguously detected and quantified by immunoenrichment-SRM assay in seminal plasma and blood serum samples from individuals with confirmed prostate cancer and negative biopsy. Mass spectrometry revealed exclusively the presence of a secreted isoform and thus unequivocally resolved earlier disputes about KLK4 identity in seminal plasma. Measurements of KLK4 in either 41 seminal plasma or 58 blood serum samples

  11. Tissue kallikrein induces SH-SY5Y cell proliferation via epidermal growth factor receptor and extracellular signal-regulated kinase1/2 pathway

    SciTech Connect

    Lu, Zhengyu; Yang, Qi; Cui, Mei; Liu, Yanping; Wang, Tao; Zhao, Hong; Dong, Qiang

    2014-03-28

    Highlights: • TK promotes EGFR phosphorylation in SH-SY5Y cells. • TK activates ERK1/2 and p38 phosphorylation in SH-SY5Y cells. • TK mediates SH-SY5Y cell proliferation via EGFR and ERK1/2 pathway. - Abstract: Tissue kallikrein (TK) is well known to take most of its biological functions through bradykinin receptors. In the present study, we found a novel signaling pathway mediated by TK through epidermal growth factor receptor (EGFR) in human SH-SY5Y cells. We discovered that TK facilitated the activation of EGFR, extracellular signal-regulated kinase (ERK) 1/2 and p38 cascade. Interestingly, not p38 but ERK1/2 phosphorylation was severely compromised in cells depleted of EGFR. Nevertheless, impairment of signaling of ERK1/2 seemed not to be restricted to EGFR phosphorylation. We also observed that TK stimulation could induce SH-SY5Y cell proliferation, which was reduced by EGFR down-regulation or ERK1/2 inhibitor. Overall, our findings provided convincing evidence that TK could mediate cell proliferation via EGFR and ERK1/2 pathway in vitro.

  12. The kallikrein-kinin system in diabetic retinopathy.

    PubMed

    Bhat, Menakshi; Pouliot, Mylène; Couture, Réjean; Vaucher, Elvire

    2014-01-01

    Diabetic retinopathy (DR) is a major microvascular complication associated with type 1 and type 2 diabetes mellitus, which can lead to visual impairment and blindness. Current treatment strategies for DR are mostly limited to laser therapies, steroids, and anti-VEGF agents, which are often associated with unwanted side effects leading to further complications. Recent evidence suggests that kinins play a primary role in the development of DR through enhanced vascular permeability, leukocytes infiltration, and other inflammatory mechanisms. These deleterious effects are mediated by kinin B1 and B2 receptors, which are expressed in diabetic human and rodent retina. Importantly, kinin B1 receptor is virtually absent in sane tissue, yet it is induced and upregulated in diabetic retina. These peptides belong to the kallikrein-kinin system (KKS), which contains two separate and independent pathways of regulated serine proteases, namely plasma kallikrein (PK) and tissue kallikrein (TK) that are involved in the biosynthesis of bradykinin (BK) and kallidin (Lys-BK), respectively. Hence, ocular inhibition of kallikreins or antagonism of kinin receptors offers new therapeutic avenues in the treatment and management of DR. Herein, we present an overview of the principal features and known inflammatory mechanisms associated with DR along with the current therapeutic approaches and put special emphasis on the KKS as a new and promising therapeutic target due to its link with key pathways directly associated with the development of DR. PMID:25130041

  13. A novel signaling pathway of tissue kallikrein in promoting keratinocyte migration: Activation of proteinase-activated receptor 1 and epidermal growth factor receptor

    SciTech Connect

    Gao, Lin; Chao, Lee; Chao, Julie

    2010-02-01

    Biological functions of tissue kallikrein (TK, KLK1) are mainly mediated by kinin generation and subsequent kinin B2 receptor activation. In this study, we investigated the potential role of TK and its signaling pathways in cultured human keratinocyte migration and in a rat skin wound healing model. Herein, we show that TK promoted cell migration and proliferation in a concentration- and time-dependent manner. Inactive TK or kinin had no significant effect on cell migration. Interestingly, cell migration induced by active TK was not blocked by icatibant or L-NAME, indicating an event independent of kinin B2 receptor and nitric oxide formation. TK's stimulatory effect on cell migration was inhibited by small interfering RNA for proteinase-activated receptor 1 (PAR{sub 1}), and by PAR{sub 1} inhibitor. TK-induced migration was associated with increased phosphorylation of epidermal growth factor receptor (EGFR) and extracellular signal-regulated kinase (ERK), which was blocked by inhibition of protein kinase C (PKC), Src, EGFR and ERK. TK-induced cell migration and EGFR phosphorylation were blocked by metalloproteinase (MMP) inhibitor, heparin, and antibodies against EGFR external domain, heparin-binding EGF-like growth factor (HB-EGF) and amphiregulin (AR). Local application of TK promoted skin wound healing in rats, whereas icatibant and EGFR inhibitor blocked TK's effect. Skin wound healing was further delayed by aprotinin and neutralizing TK antibody. This study demonstrates a novel role of TK in skin wound healing and uncovers new signaling pathways mediated by TK in promoting keratinocyte migration through activation of the PAR{sub 1}-PKC-Src-MMP pathway and HB-EGF/AR shedding-dependent EGFR transactivation.

  14. A Single Glycan at the 99-Loop of Human Kallikrein-related Peptidase 2 Regulates Activation and Enzymatic Activity*

    PubMed Central

    Guo, Shihui; Skala, Wolfgang; Magdolen, Viktor; Briza, Peter; Biniossek, Martin L.; Schilling, Oliver; Kellermann, Josef; Brandstetter, Hans; Goettig, Peter

    2016-01-01

    Human kallikrein-related peptidase 2 (KLK2) is a key serine protease in semen liquefaction and prostate cancer together with KLK3/prostate-specific antigen. In order to decipher the function of its potential N-glycosylation site, we produced pro-KLK2 in Leishmania tarentolae cells and compared it with its non-glycosylated counterpart from Escherichia coli expression. Mass spectrometry revealed that Asn-95 carries a core glycan, consisting of two GlcNAc and three hexoses. Autocatalytic activation was retarded in glyco-pro-KLK2, whereas the activated glyco-form exhibited an increased proteolytic resistance. The specificity patterns obtained by the PICS (proteomic identification of protease cleavage sites) method are similar for both KLK2 variants, with a major preference for P1-Arg. However, glycosylation changes the enzymatic activity of KLK2 in a drastically substrate-dependent manner. Although glyco-KLK2 has a considerably lower catalytic efficiency than glycan-free KLK2 toward peptidic substrates with P2-Phe, the situation was reverted toward protein substrates, such as glyco-pro-KLK2 itself. These findings can be rationalized by the glycan-carrying 99-loop that prefers to cover the active site like a lid. By contrast, the non-glycosylated 99-loop seems to favor a wide open conformation, which mostly increases the apparent affinity for the substrates (i.e. by a reduction of Km). Also, the cleavage pattern and kinetics in autolytic inactivation of both KLK2 variants can be explained by a shift of the target sites due to the presence of the glycan. These striking effects of glycosylation pave the way to a deeper understanding of kallikrein-related peptidase biology and pathology. PMID:26582203

  15. Helodermatine, a kallikrein-like, hypotensive enzyme from the venom of Heloderma horridum horridum (Mexican beaded lizard)

    PubMed Central

    1986-01-01

    We have purified and characterized the major N-benzoyl-L-arginine ethyl ester hydrolase from the venom of Heloderma horridum horridum. The enzyme belongs to the serine proteinase family, and its activity vs. peptide amide substrates and human high-molecular-weight kininogen suggests a similarity to the family of kallikreins. This interpretation is corroborated by its reactivity with the natural inhibitors soybean trypsin inhibitor and Kunitz-type bovine pancreatic trypsin inhibitor (aprotinin). Injection of the enzyme (2-16 micrograms/kg) into anesthetized rabbits leads to a rapid dose-dependent transient decrease of the arterial blood pressure. Like glandular kallikrein it specifically converts single-chain tissue type plasminogen activator into its double chain form. In contrast to other kallikrein-like enzymes from snake venoms it shows no thrombin-like or plasminogen activator activity. The enzyme is a single-chain glycoprotein (Mr 63,000). The N-terminal sequence revealed significant homology to pig pancreatic kallikrein and to kallikrein like enzymes from Crotalus atrox and Crotalus adamanteus venom. This enzyme, which we name Helodermatine, is the first purified from Sauria with kallikrein-like properties. PMID:3537191

  16. Synergistic effect of a tissue kallikrein 1 and tissue inhibitor of matrix metalloproteinase 1 co‑expression vector on the proliferation of rat vascular smooth muscle cells.

    PubMed

    Zhu, Pengli; Yu, Huizhen; Huang, Shujie; Xiang, Hong; Li, Feng; Zheng, Weiping

    2015-10-01

    Tissue kallikrein 1 (TK1) and tissue inhibitor of matrix metalloproteinase 1 (TIMP1) are important in inhibiting vascular smooth muscle cell (VSMC) proliferation and improving vascular remodeling, respectively. It was hypothesized that a combination of TK1 and TIMP1 genes, mediated by an adenovirus vector could augment or act in synergy to enhance the inhibitory effects. The promoter, mCMV carrying hTIMP1 cDNA was subcloned into pDC316‑hTK1 to construct a recombinant plasmid carrying hTK1 and hTIMP1 genes. Subsequently, the double gene plasmid and adenovirus backbone plasmid were packaged into HEK293A cells. Gene transcription and protein expression were examined, respectively using reverse transcription‑quantitative polymerase chain reaction (PCR) and western blotting assays. VSMC proliferation was assessed using cell counting and methyl‑thiazolyl‑tetrazoliuin methods. The constructed plasmid containing hTK1 and hTIMP1 genes was correctly identified by means of PCR, double digestion and sequencing analysis. The co‑expression vector, Ad‑hTK1‑hTIMP1 was successfully constructed and packaged into HEK293A cells. When VSMCs were transfected with the co‑expression vector, the mRNA transcription and protein expression of hTK1 and hTIMP1 exhibited abundant expression in a concentration‑dependent and time‑dependent manner, independently. In conclusion, the co‑expression vector synergistically inhibited the cell growth and proliferation induced by platelet‑derived growth factor‑BB compared with the single gene vector.

  17. Kallikrein isolated from commercial crystalline pepsin preparations.

    PubMed

    Ferreira, L A; Henriques, O B

    1987-01-01

    1. An experiment designed to study the relationship between pH and kininogenase activity of three commercial preparations of porcine crystallized pepsin showed that each preparation had two well separated pH optima, pH 4 and 8. 2. From the inhibition spectrum of the pH 8 kininogenase it was concluded that it is a kallikrein of the glandular type, since it proved to be a serine protease and was insensitive to protein trypsin inhibitors. 3. Kallikrein activity can be separated from pepsin by affinity chromatography on Sepharose-4B-Pro-Phe-agmatine. This separation permitted us to obtain purified material with kallikrein specific activity 43 times higher than that of crude pepsin and which showed a single band on polyacrylamide gel electrophoresis. 4. The kallikrein activity was found to have a molecular weight of 36 kDal and a Michaelis constant of 25 microM when acting on Bz-Pro-Phe-Arg-p-nitroanilide at pH 8.6. 5. On the basis of these properties, kallikrein from commercial pepsin resembles the kallikreins previously described from rat or human stomach. PMID:3133004

  18. Tissue Kallikrein Alleviates Cerebral Ischemia-Reperfusion Injury by Activating the B2R-ERK1/2-CREB-Bcl-2 Signaling Pathway in Diabetic Rats

    PubMed Central

    Yuan, Kunxiong; Hu, Bin; Sang, Hongfei; Xie, Yi; Xu, Lili; Cao, Qinqin; Chen, Xin; Zhao, Lingling; Liu, Xinfeng; Liu, Ling; Zhang, Renliang

    2016-01-01

    Diabetes mellitus (DM) substantially increases the risk of ischemic stroke and reduces the tolerance to ischemic insults. Tissue kallikrein (TK) has been demonstrated to protect neurons from ischemia/reperfusion (I/R) injury in orthoglycemic model by activating the bradykinin B2 receptor (B2R). Considering the differential effects of B2R or bradykinin B1 receptor (B1R) on cardioprotection and neuroprotection in I/R with or without diabetes, this study was designed to investigate the role of TK during cerebral I/R injury in streptozotocin-induced diabetic rats. Intravenous injection of TK inhibited apoptosis in neurons, alleviated edema and inflammatory reactions after focal cerebral I/R, significantly reduced the infarct volume, and improved functional recovery. These beneficial effects were accompanied by activation of the extracellular signal-regulated kinase 1/2 (ERK1/2), cAMP response element-binding (CREB), and Bcl-2 signal proteins. Inhibition of the B2R or ERK1/2 pathway abated the effects of TK, whereas an antagonist of B1R enhanced the effects. These findings reveal that the neuroprotective effect of TK against cerebral I/R injury in streptozotocin-induced diabetic rats mainly involves the enhancement of B2R and ERK1/2-CREB-Bcl-2 signaling pathway activity. PMID:27446506

  19. Tissue Kallikrein Alleviates Cerebral Ischemia-Reperfusion Injury by Activating the B2R-ERK1/2-CREB-Bcl-2 Signaling Pathway in Diabetic Rats

    PubMed Central

    Yuan, Kunxiong; Hu, Bin; Sang, Hongfei; Xie, Yi; Xu, Lili; Cao, Qinqin; Chen, Xin; Zhao, Lingling; Liu, Xinfeng; Liu, Ling; Zhang, Renliang

    2016-01-01

    Diabetes mellitus (DM) substantially increases the risk of ischemic stroke and reduces the tolerance to ischemic insults. Tissue kallikrein (TK) has been demonstrated to protect neurons from ischemia/reperfusion (I/R) injury in orthoglycemic model by activating the bradykinin B2 receptor (B2R). Considering the differential effects of B2R or bradykinin B1 receptor (B1R) on cardioprotection and neuroprotection in I/R with or without diabetes, this study was designed to investigate the role of TK during cerebral I/R injury in streptozotocin-induced diabetic rats. Intravenous injection of TK inhibited apoptosis in neurons, alleviated edema and inflammatory reactions after focal cerebral I/R, significantly reduced the infarct volume, and improved functional recovery. These beneficial effects were accompanied by activation of the extracellular signal-regulated kinase 1/2 (ERK1/2), cAMP response element-binding (CREB), and Bcl-2 signal proteins. Inhibition of the B2R or ERK1/2 pathway abated the effects of TK, whereas an antagonist of B1R enhanced the effects. These findings reveal that the neuroprotective effect of TK against cerebral I/R injury in streptozotocin-induced diabetic rats mainly involves the enhancement of B2R and ERK1/2-CREB-Bcl-2 signaling pathway activity.

  20. Gene structure and chromosomal localization of plasma kallikrein

    SciTech Connect

    Beaubien, G.; Mbikay, M.; Chretien, M.; Seidah, N.G. ); Rosinski-Chupin, I. ); Mattei, M.G. )

    1991-02-12

    Plasma kallikrein (Fletcher factor) is a hepatic serine proteinase that participates in the early phase of blood coagulation. From two genomic libraries, the authors succeeded to isolate four overlapping clones representing the entire rat plasma kallikrein gene. Using selective DNA sequencing, polymerase chain reactions, and restriction mapping, the authors demonstrated that the gene for rat plasma kallikrein was 22 kb in length. Similar to human factor XI the authors also found that the plasma kallikrein gene is composed of 15 exons and 14 introns. A potential transcription initiation step was determined by a novel application of the polymerase chain reaction technique. Computer analysis of the 5{prime}-promoter region of this gene revealed some putative control elements that might regulate the rat plasma kallikrein gene expression. These data and the results of chromosomal localization reported in the present study for mouse (chromosome 8) and human (chromosome 4) plasma kallikrein genes strongly corroborate a genic duplication event from a common ancestor to both plasma kallikrein and factor XI.

  1. A Highly Sensitive Porous Silicon (P-Si)-Based Human Kallikrein 2 (hK2) Immunoassay Platform toward Accurate Diagnosis of Prostate Cancer.

    PubMed

    Lee, Sang Wook; Hosokawa, Kazuo; Kim, Soyoun; Jeong, Ok Chan; Lilja, Hans; Laurell, Thomas; Maeda, Mizuo

    2015-05-22

    Levels of total human kallikrein 2 (hK2), a protein involved the pathology of prostate cancer (PCa), could be used as a biomarker to aid in the diagnosis of this disease. In this study, we report on a porous silicon antibody immunoassay platform for the detection of serum levels of total hK2. The surface of porous silicon has a 3-dimensional macro- and nanoporous structure, which offers a large binding capacity for capturing probe molecules. The tailored pore size of the porous silicon also allows efficient immobilization of antibodies by surface adsorption, and does not require chemical immobilization. Monoclonal hK2 capture antibody (6B7) was dispensed onto P-Si chip using a piezoelectric dispenser. In total 13 × 13 arrays (169 spots) were spotted on the chip with its single spot volume of 300 pL. For an optimization of capture antibody condition, we firstly performed an immunoassay of the P-Si microarray under a titration series of hK2 in pure buffer (PBS) at three different antibody densities (75, 100 and 145 µg/mL). The best performance of the microarray platform was seen at 100 µg/mL of the capture antibody concentration (LOD was 100 fg/mL). The platform then was subsequently evaluated for a titration series of serum-spiked hK2 samples. The developed platform utilizes only 15 µL of serum per test and the total assay time is about 3 h, including immobilization of the capture antibody. The detection limit of the hK2 assay was 100 fg/mL in PBS buffer and 1 pg/mL in serum with a dynamic range of 106 (10(-4) to 10(2) ng/mL).

  2. [Human brown adipose tissue].

    PubMed

    Virtanen, Kirsi A; Nuutila, Pirjo

    2015-01-01

    Adult humans have heat-producing and energy-consuming brown adipose tissue in the clavicular region of the neck. There are two types of brown adipose cells, the so-called classic and beige adipose cells. Brown adipose cells produce heat by means of uncoupler protein 1 (UCP1) from fatty acids and sugar. By applying positron emission tomography (PET) measuring the utilization of sugar, the metabolism of brown fat has been shown to multiply in the cold, presumably influencing energy consumption. Active brown fat is most likely present in young adults, persons of normal weight and women, least likely in obese persons.

  3. Human Tissue Stimulator

    NASA Technical Reports Server (NTRS)

    1982-01-01

    Neurodyne Corporation Human Tissue Stimulator (HTS) is a totally implantable system used for treatment of chronic pain and involuntary motion disorders by electrical stimulation. It was developed by Pacesetter Systems, Inc. in cooperation with the Applied Physics Laboratory. HTS incorporates a nickel cadmium battery, telemetry and command systems technologies of the same type as those used in NASA's Small Astronomy Satellite-3 in microminiature proportions so that the implantable element is the size of a deck of cards. The stimulator includes a rechargeable battery, an antenna and electronics to receive and process commands and to report on its own condition via telemetry, a wireless process wherein instrument data is converted to electrical signals and sent to a receiver where signals are presented as usable information. The HTS is targeted to nerve centers or to particular areas of the brain to provide relief from intractable pain or arrest involuntary motion. The nickel cadmium battery can be recharged through the skin. The first two HTS units were implanted last year and have been successful. Extensive testing is required before HTS can be made available for general use.

  4. The relationship between kallikrein and water excretion and the conditional relationship between kallikrein and sodium excretion.

    PubMed Central

    Mills, I H; Ward, P E

    1975-01-01

    1. The renal kallikrein-kinin system has previously been linked with renal control of sodium and water excretion. The present investigations were carried out to examine more closely these relationships. 2. In physiological studies with rabbits, urinary kallikrein was measured by a modification of the [3-H]TAME method. 3. With rabbits on free sodium and water intake, urinary kallikrein was positively correlated with both sodium and water excretion. Kallikrein excretion was also negatively correlated with urinary osmolality. 4. In rabbits on chronic high and low sodium diets, urinary kallikrein was positively correlated with urinary volume but not with sodium excretion. 5. In rabbits held to a constant fluid intake but with sodium intake changed, urinary kallikrein was not correlated with sodium excretion. 6. These results indicate that the positive correlation of kallikrein excretion with sodium excretion under conditions of free sodium and water intake may be only secondary to the positive relationship of kallikrein excretion with urinary volume. 7. The results of the present investigations do not support the hypothesis that the renal kallikrein-kinin system is necessarily involved in renal control of sodium excretion under normal conditions but it is where a change in sodium intake leads to a change in fluid intake and consequently of urinary volume. 8. In the above experiments, urinary kallikrein was always positively correlated with urinary volume and negatively correlated with urinary osmolality. This may indicate a functional relationship between renal kallikrein and water excretion. PMID:1133793

  5. Trade in human tissue products.

    PubMed

    Tonti-Filippini, Nicholas; Zeps, Nikolajs

    2011-03-01

    Trade in human tissue in Australia is prohibited by state law, and in ethical guidelines by the National Health and Medical Research Council: National statement on ethical conduct in human research; Organ and tissue donation by living donors: guidelines for ethical practice for health professionals. However, trade in human tissue products is a common practice especially for: reconstructive orthopaedic or plastic surgery; novel human tissue products such as a replacement trachea created by using human mesenchymal stem cells; biomedical research using cell lines, DNA and protein provided through biobanks. Cost pressures on these have forced consideration of commercial models to sustain their operations. Both the existing and novel activities require a robust framework to enable commercial uses of human tissue products while maintaining community acceptability of such practices, but to date no such framework exists. In this article, we propose a model ethical framework for ethical governance which identifies specific ethical issues such as: privacy; unique value of a person's tissue; commodification of the body; equity and benefit to the community; perverse incentives; and "attenuation" as a potentially useful concept to help deal with the broad range of subjective views relevant to whether it is acceptable to commercialise certain human tissue products. PMID:21382003

  6. Trade in human tissue products.

    PubMed

    Tonti-Filippini, Nicholas; Zeps, Nikolajs

    2011-03-01

    Trade in human tissue in Australia is prohibited by state law, and in ethical guidelines by the National Health and Medical Research Council: National statement on ethical conduct in human research; Organ and tissue donation by living donors: guidelines for ethical practice for health professionals. However, trade in human tissue products is a common practice especially for: reconstructive orthopaedic or plastic surgery; novel human tissue products such as a replacement trachea created by using human mesenchymal stem cells; biomedical research using cell lines, DNA and protein provided through biobanks. Cost pressures on these have forced consideration of commercial models to sustain their operations. Both the existing and novel activities require a robust framework to enable commercial uses of human tissue products while maintaining community acceptability of such practices, but to date no such framework exists. In this article, we propose a model ethical framework for ethical governance which identifies specific ethical issues such as: privacy; unique value of a person's tissue; commodification of the body; equity and benefit to the community; perverse incentives; and "attenuation" as a potentially useful concept to help deal with the broad range of subjective views relevant to whether it is acceptable to commercialise certain human tissue products.

  7. Lack of plasma kallikrein-kinin system cascade in teleosts.

    PubMed

    Wong, Marty Kwok-Shing; Takei, Yoshio

    2013-01-01

    The kallikrein-kinin system (KKS) consists of two major cascades in mammals: "plasma KKS" consisting of high molecular-weight (HMW) kininogen (KNG), plasma kallikrein (KLKB1), and bradykinin (BK); and "tissue KKS" consisting of low molecular-weight (LMW) KNG, tissue kallikreins (KLKs), and [Lys(0)]-BK. Some components of the KKS have been identified in the fishes, but systematic analyses have not been performed, thus this study aims to define the KKS components in teleosts and pave a way for future physiological and evolutionary studies. Through a combination of genomics, molecular, and biochemical methods, we showed that the entire plasma KKS cascade is absent in teleosts. Instead of two KNGs as found in mammals, a single molecular weight KNG was found in various teleosts, which is homologous to the mammalian LMW KNG. Results of molecular phylogenetic and synteny analyses indicated that the all current teleost genomes lack KLKB1, and its unique protein structure, four apple domains and one trypsin domain, could not be identified in any genome or nucleotide databases. We identified some KLK-like proteins in teleost genomes by synteny and conserved domain analyses, which could be the orthologs of tetrapod KLKs. A radioimmunoassay system was established to measure the teleost BK and we found that [Arg(0)]-BK is the major circulating form instead of BK, which supports that the teleost KKS is similar to the mammalian tissue KKS. Coincidently, coelacanths are the earliest vertebrate that possess both HMW KNG and KLKB1, which implies that the plasma KKS could have evolved in the early lobe-finned fish and descended to the tetrapod lineage. The co-evolution of HMW KNG and KLKB1 in lobe-finned fish and early tetrapods may mark the emergence of the plasma KKS and a contact activation system in blood coagulation, while teleosts may have retained a single KKS cascade. PMID:24278376

  8. Genetic manipulation and genetic variation of the kallikrein-kinin system: impact on cardiovascular and renal diseases.

    PubMed

    Girolami, Jean-Pierre; Blaes, Nelly; Bouby, Nadine; Alhenc-Gelas, François

    2014-01-01

    Genetic manipulation of the kallikrein-kinin system (KKS) in mice, with either gain or loss of function, and study of human genetic variability in KKS components which has been well documented at the phenotypic and genomic level, have allowed recognizing the physiological role of KKS in health and in disease. This role has been especially documented in the cardiovascular system and the kidney. Kinins are produced at slow rate in most organs in resting condition and/or inactivated quickly. Yet the KKS is involved in arterial function and in renal tubular function. In several pathological situations, kinin production increases, kinin receptor synthesis is upregulated, and kinins play an important role, whether beneficial or detrimental, in disease outcome. In the setting of ischemic, diabetic or hemodynamic aggression, kinin release by tissue kallikrein protects against organ damage, through B2 and/or B1 bradykinin receptor activation, depending on organ and disease. This has been well documented for the ischemic or diabetic heart, kidney and skeletal muscle, where KKS activity reduces oxidative stress, limits necrosis or fibrosis and promotes angiogenesis. On the other hand, in some pathological situations where plasma prekallikrein is inappropriately activated, excess kinin release in local or systemic circulation is detrimental, through oedema or hypotension. Putative therapeutic application of these clinical and experimental findings through current pharmacological development is discussed in the chapter. PMID:25130042

  9. Radiation Effect on Human Tissue

    NASA Technical Reports Server (NTRS)

    Richmond, Robert C.; Cruz, Angela; Bors, Karen; Curreri, Peter A. (Technical Monitor)

    2002-01-01

    Predicting the occurrence of human cancer following exposure of an epidemiologic population to any agent causing genetic damage is a difficult task. To an approximation, this is because the uncertainty of uniform exposure to the damaging agent, and the uncertainty of uniform processing of that damage within a complex set of biological variables, degrade the confidence of predicting the delayed expression of cancer as a relatively rare event within clinically normal individuals. This situation begs the need for alternate controlled experimental models that are predictive for the development of human cancer following exposures to agents causing genetic damage. Such models historically have not been of substantial proven value. It is more recently encouraging, however, that developments in molecular and cell biology have led to an expanded knowledge of human carcinogenesis, and of molecular markers associated with that process. It is therefore appropriate to consider new laboratory models developed to accomodate that expanded knowledge in order to assess the cancer risks associated with exposures to genotoxic agents. When ionizing radiation of space is the genotoxic agent, then a series of additional considerations for human cancer risk assessment must also be applied. These include the dose of radiation absorbed by tissue at different locations in the body, the quality of the absorbed radiation, the rate at which absorbed dose accumulates in tissue, the way in which absorbed dose is measured and calculated, and the alterations in incident radiation caused by shielding materials. It is clear that human cancer risk assessment for damage caused by ionizing radiation is a multidisciplinary responsibility, and that within this responsibility no single discipline can hold disproportionate sway if a risk assessment model of radiation-induced human cancer is to be developed that has proven value. Biomolecular and cellular markers from the work reported here are considered

  10. Plasma kallikrein-kinin system and diabetic retinopathy.

    PubMed

    Liu, Jia; Feener, Edward P

    2013-03-01

    Diabetic retinopathy (DR) occurs, to some extent, in most people with at least 20 years' duration of diabetes mellitus. The progression of DR to its sight-threatening stages is usually associated with the worsening of underlying retinal vascular dysfunction and disease. The plasma kallikrein-kinin system (KKS) is activated during vascular injury, where it mediates important functions in innate inflammation, blood flow, and coagulation. Recent findings from human vitreous proteomics and experimental studies on diabetic animal models have implicated the KKS in contributing to DR. Vitreous fluid from people with advanced stages of DR contains increased levels of plasma KKS components, including plasma kallikrein (PK), coagulation factor XII, and high-molecular-weight kininogen. Both bradykinin B1 and B2 receptor isoforms (B1R and B2R, respectively) are expressed in human retina, and retinal B1R levels are increased in diabetic rodents. The activation of the intraocular KKS induces retinal vascular permeability, vasodilation, and retinal thickening, and these responses are exacerbated in diabetic rats. Preclinical studies have shown that the administration of PK inhibitors and B1R antagonists to diabetic rats ameliorates retinal vascular hyperpermeability and inflammation. These findings suggest that components of plasma KKS are potential therapeutic targets for diabetic macular edema. PMID:23362193

  11. Potassium supplement upregulates the expression of renal kallikrein and bradykinin B2 receptor in SHR.

    PubMed

    Jin, L; Chao, L; Chao, J

    1999-03-01

    High potassium intake is known to attenuate hypertension, glomerular lesion, ischemic damage, and stroke-associated death. Our recent studies showed that expression of recombinant kallikrein by somatic gene delivery reduced high blood pressure, cardiac hypertrophy, and renal injury in hypertensive animal models. The aim of this study is to explore the potential role of the tissue kallikrein-kinin system in blood pressure reduction and renal protection in spontaneously hypertensive rats (SHR) on a high-potassium diet. Young SHR were given drinking water with or without 1% potassium chloride for 6 wk. Systolic blood pressure was significantly reduced beginning at 1 wk, and the effect lasted for 6 wk in the potassium-supplemented group compared with that in the control group. Potassium supplement induced 70 and 40% increases in urinary kallikrein levels and renal bradykinin B2 receptor density, respectively (P < 0.05), but did not change serum kininogen levels. Similarly, Northern blot analysis showed that renal kallikrein mRNA levels increased 2.7-fold, whereas hepatic kininogen mRNA levels remained unchanged in rats with high potassium intake. No difference was observed in beta-actin mRNA levels in the kidney or liver of either group. Competitive RT-PCR showed a 1.7-fold increase in renal bradykinin B2 receptor mRNA levels in rats with high potassium intake. Potassium supplement significantly increased water intake, urine excretion, urinary kinin, cAMP, and cGMP levels. This study suggests that upregulation of the tissue kallikrein-kinin system may be attributed, in part, to blood pressure-lowering and diuretic effects of high potassium intake.

  12. The kallikrein-kinin system in early state of diabetes.

    PubMed

    Vila, S B; Peluffo, V A; Alvarado, C; Cresto, J C; Zuccollo, A; Catanzaro, O L

    1992-01-01

    The kallikrein-kinin system was studied in 9 normals, healthy subjects (6 men, 3 women, age range 1 to 14 years) and 15 diabetic patients (9 men, 6 women age range 2 to 14 years) with an evolution of the disease between 1 to 14 years. Diabetic patients with low microalbuminuria (6.62 +/- 0.97 mg/24 h) show increased total and pre-kallikrein respect to control (3 and 2 fold respectively). On the other hand patients with high microalbuminuria (44.7 +/- 13.2 mg/24 h) show a total and pre-kallikrein of more than 4 and 8 fold increased respectively, compare with the control. According with these results we can concluded: 1) The total kallikrein and pre-kallikrein is increased in the diabetic state. 2) When microalbuminuria is high, the total and pre-kallikrein correlates with those increasing. 3) These changes could modified the renal hemodynamic in diabetes.

  13. Characterization of elements of the kallikrein-kinin system in rainbow trout (Salmo gairdneri)

    SciTech Connect

    Lipke, D.W.

    1988-01-01

    Elements of kallikrein-kinin system (KKS) are found in anamniotic vertebrates, however, research has failed to substantiate the presence of a KKS in these animals. This research was conducted to verify the presence of elements of the KKS in rainbow trout. Trout branchial angiotensin-converting enzyme (ACE, kininase II) was similar to mammalian ACE in many physical, chemical and kinetic parameters. Angiotensin-converting enzyme-like activity (ACELA) was also demonstrated in the most primitive vertebrates. ACELA was consistently found in vertebrate respiratory tissues and was prevalent in salt and water exchanging organs. /sup 3/H-bradykinin, perfused through the trout gill was not metabolized, as demonstrated by high voltage paper electrophoresis. However, trout gills extracted and retained approximately 40% of the /sup 3/H-bradykinin perfused into the branchial vasculature. Gill tissue homogenates metabolized both /sup 3/H-bradykinin and the vasoactive substance produced by incubating trout plasma with glandular kallikrein.

  14. Human dignity and human tissue: a meaningful ethical relationship?

    PubMed

    Kirchhoffer, David G; Dierickx, Kris

    2011-09-01

    Human dignity has long been used as a foundational principle in policy documents and ethical guidelines intended to govern various forms of biomedical research. Despite the vast amount of literature concerning human dignity and embryonic tissues, the majority of biomedical research uses non-embryonic human tissue. Therefore, this contribution addresses a notable lacuna in the literature: the relationship, if any, between human dignity and human tissue. This paper first elaborates a multidimensional understanding of human dignity that overcomes many of the shortcomings associated with the use of human dignity in other ethical debates. Second, it discusses the relationship between such an understanding of human dignity and 'non-embryonic' human tissue. Finally, it considers the implications of this relationship for biomedical research and practice involving human tissue. The contribution demonstrates that while human tissue cannot be said to have human dignity, human dignity is nevertheless implicated by human tissue, making what is done with human tissue and how it is done worthy of moral consideration.

  15. Sustainable three-dimensional tissue model of human adipose tissue.

    PubMed

    Bellas, Evangelia; Marra, Kacey G; Kaplan, David L

    2013-10-01

    The need for physiologically relevant sustainable human adipose tissue models is crucial for understanding tissue development, disease progression, in vitro drug development and soft tissue regeneration. The coculture of adipocytes differentiated from human adipose-derived stem cells, with endothelial cells, on porous silk protein matrices for at least 6 months is reported, while maintaining adipose-like outcomes. Cultures were assessed for structure and morphology (Oil Red O content and CD31 expression), metabolic functions (leptin, glycerol production, gene expression for GLUT4, and PPARγ) and cell replication (DNA content). The cocultures maintained size and shape over this extended period in static cultures, while increasing in diameter by 12.5% in spinner flask culture. Spinner flask cultures yielded improved adipose tissue outcomes overall, based on structure and function, when compared to the static cultures. This work establishes a tissue model system that can be applied to the development of chronic metabolic dysfunction systems associated with human adipose tissue, such as obesity and diabetes, due to the long term sustainable functions demonstrated here.

  16. Synchronous luminescence spectroscopy of human breast tissues

    NASA Astrophysics Data System (ADS)

    Majumdar, S. K.; Gupta, P. K.

    1998-06-01

    We report, to our knowledge, the first use of synchronous luminescence (SL) spectroscopy for autofluorescence diagnosis of cancer. The spectral narrowing effect of the SL spectroscopy led to an easier identification of the different fluorophores present in human breast tissues and provided relative estimate of their concentration in qualitative agreement with the estimates obtained from conventional excitation and emission spectroscopy. Further, the SL spectra from human breast tissues could discriminate cancerous tissues from benign tumors and normal tissues with a sensitivity and specificity of 100% in a study involving 34 patients with breast tumor (19 ductal carcinomas and 15 fibroadenomas).

  17. TiO2 nanoparticles tested in a novel screening whole human blood model of toxicity trigger adverse activation of the kallikrein system at low concentrations.

    PubMed

    Ekstrand-Hammarström, Barbro; Hong, Jaan; Davoodpour, Padideh; Sandholm, Kerstin; Ekdahl, Kristina N; Bucht, Anders; Nilsson, Bo

    2015-05-01

    There is a compelling need to understand and assess the toxicity of industrially produced nanoparticles (NPs). In order to appreciate the long-term effects of NPs, sensitive human-based screening tests that comprehensively map the NP properties are needed to detect possible toxic mechanisms. Animal models can only be used in a limited number of test applications and are subject to ethical concerns, and the interpretation of experiments in animals is also distorted by the species differences. Here, we present a novel easy-to-perform highly sensitive whole-blood model using fresh non-anticoagulated human blood, which most justly reflects complex biological cross talks in a human system. As a demonstrator of the tests versatility, we evaluated the toxicity of TiO2 NPs that are widely used in various applications and otherwise considered to have relatively low toxic properties. We show that TiO2 NPs at very low concentrations (50 ng/mL) induce strong activation of the contact system, which in this model elicits thromboinflammation. These data are in line with the finding of components of the contact system in the protein corona of the TiO2 NPs after exposure to blood. The contact system activation may lead to both thrombotic reactions and generation of bradykinin, thereby representing fuel for chronic inflammation in vivo and potentially long-term risk of autoimmunity, arteriosclerosis and cancer. These results support the notion that this novel whole-blood model represents an important contribution to testing of NP toxicity.

  18. TiO2 nanoparticles tested in a novel screening whole human blood model of toxicity trigger adverse activation of the kallikrein system at low concentrations.

    PubMed

    Ekstrand-Hammarström, Barbro; Hong, Jaan; Davoodpour, Padideh; Sandholm, Kerstin; Ekdahl, Kristina N; Bucht, Anders; Nilsson, Bo

    2015-05-01

    There is a compelling need to understand and assess the toxicity of industrially produced nanoparticles (NPs). In order to appreciate the long-term effects of NPs, sensitive human-based screening tests that comprehensively map the NP properties are needed to detect possible toxic mechanisms. Animal models can only be used in a limited number of test applications and are subject to ethical concerns, and the interpretation of experiments in animals is also distorted by the species differences. Here, we present a novel easy-to-perform highly sensitive whole-blood model using fresh non-anticoagulated human blood, which most justly reflects complex biological cross talks in a human system. As a demonstrator of the tests versatility, we evaluated the toxicity of TiO2 NPs that are widely used in various applications and otherwise considered to have relatively low toxic properties. We show that TiO2 NPs at very low concentrations (50 ng/mL) induce strong activation of the contact system, which in this model elicits thromboinflammation. These data are in line with the finding of components of the contact system in the protein corona of the TiO2 NPs after exposure to blood. The contact system activation may lead to both thrombotic reactions and generation of bradykinin, thereby representing fuel for chronic inflammation in vivo and potentially long-term risk of autoimmunity, arteriosclerosis and cancer. These results support the notion that this novel whole-blood model represents an important contribution to testing of NP toxicity. PMID:25770998

  19. Tissue Specificity of Human Disease Module

    PubMed Central

    Kitsak, Maksim; Sharma, Amitabh; Menche, Jörg; Guney, Emre; Ghiassian, Susan Dina; Loscalzo, Joseph; Barabási, Albert-László

    2016-01-01

    Genes carrying mutations associated with genetic diseases are present in all human cells; yet, clinical manifestations of genetic diseases are usually highly tissue-specific. Although some disease genes are expressed only in selected tissues, the expression patterns of disease genes alone cannot explain the observed tissue specificity of human diseases. Here we hypothesize that for a disease to manifest itself in a particular tissue, a whole functional subnetwork of genes (disease module) needs to be expressed in that tissue. Driven by this hypothesis, we conducted a systematic study of the expression patterns of disease genes within the human interactome. We find that genes expressed in a specific tissue tend to be localized in the same neighborhood of the interactome. By contrast, genes expressed in different tissues are segregated in distinct network neighborhoods. Most important, we show that it is the integrity and the completeness of the expression of the disease module that determines disease manifestation in selected tissues. This approach allows us to construct a disease-tissue network that confirms known and predicts unexpected disease-tissue associations. PMID:27748412

  20. Human Postmortem Tissue: What Quality Markers Matter?

    PubMed Central

    Stan, Ana D.; Ghose, Subroto; Gao, Xue-Min; Roberts, Rosalinda C.; Lewis-Amezcua, Kelly; Hatanpaa, Kimmo J.; Tamminga, Carol A.

    2007-01-01

    Post mortem human brain tissue is used for the study of many different brain diseases. A key factor in conducting postmortem research is the quality of the tissue. Unlike animal tissue, whose condition at death can be controlled and influenced, human tissue can only be collected naturalistically. This introduces potential confounds, based both on pre- and postmortem conditions, that may influence the quality of tissue and its ability to yield accurate results. The traditionally recognized confounds that reduce tissue quality are agonal factors (e.g., coma, hypoxia, hyperpyrexia at the time of death), and long postmortem interval (PMI). We measured tissue quality parameters in over 100 postmortem cases collected from different sources and correlated them with RNA quality (as indicated by the RNA Integrity Number (RIN)) and with protein quality (as measured by the level of representative proteins). Our results show that the most sensible indicator of tissue quality is RIN and that there is a good correlation between RIN and the pH. No correlation developed between protein levels and the aforementioned factors. Moreover, even when RNA was degraded, the protein levels remained stable. However, these correlations did not prove true under all circumstances (e.g. thawed tissue, surgical tissue), that yielded unexpected quality indicators. These data also suggest that cases whose source was a Medical Examiner’s office represent high tissue quality. PMID:17045977

  1. Grating-based tomography of human tissues

    NASA Astrophysics Data System (ADS)

    Müller, Bert; Schulz, Georg; Mehlin, Andrea; Herzen, Julia; Lang, Sabrina; Holme, Margaret; Zanette, Irene; Hieber, Simone; Deyhle, Hans; Beckmann, Felix; Pfeiffer, Franz; Weitkamp, Timm

    2012-07-01

    The development of therapies to improve our health requires a detailed knowledge on the anatomy of soft tissues from the human body down to the cellular level. Grating-based phase contrast micro computed tomography using synchrotron radiation provides a sensitivity, which allows visualizing micrometer size anatomical features in soft tissue without applying any contrast agent. We show phase contrast tomography data of human brain, tumor vessels and constricted arteries from the beamline ID 19 (ESRF) and urethral tissue from the beamline W2 (HASYLAB/DESY) with micrometer resolution. Here, we demonstrate that anatomical features can be identified within brain tissue as well known from histology. Using human urethral tissue, the application of two photon energies is compared. Tumor vessels thicker than 20 μm can be perfectly segmented. The morphology of coronary arteries can be better extracted in formalin than after paraffin embedding.

  2. Copy Number Variants in the Kallikrein Gene Cluster

    PubMed Central

    Lindahl, Pernilla; Säll, Torbjörn; Bjartell, Anders; Johansson, Anna M.; Lilja, Hans; Halldén, Christer

    2013-01-01

    The kallikrein gene family (KLK1-KLK15) is the largest contiguous group of protease genes within the human genome and is associated with both risk and outcome of cancer and other diseases. We searched for copy number variants in all KLK genes using quantitative PCR analysis and analysis of inheritance patterns of single nucleotide polymorphisms. Two deletions were identified: one 2235-bp deletion in KLK9 present in 1.2% of alleles, and one 3394-bp deletion in KLK15 present in 4.0% of alleles. Each deletion eliminated one complete exon and created out-of-frame coding that eliminated the catalytic triad of the resulting truncated gene product, which therefore likely is a non-functional protein. Deletion breakpoints identified by DNA sequencing located the KLK9 deletion breakpoint to a long interspersed element (LINE) repeated sequence, while the deletion in KLK15 is located in a single copy sequence. To search for an association between each deletion and risk of prostate cancer (PC), we analyzed a cohort of 667 biopsied men (266 PC cases and 401 men with no evidence of PC at biopsy) using short deletion-specific PCR assays. There was no association between evidence of PC in this cohort and the presence of either gene deletion. Haplotyping revealed a single origin of each deletion, with most recent common ancestor estimates of 3000-8000 and 6000-14 000 years for the deletions in KLK9 and KLK15, respectively. The presence of the deletions on the same haplotypes in 1000 Genomes data of both European and African populations indicate an early origin of both deletions. The old age in combination with homozygous presence of loss-of-function variants suggests that some kallikrein-related peptidases have non-essential functions. PMID:23894413

  3. NCI’s Cooperative Human Tissue Network

    Cancer.gov

    Quality biospecimens are a foundational resource for cancer research. One of NCI’s longest running biospecimen programs is the Cooperative Human Tissue Network, a resource mainly for basic discovery and early translational research.

  4. Involvement of Kallikrein-Related Peptidases in Normal and Pathologic Processes

    PubMed Central

    Stefanini, Ana Carolina B.; da Cunha, Bianca Rodrigues; Henrique, Tiago; Tajara, Eloiza H.

    2015-01-01

    Human kallikrein-related peptidases (KLKs) are a subgroup of serine proteases that participate in proteolytic pathways and control protein levels in normal physiology as well as in several pathological conditions. Their complex network of stimulatory and inhibitory interactions may induce inflammatory and immune responses and contribute to the neoplastic phenotype through the regulation of several cellular processes, such as proliferation, survival, migration, and invasion. This family of proteases, which includes one of the most useful cancer biomarkers, kallikrein-related peptidase 3 or PSA, also has a protective effect against cancer promoting apoptosis or counteracting angiogenesis and cell proliferation. Therefore, they represent attractive therapeutic targets and may have important applications in clinical oncology. Despite being intensively studied, many gaps in our knowledge on several molecular aspects of KLK functions still exist. This review aims to summarize recent data on their involvement in different processes related to health and disease, in particular those directly or indirectly linked to the neoplastic process. PMID:26783378

  5. Variation in alternative splicing across human tissues

    PubMed Central

    Yeo, Gene; Holste, Dirk; Kreiman, Gabriel; Burge, Christopher B

    2004-01-01

    Background Alternative pre-mRNA splicing (AS) is widely used by higher eukaryotes to generate different protein isoforms in specific cell or tissue types. To compare AS events across human tissues, we analyzed the splicing patterns of genomically aligned expressed sequence tags (ESTs) derived from libraries of cDNAs from different tissues. Results Controlling for differences in EST coverage among tissues, we found that the brain and testis had the highest levels of exon skipping. The most pronounced differences between tissues were seen for the frequencies of alternative 3' splice site and alternative 5' splice site usage, which were about 50 to 100% higher in the liver than in any other human tissue studied. Quantifying differences in splice junction usage, the brain, pancreas, liver and the peripheral nervous system had the most distinctive patterns of AS. Analysis of available microarray expression data showed that the liver had the most divergent pattern of expression of serine-arginine protein and heterogeneous ribonucleoprotein genes compared to the other human tissues studied, possibly contributing to the unusually high frequency of alternative splice site usage seen in liver. Sequence motifs enriched in alternative exons in genes expressed in the brain, testis and liver suggest specific splicing factors that may be important in AS regulation in these tissues. Conclusions This study distinguishes the human brain, testis and liver as having unusually high levels of AS, highlights differences in the types of AS occurring commonly in different tissues, and identifies candidate cis-regulatory elements and trans-acting factors likely to have important roles in tissue-specific AS in human cells. PMID:15461793

  6. Melanin content of hamster tissues, human tissues, and various melanomas

    SciTech Connect

    Watts, K.P.; Fairchild, R.G.; Slatkin, D.N.; Greenberg, D.; Packer, S.; Atkins, H.L.; Hannon, S.J.

    1981-02-01

    Melanin content (percentage by weight) was determined in both pigmented and nonpigmented tissues of Syrian golden hamsters bearing Greene melanoma. Melanin content was also measured in various other melanoma models (B-16 in C57 mice, Harding-Passey in BALB/c mice, and KHDD in C3H mice) and in nine human melanomas, as well as in selected normal tissues. The purpose was to evaluate the possible efficacy of chlorpromazine, which is known to bind to melanin, as a vehicle for boron transport in neutron capture therapy. Successful therapy would depend upon selective uptake and absolute concentration of borated compounds in tumors; these parameters will in turn depend upon melanin concentration in melanomas and nonpigmented ''background'' tissues. Hamster whole eyes, hamster melanomas, and other well-pigmented animal melanomas were found to contain 0.3 to 0.8% melanin by weight, whereas human melanomas varied from 0.1 to 0.9% (average, 0.35%). Other tissues, with the exception of skin, were lower in content by a factor of greater than or equal to30. Melanin pigment was extracted from tissues, and the melanin content was determined spectrophotometrically. Measurements were found to be sensitive to the presence of other proteins. Previous procedures for isolating and quantifying melanin often neglected the importance of removing proteins and other interfering nonmelanic substances.

  7. Epidermal growth factor (urogastrone) in human tissues.

    PubMed

    Hirata, Y; Orth, D N

    1979-04-01

    Human epidermal growth factor (hEGF), which stimulates the growth of a variety of tissues, was first isolated from mouse submandibular glands, but is also excreted in large amounts (about 50 micrograms/day) in human urine and is probably identical to human beta-urogastrone (hUG), a potent inhibitor of stimulated gastric acid secretion. However, the primary tissue source of hEGF/hUG is as yet unknown. The hEGF/hUG in homogenates of human salivary glands and a wide variety of other endocrine and nonendocrine tissues was extracted by Amberlite CG-50 cation exchange chromatography and immune affinity chromatography using the immunoglobulin fraction of rabbit anti-hEGF serum covalently bound to agarose. The extracts were subjected to homologous hEGF RIA. Immunoreactive hEGF was found in extracts of adult submandibular gland, thyroid gland, duodenum, jejunum, and kidney, but not in several fetal tissues. The tissue immunoreactive hEGF was similar to standard hEGF in terms of immunoreactivity and elution from Sephadex G-50 Fine resin, but its concentrations were very low (1.3-5.5 ng/g wet tissue). Thus, it is not certain that these tissues represent the only source of the large amounts of hEGF/hUG that appear to be filtered by the kidneys each day.

  8. Pre-stimulation of the kallikrein system in cisplatin-induced acute renal injury: An approach to renoprotection

    SciTech Connect

    Aburto, Andrés; Barría, Agustín; Cárdenas, Areli; Carpio, Daniel; Figueroa, Carlos D.; Burgos, Maria E.; Ardiles, Leopoldo

    2014-10-15

    Antineoplastic treatment with cisplatin is frequently complicated by nephrotoxicity. Although oxidative stress may be involved, the pathogenic mechanisms responsible for renal damage have not been completely clarified. In order to investigate the role of the renal kinin system in this condition, a group of rats was submitted to high potassium diet to stimulate the synthesis and excretion of tissue kallikrein 1 (rKLK1) previous to an intraperitoneal injection of 7 mg/kg cisplatin. A significant reduction in lipoperoxidation, evidenced by urinary excretion of malondialdehyde and renal immunostaining of hidroxy-nonenal, was accompanied by a decline in apoptosis. Coincident with these findings we observed a reduction in the expression of renal KIM-1 suggesting that renoprotection may be occurring. Stimulation or indemnity of the renal kinin system deserves to be evaluated as a complementary pharmacological measure to diminish cisplatin nephrotoxicity. - Highlights: • Mechanisms of cisplatin-induced-renal damage have not been completely clarified. • Cisplatin induces oxidative stress and apoptosis. • The renal kallikrein-kinin system is protective in experimental acute renal damage. • Kallikrein stimulation reduces oxidative stress and apoptosis induced by cisplatin. • Protection of the kallikrein-kinin system may reduce cisplatin toxicity.

  9. Human histocultures (tissue explants) in retrovirology

    PubMed Central

    Arakelyan, Anush; Fitzgerald, Wendy; Grivel, Jean-Charles; Vanpouille, Christophe; Margolis, Leonid

    2014-01-01

    Summary Viral pathogenesis is studied predominantly in cultures of primary isolated cells or cell lines. Many retroviruses efficiently replicate only in activated cells. Therefore, in order to become efficient viral producers cells should be artificially activated, a procedure which significantly changes cell physiology. However, for many viral diseases, like HIV-1 and other retroviruses’ diseases, critical pathogenic events occur in tissues and cell isolation from their native microenvironment prevents single cell cultures from faithfully reflecting important aspects of cell-cell and cell-pathogen interactions that occur in the context of complex tissue cytoarchitecture. Tissue explants (histocultures) that retain tissue cytoarchitecture and many aspects of cell-cell interactions more faithfully represent in vivo tissue features. Human histocultures constitute an adequate model for studying viral pathogenesis under controlled laboratory conditions. Protocols for various human histocultures as applied to study retroviral pathogenesis, in particular of HIV-1, have been refined by our laboratory and are described in the present publication. Human histocultures of human tonsils and lymph nodes, as well as of recto-sigmoid and cervico-vaginal tissues can be used to study viral transmission, pathogenesis and as a pre-clinical platform for antivirals evaluation. PMID:24158827

  10. Demand for human allograft tissue in Canada.

    PubMed

    Lakey, Jonathan R T; Mirbolooki, Mohammadreza; Rogers, Christina; Mohr, Jim

    2007-01-01

    There is relatively little known about the demand for allograft tissues in Canada. The Canadian Council for Donation and Transplantation (CCDT) is a national advisory body that undertook a comprehensive "market survey" to estimate surgical demand for human allograft tissues in Canada. The report "Demand for Human Allograft Tissue in Canada" reflects survey results sent to 5 prominent User Groups. User Groups were identified as orthopaedic surgeons; neurosurgeons; corneal transplant surgeons; plastic surgeons, specifically those at Canadian Burn Units; and cardiac surgeons (adult and paediatric surgery). The demand for allograft grafts was determined and then extrapolated across the total User Group and then increases in allograft tissue use over the next 1-2 years across User Groups were predicted. The overall response rate for the survey was 21.4%. It varied from a low of 19.6% for the orthopaedic survey to a high of 40.5% for the corneal survey. The estimated current demand for allograft tissue in Canada ranges from a low of 34,442 grafts per year to a high of 62,098 grafts per year. The predicted increase in use of allograft tissue over the next 1-2 year period would suggest that annual demand could rise to somewhere in the range of 42,589-72,210 grafts. The highest rated preferences (98% and 94%) were for accredited and Canadian tissue banks, respectively. This study represents a key step in addressing the paucity of information concerning the demand for allograft tissue in Canada.

  11. Kallikrein-related peptidases in cancers of gastrointestinal tract: an inside view of their role and clinical significance.

    PubMed

    Linardoutsos, Dimitrios; Gazouli, Maria; Machairas, Anastasios; Bramis, Ioannis; Zografos, Georgios C

    2014-01-01

    Human tissue kallikrein (KLK1) and is related peptidases (KLK2-KLK15) are a family of 15 homologous serine proteases, participating in numerous processes of normal physiology. Considering the irreversible impact of proteases on substrates, the tissue-dependent regulation of KLKs activity becomes crucial for their beneficial role in normal homeostasis. Moreover, KLKs expression is strongly regulated at the transcriptional and post-transcriptional level by steroid hormones and miRNAs, respectively. Deregulation of KLKs expression, secretion and/or activation has been observed in most human malignancies and there is a trend to identify their role in the multi-complex process of cancer development. The identification of extracellular matrix (ECM) proteins, cell-surface receptors, cell-surface adhesion molecules and growth factors among substrates, clearly support the driving role of KLK abnormal expression and function during tumorigenesis and cancer progression. KLKs have also clinical utility in cancer diagnosis and monitoring like KLK 3 (PSA) in prostate cancer. In this review, we tried to summarize the existing literature about the role of KLKs in gastrointestinal cancers as well as to emphasize their clinical significance for patients' prognosis.

  12. Extensive genetic variation in somatic human tissues.

    PubMed

    O'Huallachain, Maeve; Karczewski, Konrad J; Weissman, Sherman M; Urban, Alexander Eckehart; Snyder, Michael P

    2012-10-30

    Genetic variation between individuals has been extensively investigated, but differences between tissues within individuals are far less understood. It is commonly assumed that all healthy cells that arise from the same zygote possess the same genomic content, with a few known exceptions in the immune system and germ line. However, a growing body of evidence shows that genomic variation exists between differentiated tissues. We investigated the scope of somatic genomic variation between tissues within humans. Analysis of copy number variation by high-resolution array-comparative genomic hybridization in diverse tissues from six unrelated subjects reveals a significant number of intraindividual genomic changes between tissues. Many (79%) of these events affect genes. Our results have important consequences for understanding normal genetic and phenotypic variation within individuals, and they have significant implications for both the etiology of genetic diseases such as cancer and for immortalized cell lines that might be used in research and therapeutics.

  13. Humanized mice with ectopic artificial liver tissues

    PubMed Central

    Chen, Alice A.; Thomas, David K.; Ong, Luvena L.; Schwartz, Robert E.; Golub, Todd R.; Bhatia, Sangeeta N.

    2011-01-01

    “Humanized” mice offer a window into aspects of human physiology that are otherwise inaccessible. The best available methods for liver humanization rely on cell transplantation into immunodeficient mice with liver injury but these methods have not gained widespread use due to the duration and variability of hepatocyte repopulation. In light of the significant progress that has been achieved in clinical cell transplantation through tissue engineering, we sought to develop a humanized mouse model based on the facile and ectopic implantation of a tissue-engineered human liver. These human ectopic artificial livers (HEALs) stabilize the function of cryopreserved primary human hepatocytes through juxtacrine and paracrine signals in polymeric scaffolds. In contrast to current methods, HEALs can be efficiently established in immunocompetent mice with normal liver function. Mice transplanted with HEALs exhibit humanized liver functions persistent for weeks, including synthesis of human proteins, human drug metabolism, drug–drug interaction, and drug-induced liver injury. Here, mice with HEALs are used to predict the disproportionate metabolism and toxicity of “major” human metabolites using multiple routes of administration and monitoring. These advances may enable manufacturing of reproducible in vivo models for diverse drug development and research applications. PMID:21746904

  14. Dolichol and dolichyl phosphate in human tissues.

    PubMed Central

    Tollbom, O.; Dallner, G.

    1986-01-01

    The content of dolichol and dolichyl phosphate in various human organs was analysed using autopsy samples. The reliability of these measurements was demonstrated by comparison with values for fresh biopsy material. Dolichol was present in all tissues investigated and the content was highest in the adrenal gland, pancreas, pituitary gland, testis and thyroid gland, ranging between 1.5 and 7.1 mg/g tissue. Dolichyl-P was detected in the various organs in highly variable amounts, ranging between 1 and 9% of the total dolichol content. While the main pattern of isoprene composition for dolichol and dolichyl-P was similar in individual organs, some variation was observed between tissues. Dolichol represents the largest lipid component in the pituitary gland, exceeding the total phospholipid content. The high concentrations of dolichol and dolichyl-P in human organs indicate that these lipids may play important roles in physiological and pathological cellular functions. PMID:3641633

  15. Lubricin in human breast tissue expander capsules.

    PubMed

    Cheriyan, Thomas; Guo, Lifei; Orgill, Dennis P; Padera, Robert F; Schmid, Thomas M; Spector, Myron

    2012-10-01

    Capsular contraction is the most common complication of breast reconstruction surgery. While presence of the contractile protein alpha smooth muscle actin (α-SMA) is considered among the causes of capsular contraction, the exact etiology and pathophysiology is not fully understood. The objective of this study was to investigate the possible role of lubricin in capsular formation and contraction by determining the presence and distribution of the lubricating protein lubricin in human breast tissue expander capsules. Related aims were to evaluate select histopathologic features of the capsules, and the percentage of cells expressing α-SMA, which reflects the myofibroblast phenotype. Capsules from tissue expanders were obtained from eight patients. Lubricin, at the tissue-implant interface, in the extracellular matrix, and in cells, and α-SMA-containing cells were evaluated immunohistochemically. The notable finding was that lubricin was identified in all tissue expander capsules: as a discrete layer at the tissue-implant interface, extracellular, and intracellular. There was a greater amount of lubricin in the extracellular matrix in the intimal-subintimal zone when compared with the tissue away from the implant. Varying degrees of synovial metaplasia were seen at the tissue-implant interface. α-SMA-containing cells were also seen in all but one patient. The findings might help us better understand factors involved in capsule formation.

  16. Initiation of contact system activation in plasma is dependent on factor XII autoactivation and not on enhanced susceptibility of factor XII for kallikrein cleavage.

    PubMed

    Citarella, F; Wuillemin, W A; Lubbers, Y T; Hack, C E

    1997-10-01

    Various mechanisms have been hypothesized to explain the initiation of contact system activation in plasma. We investigated the capability of dextran sulphate (DS) of different molecular weights to initiate contact system activation in normal human plasma, and compared this with their capability to support factor XII autoactivation and to enhance factor XII susceptibility for cleavage by kallikrein. Dextran sulphate of Mr 500,000 (DS500) and 50,000 (DS50) was able to initiate contact system activation in plasma (determined by measuring the amount of factor XIIa-C1-inhibitor, kallikrein-C1-inhibitor and factor XIa-C1-inhibitor complexes generated) as well as to support factor XII autoactivation and to enhance factor XII susceptibility for cleavage by kallikrein (as measured with amidolytic assays using purified proteins). In contrast, dextran sulphate of Mr 15,000 (DS15) and 5000 (DS5) neither induced contact system activation in plasma, nor supported autoactivation of factor XII, although both of these DS species enhanced the rate of activation of factor XII by kallikrein in the purified system. Based on these properties (i.e. binding of factor XII without inducing autoactivation), DS15 and DS5 were predicted to be inhibitors of contact system activation induced in plasma by DS500, which indeed was observed. We conclude that enhanced factor XII susceptibility for kallikrein activation and factor XII autoactivation are distinct phenomena, the latter being necessary to support activation of the contact system in plasma.

  17. Alpha-dispersion in human tissue

    NASA Astrophysics Data System (ADS)

    Grimnes, Sverre; Martinsen, Ørjan G.

    2010-04-01

    Beta dispersion is found in living tissue in the kilohertz - megahertz range and is caused by the cellular structure of biological materials with low frequency properties caused by cell membranes. Alpha dispersion is found in the hertz range and the causes are not so well known. Alpha dispersions are the first to disappear when tissue dies. Tissue data have often been based upon excised specimen from animals and are therefore not necessarily representative for human tissue alpha dispersions. Here we present data obtained with non-invasive skin surface electrodes for different segments of the living human body. We found alpha dispersions in all cases; the ankle-wrist results had the smallest. Large alpha dispersions were found where the distance between the electrodes and muscle masses was small, e.g. on the calf. Further studies on electrode technique and reciprocity, electrode positioning, statistical variations, gender, age and bodily constitutions are necessary in order to reveal more about the alpha dispersion, its appearance and disappearance.

  18. Tissue microarray profiling in human heart failure.

    PubMed

    Lal, Sean; Nguyen, Lisa; Tezone, Rhenan; Ponten, Fredrik; Odeberg, Jacob; Li, Amy; Dos Remedios, Cristobal

    2016-09-01

    Tissue MicroArrays (TMAs) are a versatile tool for high-throughput protein screening, allowing qualitative analysis of a large number of samples on a single slide. We have developed a customizable TMA system that uniquely utilizes cryopreserved human cardiac samples from both heart failure and donor patients to produce formalin-fixed paraffin-embedded sections. Confirmatory upstream or downstream molecular studies can then be performed on the same (biobanked) cryopreserved tissue. In a pilot study, we applied our TMAs to screen for the expression of four-and-a-half LIM-domain 2 (FHL2), a member of the four-and-a-half LIM family. This protein has been implicated in the pathogenesis of heart failure in a variety of animal models. While FHL2 is abundant in the heart, not much is known about its expression in human heart failure. For this purpose, we generated an affinity-purified rabbit polyclonal anti-human FHL2 antibody. Our TMAs allowed high-throughput profiling of FHL2 protein using qualitative and semiquantitative immunohistochemistry that proved complementary to Western blot analysis. We demonstrated a significant relative reduction in FHL2 protein expression across different forms of human heart failure.

  19. Monoclonal antibody F1 binds to the kringle domain of factor XII and induces enhanced susceptibility for cleavage by kallikrein.

    PubMed

    Ravon, D M; Citarella, F; Lubbers, Y T; Pascucci, B; Hack, C E

    1995-12-01

    In a previous study we have shown that monoclonal antibody F1 (MoAb F1), directed against an epitope on the heavy chain of factor XII distinct from the binding site for anionic surfaces, is able to activate factor XII in plasma (Nuijens JH, et al: J Biol Chem 264; 12941, 1989). Here, we studied in detail the mechanism underlying the activation of factor XII by MoAb F1 using purified proteins. Formation of factor XIIa was assessed by measuring its amidolytic activity towards the chromogenic substrate H-D-Pro-Phe-Arg-pNA (S-2302) in the presence of soybean trypsin inhibitor and by assessing cleavage on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Upon incubation with MoAb F1 alone, factor XII was auto-activated in a time-dependent fashion, activation being maximal after 30 hours. Factor XII incubated in the absence of MoAb F1 was hardly activated by kallikrein, whereas in the presence of MoAb F1, but not in that of a control MoAb, the rate of factor XII activation by kallikrein was promoted at least 60-fold. Maximal activation of factor XII with kallikrein in the presence of MoAb F1 was reached within 1 hour. This effect of kallikrein on the cleavage of factor XII bound to MoAb F1 was specific because the fibrinolytic enzymes plasmin, urokinase, and tissue-type plasminogen activator could not substitute for kallikrein. Also, trypsin could easily activate factor XII, but in contrast to kallikrein, this activation was independent of MoAb F1. SDS-PAGE analysis showed that the appearance of amidolytic activity correlated well with cleavage of factor XII. MoAb F1-induced activation of factor XII in this purified system was not dependent on the presence of high-molecular-weight kininogen (HK), in contrast to the activation of the contact system in plasma by MoAb F1. Experiments with deletion mutants revealed that the epitopic region for MoAb F1 on factor XII is located on the kringle domain. Thus, this study shows that binding of ligands to the

  20. Kallikrein-related peptidase 8 is expressed in myocardium and induces cardiac hypertrophy

    PubMed Central

    Cao, Buqing; Yu, Qing; Zhao, Wei; Tang, Zhiping; Cong, Binghai; Du, Jiankui; Lu, Jianqiang; Zhu, Xiaoyan; Ni, Xin

    2016-01-01

    The tissue kallikrein-related peptidase family (KLK) is a group of trypsin- and chymotrypsin-like serine proteases that share a similar homology to parent tissue kallikrein (KLK1). KLK1 is identified in heart and has anti-hypertrophic effects. However, whether other KLK family members play a role in regulating cardiac function remains unknown. In the present study, we demonstrated for the first time that KLK8 was expressed in myocardium. KLK8 expression was upregulated in left ventricle of cardiac hypertrophy models. Both intra-cardiac adenovirus-mediated and transgenic-mediated KLK8 overexpression led to cardiac hypertrophy in vivo. In primary neonatal rat cardiomyocytes, KLK8 knockdown inhibited phenylephrine (PE)-induced cardiomyocyte hypertrophy, whereas KLK8 overexpression promoted cardiomyocyte hypertrophy via a serine protease activity-dependent but kinin receptor-independent pathway. KLK8 overexpression increased epidermal growth factor (EGF) production, which was blocked by the inhibitors of serine protease. EGF receptor (EGFR) antagonist and EGFR knockdown reversed the hypertrophy induced by KLK8 overexpression. KLK8-induced cardiomyocyte hypertrophy was also significantly decreased by blocking the protease-activated receptor 1 (PAR1) or PAR2 pathway. Our data suggest that KLK8 may promote cardiomyocyte hypertrophy through EGF signaling- and PARs-dependent but a kinin receptor-independent pathway. It is implied that different KLK family members can subtly regulate cardiac function and remodeling. PMID:26823023

  1. Tissue pharmacokinetics of levofloxacin in human soft tissue infections

    PubMed Central

    Bellmann, Romuald; Kuchling, Gerald; Dehghanyar, Pejman; Zeitlinger, Markus; Minar, Erich; Mayer, Bernhard X; Müller, Markus; Joukhadar, Christian

    2004-01-01

    Aims The present study addressed the ability of levofloxacin to penetrate into subcutaneous adipose tissues in patients with soft tissue infection. Methods Tissue concentrations of levofloxacin in inflamed and healthy subcutaneous adipose tissue were measured in six patients by microdialysis after administration of a single intravenous dose of 500 mg. Levofloxacin was assayed by high-performance liquid chromatography. Results The mean concentration vs time profile of free levofloxacin in plasma was identical to that in inflamed and healthy tissues. The ratios of the mean area under the free levofloxacin concentration vs time curve from 0 to 10 h (AUC(0,10 h)) in tissue to that in plasma were 1.2 ± 1.0 for inflamed and 1.1 ± 0.6 for healthy subcutaneous adipose tissue (mean ± SD). The mean difference in the ratio of the AUCtissue : AUCplasma for inflamed and healthy tissue was 0.09 (95% confidence interval −0.58, 0.759, P > 0.05). Interindividual variability in tissue penetration was high, as indicated by a coefficient of variation of approximately 82% for AUCtissue : AUCplasma ratios. Conclusions The penetration of levofloxacin into tissue appears to be unaffected by local inflammation. Our plasma and tissue data suggest that an intravenous dose of 500 mg levofloxacin provides effective antibacterial concentrations at the target site. However, in treatment resistant patients, tissue concentrations may be sub-therapeutic. PMID:15089808

  2. Hippocampus and epilepsy: Findings from human tissues.

    PubMed

    Huberfeld, G; Blauwblomme, T; Miles, R

    2015-03-01

    Surgical removal of the epileptogenic zone provides an effective therapy for several focal epileptic syndromes. This surgery offers the opportunity to study pathological activity in living human tissue for pharmacoresistant partial epilepsy syndromes including temporal lobe epilepsies with hippocampal sclerosis, cortical dysplasias, epilepsies associated with tumors and developmental malformations. Slices of tissue from patients with these syndromes retain functional neuronal networks and may generate epileptic activities. The properties of cells in this tissue may not be greatly changed, but excitatory synaptic transmission is often enhanced and GABAergic inhibition is preserved. Typically epileptic activity is not generated spontaneously by the neocortex, whether dysplastic or not, but can be induced by convulsants. The initiation of ictal discharges in the neocortex depends on both GABAergic signaling and increased extracellular potassium. In contrast, a spontaneous interictal-like activity is generated by tissues from patients with temporal lobe epilepsies associated with hippocampal sclerosis. This activity is initiated, not in the hippocampus but in the subiculum, an output region, which projects to the entorhinal cortex. Interictal events seem to be triggered by GABAergic cells, which paradoxically excite about 20% of subicular pyramidal cells while simultaneously inhibiting the majority. Interictal discharges thus depend on both GABAergic and glutamatergic signaling. The depolarizing effects of GABA depend on a pathological elevation in levels of chloride in some subicular cells, similar to those of developmentally immature cells. Such defect is caused by a perturbed expression of the cotransporters regulating intracellular chloride concentration, the importer NKCC1 and the extruder KCC2. Blockade of NKCC1 actions by the diuretic bumetanide restores intracellular chloride and thus hyperpolarizing GABAergic actions and consequently suppressing interictal

  3. Urinary Kallikrein Excretion in Essential and Mineralocorticoid Hypertension

    PubMed Central

    Holland, O. Bryan; Chud, James M.; Braunstein, Helen

    1980-01-01

    Urinary kallikrein excretion has been reported to be decreased in patients with essential hypertension and elevated in patients with primary aldosteronism as a reflection of mineralocorticoid activity. Low renin essential hypertension (LREH) has been postulated to result from excess production of an unknown mineralocorticoid(s). Urinary kallikrein excretion was compared in outpatients with essential hypertension, mineralocorticoid hypertension (primary aldosteronism and 17α-hydroxylase deficiency), and in normal subjects of the same race. No significant difference in urinary kallikrein excretion of patients with LREH vs. normal renin essential hypertension (NREH) was found for either black (4.1±0.4 vs. 4.8±0.5 esterase units (EU)/24 h, mean±SE, for 27 LREH and 38 NREH, respectively) or white patients (12.2±2.3 vs. 11.7±1.4 EU/24 h for 13 LREH and 25 NREH, respectively). Urinary kallikrein was decreased in black vs. white hypertensive patients and normal subjects. However, in patients with normal renal function (creatinine clearance ≥80 ml/min) urinary kallikrein was not significantly decreased in either black hypertensive vs. black normal subjects (4.3±0.3 vs. 5.4±0.6 EU/24 h) or in white hypertensive vs. white normal subjects (11.9±1.2 vs. 8.4±0.9 EU/24 h). In contrast, hypertensive patients with mild renal insufficiency (creatinine clearance of 41.8±78.5 ml/min) had reduced (P < 0.05) urinary kallikrein (3.3 EU/24 h with creatinine clearance of 63.6±2.0 for 24 black patients and 4.2±0.7 EU/24 h with creatinine clearance of 67.0±3.5 for 6 white patients). These results suggest that a reduction in urinary kallikrein excretion rate is an early accompaniment of hypertensive renal injury. Urinary kallikrein excretion in response to a 6-d 10-meq sodium diet and a 3-d Florinef (0.5 mg b.i.d.) administration was compared in hypertensive patients with normal renal function vs. race and age-matched normal subjects. Stimulation of urinary kallikrein

  4. Three-Dimensional Human Tissue Models of Wounded Skin

    PubMed Central

    Egles, Christophe; Garlick, Jonathan A.; Shamis, Yulia

    2010-01-01

    Human skin equivalents (HSEs) are in vitro tissues in which a fully differentiated, stratified squamous epithelium is grown at an air–liquid interface on a Type I collagen gel harboring human dermal fibroblasts. HSEs now provide experimental human tissue models to study factors that direct re-epithelialization and epithelial–mesenchymal cross-talk following wounding. This chapter describes the fabrication of HSEs from human keratinocytes and fibroblasts and how HSEs can be modified to characterize the response of the human epithelium during wound repair. The protocols outlined first describe techniques for the generation of human tissues that closely approximate the architectural features, differentiation, and growth of human skin. This will be followed by a description of a protocol that enables HSEs to be adapted to monitor their response following wounding. These engineered human tissues provide powerful tools to study biological process in tissues that mimic the healing of human skin and of the epithelial tissue. PMID:19908015

  5. The reconstruction and analysis of tissue specific human metabolic networks.

    PubMed

    Hao, Tong; Ma, Hong-Wu; Zhao, Xue-Ming; Goryanin, Igor

    2012-02-01

    Human tissues have distinct biological functions. Many proteins/enzymes are known to be expressed only in specific tissues and therefore the metabolic networks in various tissues are different. Though high quality global human metabolic networks and metabolic networks for certain tissues such as liver have already been studied, a systematic study of tissue specific metabolic networks for all main tissues is still missing. In this work, we reconstruct the tissue specific metabolic networks for 15 main tissues in human based on the previously reconstructed Edinburgh Human Metabolic Network (EHMN). The tissue information is firstly obtained for enzymes from Human Protein Reference Database (HPRD) and UniprotKB databases and transfers to reactions through the enzyme-reaction relationships in EHMN. As our knowledge of tissue distribution of proteins is still very limited, we replenish the tissue information of the metabolic network based on network connectivity analysis and thorough examination of the literature. Finally, about 80% of proteins and reactions in EHMN are determined to be in at least one of the 15 tissues. To validate the quality of the tissue specific network, the brain specific metabolic network is taken as an example for functional module analysis and the results reveal that the function of the brain metabolic network is closely related with its function as the centre of the human nervous system. The tissue specific human metabolic networks are available at .

  6. The Kallikrein Inhibitor from Bauhinia bauhinioides (BbKI) shows antithrombotic properties in venous and arterial thrombosis models.

    PubMed

    Brito, Marlon V; de Oliveira, Cleide; Salu, Bruno R; Andrade, Sonia A; Malloy, Paula M D; Sato, Ana C; Vicente, Cristina P; Sampaio, Misako U; Maffei, Francisco H A; Oliva, Maria Luiza V

    2014-05-01

    The Bauhinia bauhinioides Kallikrein Inhibitor (BbKI) is a Kunitz-type serine peptidase inhibitor of plant origin that has been shown to impair the viability of some tumor cells and to feature a potent inhibitory activity against human and rat plasma kallikrein (Kiapp 2.4 nmol/L and 5.2 nmol/L, respectively). This inhibitory activity is possibly responsible for an effect on hemostasis by prolonging activated partial thromboplastin time (aPTT). Because the association between cancer and thrombosis is well established, we evaluated the possible antithrombotic activity of this protein in venous and arterial thrombosis models. Vein thrombosis was studied in the vena cava ligature model in Wistar rats, and arterial thrombosis in the photochemical induced endothelium lesion model in the carotid artery of C57 black 6 mice. BbKI at a concentration of 2.0 mg/kg reduced the venous thrombus weight by 65% in treated rats in comparison to rats in the control group. The inhibitor prolonged the time for total artery occlusion in the carotid artery model mice indicating that this potent plasma kallikrein inhibitor prevented thrombosis.

  7. 21 CFR 1270.42 - Human tissue offered for import.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Human tissue offered for import. 1270.42 Section 1270.42 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) REGULATIONS UNDER CERTAIN OTHER ACTS ADMINISTERED BY THE FOOD AND DRUG ADMINISTRATION HUMAN TISSUE...

  8. 21 CFR 1270.42 - Human tissue offered for import.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Human tissue offered for import. 1270.42 Section 1270.42 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) REGULATIONS UNDER CERTAIN OTHER ACTS ADMINISTERED BY THE FOOD AND DRUG ADMINISTRATION HUMAN TISSUE...

  9. 21 CFR 1270.42 - Human tissue offered for import.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Human tissue offered for import. 1270.42 Section 1270.42 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) REGULATIONS UNDER CERTAIN OTHER ACTS ADMINISTERED BY THE FOOD AND DRUG ADMINISTRATION HUMAN TISSUE...

  10. 21 CFR 1270.42 - Human tissue offered for import.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Human tissue offered for import. 1270.42 Section 1270.42 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) REGULATIONS UNDER CERTAIN OTHER ACTS ADMINISTERED BY THE FOOD AND DRUG ADMINISTRATION HUMAN TISSUE...

  11. 21 CFR 1270.42 - Human tissue offered for import.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Human tissue offered for import. 1270.42 Section 1270.42 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) REGULATIONS UNDER CERTAIN OTHER ACTS ADMINISTERED BY THE FOOD AND DRUG ADMINISTRATION HUMAN TISSUE...

  12. Properties of kallikrein-containing granules isolated from the submaxillary gland of the guinea-pig.

    PubMed

    Bhoola, K D; Heap, P F

    1970-09-01

    1. Granular fractions of high purity consisting of subcellular kallikrein- and amylase-storing organelles have been isolated from homogenates of guinea-pig submaxillary gland.2. The isolated kallikrein- and amylase-containing granules closely resembled secretory granules observed in situ in serous acinar cells in intra-granular appearance, size and histochemical reaction.3. The subcellular, histochemical and ultrastructural studies indicate that the serine protease, kallikrein, is like amylase an exocrine enzyme with a functional role in saliva. PMID:5501268

  13. Imaging the Human Body: Micro- and Nanostructure of Human Tissues

    NASA Astrophysics Data System (ADS)

    Schulz, Georg; Deyhle, Hans; Müller, Bert

    Computed tomography based on X-rays is known to provide the best spatial resolution of all clinical three-dimensional imaging facilities and currently reaches a fraction of a millimeter. Better spatial and density resolution is obtained by means of micro computed tomography well established in the field of materials science. It is also very supportive imaging human tissues down to the level of individual cells (Lareida et al. J. Microsc. 234:95, 2009). The article demonstrates the power of micro computed tomography for imaging parts of the human body such as teeth, inner ear, cerebellum, tumors, and urethral tissue with conventional X-ray sources and synchrotron radiation facilities in absorption and phase contrast modes. The second part of the chapter relies on scanning X-ray scattering of tooth slices (Müller et al. Eur. J. Clin. Nanomed. 3:30, 2010) to uncover the presence of nanostructures including their anisotropy and orientation. This imaging technique gives unrivalled insights for medical experts, which will have a major influence on fields such as dental and incontinence treatments.

  14. Prostate kallikrein markers in diagnosis, risk stratification and prognosis

    PubMed Central

    Ulmert, David; O’Brien, M. Frank; Bjartell, Anders S.; Lilja, Hans

    2015-01-01

    Summary The kallikrein, prostate-specific antigen (PSA), is one of the world’s most frequently used disease biomarkers. After almost two decades of research and clinical experience, we are now starting to comprehend its diagnostic and monitoring limitations. Most physicians are aware of PSA’s low cancer-specificity among older men with benign prostatic conditions, but fewer are aware of recent data showing that a prior negative biopsy or a prior PSA below cut-off may have even stronger negative impact. Furthermore, a subtle PSA rise at early middle age strongly correlates to prostate cancer of unquestionable significance. We thoroughly review current and past reports on prostate kallikreins in relation to pathology and epidemiology. PMID:19578355

  15. Regulation of kallikrein-related peptidases in the skin - from physiology to diseases to therapeutic options.

    PubMed

    Fischer, J; Meyer-Hoffert, U

    2013-09-01

    Kallikrein-related peptidases (KLKs) constitute a family of 15 highly conserved serine proteases, which show a tissue-specific expression profile. This made them valuable tumour expression markers. It became evident that KLKs are involved in many physiological processes like semen liquefaction and skin desquamation. More recently, we have learnt that they are involved in many pathophysiological conditions and diseases making them promising target of therapeutic intervention. Therefore, regulation of KLKs raised the interest of numerous reports. Herein, we summarise the current knowledge on KLKs regulation with an emphasis on skin-relevant KLKs regulation processes. Regulation of KLKs takes place on the level of transcription, on protease activation and on protease inactivation. A variety of protease inhibitors has been described to interact with KLKs including the irreversible serine protease inhibitors (SERPINs) and the reversible serine protease inhibitors of Kazal-type (SPINKs). In an attempt to integrate current knowledge, we propose that KLK regulation has credentials as targets for therapeutic intervention.

  16. Studies on the Interaction between Collagen and a Plasma Kallikrein-Like Activity EVIDENCE FOR A SURFACE-ACTIVE ENZYME SYSTEM

    PubMed Central

    Harpel, Peter C.

    1972-01-01

    This study has demonstrated that collagen particles, after exposure to platelet-poor human plasma and subsequent washing, generate a kinin-like agent when incubated with prekinin substrate. The binding of kinin-generating activity to collagen in the plasma collagen incubation mixture occurs rapidly, whereas the loss of this activity in the incubation mixture occurs relatively slowly. The Hageman factor appeared to be necessary for the surface-bound kinin-generating activity, as this activity was absent in collagen exposed to Hageman factor-deficient plasma. These studies have partially characterized the plasma-derived enzymatic activity bound to collagen. Incubation of collagen with plasma caused a concentration-dependent reduction in the kinin-producing activity which was generated by the addition of ellagic acid, a known activator of plasma kallikrein. The kinin-inducing activity bound to collagen is inhibited by soybean trypsin inhibitor, Trasylol, serum C1̄ inactivator and the plasma α2-macroglobulin, but not by lima bean trypsin inhibitor. An eluate prepared from plasma-treated collagen, when compared with purified plasma kallikrein, shared a similar inhibitor profile. Selective chemical blockage of the free carboxyl groups on the collagen molecule, or heat denaturation, inactivated the ability of the collagen to generate kinin-like activity after incubation with plasma. Removal of the collagen telopeptides or blockage of the free amino groups failed to affect the collagen-plasma interaction. The binding of partially purified plasma kallikrein to collagen was found to have similar structural and chemical requirements. These data indicate that there is a structural and chemical specificity for the activation and binding of plasma kallikrein-like activity by collagen. These studies suggest that a plasma kallikrein may function as a surface-bound enzyme system. PMID:4338122

  17. Kinins produced from bovine colostrum by kallikrein and saliva

    PubMed Central

    Guth, Paul S.

    1959-01-01

    Substances capable of stimulating smooth muscle are produced on the incubation of bovine colostrum with urinary kallikrein or calf saliva. These substances, called urine- and saliva-colostrokinin, have been differentiated from kallidin, substance A and similar smooth muscle activating agents. Saliva-colostrokinin is likely to be formed in the suckling calf. Further, as colostrum became milk, the ability to form colostrokinin diminished. A function for saliva-colostrokinin in the newborn is suggested. PMID:13830444

  18. Identification of human connective tissue in transplant of human oral mucosa in nude mice.

    PubMed

    Holmstrup, P; Hansen, I L; Harder, F; Dabelsteen, E

    1984-01-01

    The present study describes a method for identification of connective tissue of human oral mucosal transplants in nude mice. The method was based on the development of a murine antiserum to human fibroblasts. After absorption with murine fibroblasts the antiserum in an immunofluorescence method appeared to react specifically with human connective tissue of frozen sections, whereas the antiserum did not react with murine connective tissue. The antiserum, applied to frozen sections of human oral mucosal transplants in nude mice, could distinguish between human and murine connective tissue in the sections. The ability to distinguish between the two types of tissue was utilized to elucidate a possible relation between epithelial morphology and underlying type of connective tissue. It was found that the formation of rete ridges of transplanted human oral epithelium was dependent on the presence of subepithelial human connective tissue. The method described may be useful for the recognition of human tissue in experimental studies of human transplants to other species.

  19. The use of animal tissues alongside human tissue: Cultural and ethical considerations.

    PubMed

    Kaw, Anu; Jones, D Gareth; Zhang, Ming

    2016-01-01

    Teaching and research facilities often use cadaveric material alongside animal tissues, although there appear to be differences in the way we handle, treat, and dispose of human cadaveric material compared to animal tissue. This study sought to analyze cultural and ethical considerations and provides policy recommendations on the use of animal tissues alongside human tissue. The status of human and animal remains and the respect because of human and animal tissues were compared and analyzed from ethical, legal, and cultural perspectives. The use of animal organs and tissues is carried out within the context of understanding human anatomy and function. Consequently, the interests of human donors are to be pre-eminent in any policies that are enunciated, so that if any donors find the presence of animal remains unacceptable, the latter should not be employed. The major differences appear to lie in differences in our perceptions of their respective intrinsic and instrumental values. Animals are considered to have lesser intrinsic value and greater instrumental value than humans. These differences stem from the role played by culture and ethical considerations, and are manifested in the resulting legal frameworks. In light of this discussion, six policy recommendations are proposed, encompassing the nature of consent, respect for animal tissues as well as human remains, and appropriate separation of both sets of tissues in preparation and display.

  20. Successful cryopreservation of human ovarian cortex tissues using supercooling.

    PubMed

    Moriguchi, Hisashi; Zhang, Yue; Mihara, Makoto; Sato, Chifumi

    2012-01-01

    The development of new method to cryopreserve human ovarian cortex tissues without damage is needed for the improvement of quality of life (QOL) of female cancer patients. Here we show novel cryopreservation method of human ovarian cortex tissues by using supercooling (S.C.) procedure. Our method will be helpful in order to preserve fertility of female cancer patients.

  1. Transplantation of human adipose tissue to nude mice.

    PubMed

    Bach-Mortensen, N; Romert, P; Ballegaard, S

    1976-08-01

    Human adipose tissue was transplanted to the mouse mutant nude (nu/nu). All the grafts were accepted and contained fat cells easily distinguishable from those of the mouse. No detectable relation between the histological pictures before and after grafting was found. In some transplants nerve tissue, and in others macrophages containing fat droplets, were found. The fat tissue graft might be useful for investigation of the influence of various hormones on human fat cells.

  2. Predicting DNA methylation level across human tissues.

    PubMed

    Ma, Baoshan; Wilker, Elissa H; Willis-Owen, Saffron A G; Byun, Hyang-Min; Wong, Kenny C C; Motta, Valeria; Baccarelli, Andrea A; Schwartz, Joel; Cookson, William O C M; Khabbaz, Kamal; Mittleman, Murray A; Moffatt, Miriam F; Liang, Liming

    2014-04-01

    Differences in methylation across tissues are critical to cell differentiation and are key to understanding the role of epigenetics in complex diseases. In this investigation, we found that locus-specific methylation differences between tissues are highly consistent across individuals. We developed a novel statistical model to predict locus-specific methylation in target tissue based on methylation in surrogate tissue. The method was evaluated in publicly available data and in two studies using the latest IlluminaBeadChips: a childhood asthma study with methylation measured in both peripheral blood leukocytes (PBL) and lymphoblastoid cell lines; and a study of postoperative atrial fibrillation with methylation in PBL, atrium and artery. We found that our method can greatly improve accuracy of cross-tissue prediction at CpG sites that are variable in the target tissue [R(2) increases from 0.38 (original R(2) between tissues) to 0.89 for PBL-to-artery prediction; from 0.39 to 0.95 for PBL-to-atrium; and from 0.81 to 0.98 for lymphoblastoid cell line-to-PBL based on cross-validation, and confirmed using cross-study prediction]. An extended model with multiple CpGs further improved performance. Our results suggest that large-scale epidemiology studies using easy-to-access surrogate tissues (e.g. blood) could be recalibrated to improve understanding of epigenetics in hard-to-access tissues (e.g. atrium) and might enable non-invasive disease screening using epigenetic profiles. PMID:24445802

  3. LRRC31 is induced by IL-13 and regulates kallikrein expression and barrier function in the esophageal epithelium

    PubMed Central

    D’Mello, RJ; Caldwell, JM; Azouz, NP; Wen, T; Sherrill, JD; Hogan, SP; Rothenberg, ME

    2015-01-01

    Eosinophilic esophagitis (EoE) is an allergic inflammatory disease of the esophagus featuring increased esophageal interleukin 13 (IL-13) levels and impaired barrier function. Herein, we investigated leucine-rich repeat–containing protein 31 (LRRC31) in human EoE esophageal tissue and IL-13–treated esophageal epithelial cells. LRRC31 had basal mRNA expression in colonic and airway mucosal epithelium. Esophageal LRRC31 mRNA and protein increased in active EoE and strongly correlated with esophageal eosinophilia and IL13 and CCL26 mRNA expression. IL-13 treatment increased LRRC31 mRNA and protein in air-liquid interface–differentiated esophageal epithelial cells (EPC2s). At baseline, differentiated LRRC31-overexpressing EPC2s had increased barrier function (1.9-fold increase in transepithelial electrical resistance [P < 0.05] and 2.8-fold decrease in paracellular flux [P < 0.05]). RNA sequencing analysis of differentiated LRRC31-overexpressing EPC2s identified 38 dysregulated genes (P < 0.05), including 5 kallikrein (KLK) serine proteases. Notably, differentiated LRRC31-overexpressing EPC2s had decreased KLK expression and activity, whereas IL-13–treated, differentiated LRRC31 gene-silenced EPC2s had increased KLK expression and suprabasal epithelial detachment. We identified similarly dysregulated KLK expression in the esophagus of patients with active EoE and in IL-13–treated esophageal epithelial cells. We propose that LRRC31 is induced by IL-13 and modulates epithelial barrier function, potentially through KLK regulation. PMID:26462420

  4. [Secondary use of human tissue: consent and better information required].

    PubMed

    Vermeulen, Eric; Geesink, Ingrid; Schmidt, Marjanka K; Steegers, Chantal; Verhue, Dieter; Brom, Frans W A; Aaronson, Neil K; van Leeuwen, Flora E

    2009-01-01

    Human tissue remaining after diagnostic procedures is important for use in scientific research. This 'secondary use' of tissue is regulated by the Dutch Medical Treatment Contracts Act and the Code of Conduct for Proper Secondary Use of Human Tissue of the Dutch Federation of Biomedical Scientific Societies. Patients have the right to opt-out of further use of their residual tissue, but the procedures for objection and the provision of information involved are not regulated by statute. Dutch patients have a positive attitude to further use of human tissue for other purposes. They prefer, however, a procedure in which they are informed verbally by their health professional about research with residual tissue. The information can be brief and is best provided early in the treatment. Administrative and technical modifications of the current registration systems are necessary to support the opting-out procedure in practice. By taking the preferences of patients into account, trust in medical practice can be maintained.

  5. Depth-resolved fluorescence of human ectocervical tissue

    NASA Astrophysics Data System (ADS)

    Wu, Yicong; Xi, Peng; Cheung, Tak-Hong; Yim, So Fan; Yu, Mei-Yung; Qu, Jianan Y.

    2005-04-01

    The depth-resolved autofluorescence of normal and dysplastic human ectocervical tissue within 120um depth were investigated utilizing a portable confocal fluorescence spectroscopy with the excitations at 355nm and 457nm. From the topmost keratinizing layer of all ectocervical tissue samples, strong keratin fluorescence with the spectral characteristics similar to collagen was observed, which created serious interference in seeking the correlation between tissue fluorescence and tissue pathology. While from the underlying non-keratinizing epithelial layer, the measured NADH fluorescence induced by 355nm excitation and FAD fluorescence induced by 457nm excitation were strongly correlated to the tissue pathology. The ratios between NADH over FAD fluorescence increased statistically in the CIN epithelial relative to the normal and HPV epithelia, which indicated increased metabolic activity in precancerous tissue. This study demonstrates that the depth-resolved fluorescence spectroscopy can reveal fine structural information on epithelial tissue and potentially provide more accurate diagnostic information for determining tissue pathology.

  6. Distribution of miRNA expression across human tissues.

    PubMed

    Ludwig, Nicole; Leidinger, Petra; Becker, Kurt; Backes, Christina; Fehlmann, Tobias; Pallasch, Christian; Rheinheimer, Steffi; Meder, Benjamin; Stähler, Cord; Meese, Eckart; Keller, Andreas

    2016-05-01

    We present a human miRNA tissue atlas by determining the abundance of 1997 miRNAs in 61 tissue biopsies of different organs from two individuals collected post-mortem. One thousand three hundred sixty-four miRNAs were discovered in at least one tissue, 143 were present in each tissue. To define the distribution of miRNAs, we utilized a tissue specificity index (TSI). The majority of miRNAs (82.9%) fell in a middle TSI range i.e. were neither specific for single tissues (TSI > 0.85) nor housekeeping miRNAs (TSI < 0.5). Nonetheless, we observed many different miRNAs and miRNA families that were predominantly expressed in certain tissues. Clustering of miRNA abundances revealed that tissues like several areas of the brain clustered together. Considering -3p and -5p mature forms we observed miR-150 with different tissue specificity. Analysis of additional lung and prostate biopsies indicated that inter-organism variability was significantly lower than inter-organ variability. Tissue-specific differences between the miRNA patterns appeared not to be significantly altered by storage as shown for heart and lung tissue. MiRNAs TSI values of human tissues were significantly (P = 10(-8)) correlated with those of rats; miRNAs that were highly abundant in certain human tissues were likewise abundant in according rat tissues. We implemented a web-based repository enabling scientists to access and browse the data (https://ccb-web.cs.uni-saarland.de/tissueatlas).

  7. Three-Dimensionally Engineered Normal Human Broncho-epithelial Tissue-Like Assemblies: Target Tissues for Human Respiratory Viral Infections

    NASA Technical Reports Server (NTRS)

    Goodwin, T. J.; McCarthy, M.; Lin, Y-H

    2006-01-01

    In vitro three-dimensional (3D) human broncho-epithelial (HBE) tissue-like assemblies (3D HBE TLAs) from this point forward referred to as TLAs were engineered in Rotating Wall Vessel (RWV) technology to mimic the characteristics of in vivo tissues thus providing a tool to study human respiratory viruses and host cell interactions. The TLAs were bioengineered onto collagen-coated cyclodextran microcarriers using primary human mesenchymal bronchial-tracheal cells (HBTC) as the foundation matrix and an adult human bronchial epithelial immortalized cell line (BEAS-2B) as the overlying component. The resulting TLAs share significant characteristics with in vivo human respiratory epithelium including polarization, tight junctions, desmosomes, and microvilli. The presence of tissue-like differentiation markers including villin, keratins, and specific lung epithelium markers, as well as the production of tissue mucin, further confirm these TLAs differentiated into tissues functionally similar to in vivo tissues. Increasing virus titers for human respiratory syncytial virus (wtRSVA2) and parainfluenza virus type 3 (wtPIV3 JS) and the detection of membrane bound glycoproteins over time confirm productive infections with both viruses. Therefore, TLAs mimic aspects of the human respiratory epithelium and provide a unique capability to study the interactions of respiratory viruses and their primary target tissue independent of the host's immune system.

  8. Three-Dimensionally Engineered Normal Human Lung Tissue-Like Assemblies: Target Tissues for Human Respiratory Viral Infections

    NASA Technical Reports Server (NTRS)

    Goodwin, Thomas J.; McCarthy, M.; Lin, Y-H.; Deatly, A. M.

    2008-01-01

    In vitro three-dimensional (3D) human lung epithelio-mesenchymal tissue-like assemblies (3D hLEM TLAs) from this point forward referred to as TLAs were engineered in Rotating Wall Vessel (RWV) technology to mimic the characteristics of in vivo tissues thus providing a tool to study human respiratory viruses and host cell interactions. The TLAs were bioengineered onto collagen-coated cyclodextran microcarriers using primary human mesenchymal bronchial-tracheal cells (HBTC) as the foundation matrix and an adult human bronchial epithelial immortalized cell line (BEAS-2B) as the overlying component. The resulting TLAs share significant characteristics with in vivo human respiratory epithelium including polarization, tight junctions, desmosomes, and microvilli. The presence of tissue-like differentiation markers including villin, keratins, and specific lung epithelium markers, as well as the production of tissue mucin, further confirm these TLAs differentiated into tissues functionally similar to in vivo tissues. Increasing virus titers for human respiratory syncytial virus (wtRSVA2) and the detection of membrane bound glycoproteins over time confirm productive infection with the virus. Therefore, we assert TLAs mimic aspects of the human respiratory epithelium and provide a unique capability to study the interactions of respiratory viruses and their primary target tissue independent of the host s immune system.

  9. Bovine Leukemia Virus DNA in Human Breast Tissue

    PubMed Central

    Shen, Hua Min; Jensen, Hanne M.; Choi, K. Yeon; Sun, Dejun; Nuovo, Gerard

    2014-01-01

    Bovine leukemia virus (BLV), a deltaretrovirus, causes B-cell leukemia/lymphoma in cattle and is prevalent in herds globally. A previous finding of antibodies against BLV in humans led us to examine the possibility of human infection with BLV. We focused on breast tissue because, in cattle, BLV DNA and protein have been found to be more abundant in mammary epithelium than in lymphocytes. In human breast tissue specimens, we identified BLV DNA by using nested liquid-phase PCR and DNA sequencing. Variations from the bovine reference sequence were infrequent and limited to base substitutions. In situ PCR and immunohistochemical testing localized BLV to the secretory epithelium of the breast. Our finding of BLV in human tissues indicates a risk for the acquisition and proliferation of this virus in humans. Further research is needed to determine whether BLV may play a direct role in human disease. PMID:24750974

  10. Altered autophagy in human adipose tissues in obesity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Context: Autophagy is a housekeeping mechanism, involved in metabolic regulation and stress response, shown recently to regulate lipid droplets biogenesis/breakdown and adipose tissue phenotype. Objective: We hypothesized that in human obesity autophagy may be altered in adipose tissue in a fat d...

  11. Microwave non-contact imaging of subcutaneous human body tissues.

    PubMed

    Kletsov, Andrey; Chernokalov, Alexander; Khripkov, Alexander; Cho, Jaegeol; Druchinin, Sergey

    2015-10-01

    A small-size microwave sensor is developed for non-contact imaging of a human body structure in 2D, enabling fitness and health monitoring using mobile devices. A method for human body tissue structure imaging is developed and experimentally validated. Subcutaneous fat tissue reconstruction depth of up to 70 mm and maximum fat thickness measurement error below 2 mm are demonstrated by measurements with a human body phantom and human subjects. Electrically small antennas are developed for integration of the microwave sensor into a mobile device. Usability of the developed microwave sensor for fitness applications, healthcare, and body weight management is demonstrated.

  12. Microwave non-contact imaging of subcutaneous human body tissues

    PubMed Central

    Chernokalov, Alexander; Khripkov, Alexander; Cho, Jaegeol; Druchinin, Sergey

    2015-01-01

    A small-size microwave sensor is developed for non-contact imaging of a human body structure in 2D, enabling fitness and health monitoring using mobile devices. A method for human body tissue structure imaging is developed and experimentally validated. Subcutaneous fat tissue reconstruction depth of up to 70 mm and maximum fat thickness measurement error below 2 mm are demonstrated by measurements with a human body phantom and human subjects. Electrically small antennas are developed for integration of the microwave sensor into a mobile device. Usability of the developed microwave sensor for fitness applications, healthcare, and body weight management is demonstrated. PMID:26609415

  13. Total DDT and dieldrin content of human adipose tissue

    SciTech Connect

    Ahmad, N.; Harsas, W.; Marolt, R.S.; Morton, M.; Pollack, J.K.

    1988-12-01

    As far as the authors could ascertain only 4 well-documented analytical studies have been carried out in Australia determining the total DDT and dieldrin content of human adipose tissue. The latest of these studies was published over 16 years ago. Therefore it is timely and important to re-examine the total DDT and dieldrin concentration within the adipose tissue of the Australian population. The present investigation has analyzed 290 samples of human adipose tissue obtained from Westmead Hospital situated in an outer suburb of Sydney, New South Wales for their content of total DDT and dieldrin.

  14. Cartilage tissue engineering identifies abnormal human induced pluripotent stem cells.

    PubMed

    Yamashita, Akihiro; Liu, Shiying; Woltjen, Knut; Thomas, Bradley; Meng, Guoliang; Hotta, Akitsu; Takahashi, Kazutoshi; Ellis, James; Yamanaka, Shinya; Rancourt, Derrick E

    2013-01-01

    Safety is the foremost issue in all human cell therapies, but human induced pluripotent stem cells (iPSCs) currently lack a useful safety indicator. Studies in chimeric mice have demonstrated that certain lines of iPSCs are tumorigenic; however a similar screen has not been developed for human iPSCs. Here, we show that in vitro cartilage tissue engineering is an excellent tool for screening human iPSC lines for tumorigenic potential. Although all human embryonic stem cells (ESCs) and most iPSC lines tested formed cartilage safely, certain human iPSCs displayed a pro-oncogenic state, as indicated by the presence of secretory tumors during cartilage differentiation in vitro. We observed five abnormal iPSC clones amoungst 21 lines derived from five different reprogramming methods using three cellular origins. We conclude that in vitro cartilage tissue engineering is a useful approach to identify abnormal human iPSC lines.

  15. Human tissue profiling with multidimensional protein identification technology.

    PubMed

    Cagney, Gerard; Park, Stephen; Chung, Clement; Tong, Bianca; O'Dushlaine, Colm; Shields, Denis C; Emili, Andrew

    2005-01-01

    Profiling of tissues and cell types through systematic characterization of expressed genes or proteins shows promise as a basic research tool, and has potential applications in disease diagnosis and classification. We used multidimensional protein identification protein identification technology (MudPIT) to analyze proteomes for enriched nuclear extracts of eight human tissues: brain, heart, liver, lung, muscle, pancreas, spleen, and testis. We show that the method is approximately 80% reproducible. We address issues of relative abundance, tissue-specificity, and selectivity, and the significance of proteins whose expression does not correlate with that of the corresponding mRNA. Surprisingly, most proteins are detected in a single tissue. These proteins tend to fulfill specialist (and potentially tissue-specific) functions compared to proteins expressed in two or more tissues.

  16. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture processing applications. (a) Identification. Tissue culture media for human ex vivo tissue and cell...

  17. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture processing applications. (a) Identification. Tissue culture media for human ex vivo tissue and cell...

  18. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture processing applications. (a) Identification. Tissue culture media for human ex vivo tissue and cell...

  19. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture processing applications. (a) Identification. Tissue culture media for human ex vivo tissue and cell...

  20. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture processing applications. (a) Identification. Tissue culture media for human ex vivo tissue and cell...

  1. A New Antigen Retrieval Technique for Human Brain Tissue

    PubMed Central

    Byne, William; Haroutunian, Vahram; García-Villanueva, Mercedes; Rábano, Alberto; García-Amado, María; Prensa, Lucía; Giménez-Amaya, José Manuel

    2008-01-01

    Immunohistochemical staining of tissues is a powerful tool used to delineate the presence or absence of an antigen. During the last 30 years, antigen visualization in human brain tissue has been significantly limited by the masking effect of fixatives. In the present study, we have used a new method for antigen retrieval in formalin-fixed human brain tissue and examined the effectiveness of this protocol to reveal masked antigens in tissues with both short and long formalin fixation times. This new method, which is based on the use of citraconic acid, has not been previously utilized in brain tissue although it has been employed in various other tissues such as tonsil, ovary, skin, lymph node, stomach, breast, colon, lung and thymus. Thus, we reported here a novel method to carry out immunohistochemical studies in free-floating human brain sections. Since fixation of brain tissue specimens in formaldehyde is a commonly method used in brain banks, this new antigen retrieval method could facilitate immunohistochemical studies of brains with prolonged formalin fixation times. PMID:18852880

  2. Predicting tissue-specific enhancers in the human genome

    PubMed Central

    Pennacchio, Len A.; Loots, Gabriela G.; Nobrega, Marcelo A.; Ovcharenko, Ivan

    2007-01-01

    Determining how transcriptional regulatory signals are encoded in vertebrate genomes is essential for understanding the origins of multicellular complexity; yet the genetic code of vertebrate gene regulation remains poorly understood. In an attempt to elucidate this code, we synergistically combined genome-wide gene-expression profiling, vertebrate genome comparisons, and transcription factor binding-site analysis to define sequence signatures characteristic of candidate tissue-specific enhancers in the human genome. We applied this strategy to microarray-based gene expression profiles from 79 human tissues and identified 7187 candidate enhancers that defined their flanking gene expression, the majority of which were located outside of known promoters. We cross-validated this method for its ability to de novo predict tissue-specific gene expression and confirmed its reliability in 57 of the 79 available human tissues, with an average precision in enhancer recognition ranging from 32% to 63% and a sensitivity of 47%. We used the sequence signatures identified by this approach to successfully assign tissue-specific predictions to ∼328,000 human–mouse conserved noncoding elements in the human genome. By overlapping these genome-wide predictions with a data set of enhancers validated in vivo, in transgenic mice, we were able to confirm our results with a 28% sensitivity and 50% precision. These results indicate the power of combining complementary genomic data sets as an initial computational foray into a global view of tissue-specific gene regulation in vertebrates. PMID:17210927

  3. Ultrastructural Changes in Human Trabecular Meshwork Tissue after Laser Trabeculoplasty

    PubMed Central

    SooHoo, Jeffrey R.; Seibold, Leonard K.; Ammar, David A.; Kahook, Malik Y.

    2015-01-01

    Purpose. To compare morphologic changes in human trabecular meshwork (TM) after selective laser trabeculoplasty (SLT) and argon laser trabeculoplasty (ALT). Design. Laboratory evaluation of ex vivo human eye TM after laser trabeculoplasty. Methods. Corneoscleral rims from human cadaver eyes were sectioned and treated with varying powers of either SLT or ALT. Specimens were examined using light microscopy, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Results. TEM of SLT at all powers resulted in disrupted TM cells with cracked and extracellular pigment granules. SEM of SLT samples treated at high power revealed tissue destruction with scrolling of trabecular beams. SEM of ALT-treated tissue showed increasing destruction with exposure to higher power. The presence or absence of “champagne” bubbles during SLT did not alter the histologic findings. Conclusions. SLT-treated human TM revealed disruption of TM cells with cracked, extracellular pigment granules, particularly at higher treatment powers. Tissue scrolling was noted at very high SLT energy levels. ALT-treated tissue showed significant damage to both the superficial and deeper TM tissues in a dose-dependent fashion. Further studies are needed to guide titration of treatment power to maximize the IOP-lowering effect while minimizing both energy delivered and damage to target tissues. PMID:26064672

  4. Human natural killer cell development in secondary lymphoid tissues.

    PubMed

    Freud, Aharon G; Yu, Jianhua; Caligiuri, Michael A

    2014-04-01

    For nearly a decade it has been appreciated that critical steps in human natural killer (NK) cell development likely occur outside of the bone marrow and potentially necessitate distinct microenvironments within extramedullary tissues. The latter include the liver and gravid uterus as well as secondary lymphoid tissues such as tonsils and lymph nodes. For as yet unknown reasons these tissues are naturally enriched with NK cell developmental intermediates (NKDI) that span a maturation continuum starting from an oligopotent CD34(+)CD45RA(+) hematopoietic precursor cell to a cytolytic mature NK cell. Indeed despite the detection of NKDI within the aforementioned tissues, relatively little is known about how, why, and when these tissues may be most suited to support NK cell maturation and how this process fits in with other components of the human immune system. With the discovery of other innate lymphoid subsets whose immunophenotypes overlap with those of NKDI, there is also need to revisit and potentially re-characterize the basic immunophenotypes of the stages of the human NK cell developmental pathway in vivo. In this review, we provide an overview of human NK cell development in secondary lymphoid tissues and discuss the many questions that remain to be answered in this exciting field.

  5. 78 FR 44134 - Submission for OMB Review; 30-day Comment Request: Financial Sustainability of Human Tissue...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-23

    ...: Financial Sustainability of Human Tissue Biobanking (NCI) SUMMARY: Under the provisions of Section 3507(a)(1... Collection: Financial Sustainability of Human Tissue Biobanking, 0925-NEW, National Cancer Institute...

  6. Predicting Tissue-Specific Enhancers in the Human Genome

    SciTech Connect

    Pennacchio, Len A.; Loots, Gabriela G.; Nobrega, Marcelo A.; Ovcharenko, Ivan

    2006-07-01

    Determining how transcriptional regulatory signals areencoded in vertebrate genomes is essential for understanding the originsof multi-cellular complexity; yet the genetic code of vertebrate generegulation remains poorly understood. In an attempt to elucidate thiscode, we synergistically combined genome-wide gene expression profiling,vertebrate genome comparisons, and transcription factor binding siteanalysis to define sequence signatures characteristic of candidatetissue-specific enhancers in the human genome. We applied this strategyto microarray-based gene expression profiles from 79 human tissues andidentified 7,187 candidate enhancers that defined their flanking geneexpression, the majority of which were located outside of knownpromoters. We cross-validated this method for its ability to de novopredict tissue-specific gene expression and confirmed its reliability in57 of the 79 available human tissues, with an average precision inenhancer recognition ranging from 32 percent to 63 percent, and asensitivity of 47 percent. We used the sequence signatures identified bythis approach to assign tissue-specific predictions to ~;328,000human-mouse conserved noncoding elements in the human genome. Byoverlapping these genome-wide predictions with a large in vivo dataset ofenhancers validated in transgenic mice, we confirmed our results with a28 percent sensitivity and 50 percent precision. These results indicatethe power of combining complementary genomic datasets as an initialcomputational foray into the global view of tissue-specific generegulation in vertebrates.

  7. Human Neural Tissue Construct Fabrication Based on Scaffold-Free Tissue Engineering.

    PubMed

    Takahashi, Hironobu; Itoga, Kazuyoshi; Shimizu, Tatsuya; Yamato, Masayuki; Okano, Teruo

    2016-08-01

    Current neural tissue engineering strategies involve the development and application of neural tissue constructs produced by using an anisotropic polymeric scaffold. This study reports a scaffold-free method of tissue engineering to create a tubular neural tissue construct containing unidirectional neuron bundles. The surface patterning of a thermoresponsive culture substrate and a coculture system of neurons with patterned astrocytes can provide an anisotropic structure and easy handling of the neural tissue construct without the use of a scaffold. Furthermore, using a gelatin gel-coated plunger, the neuron bundles can be laid out in the same direction at regulated intervals within multilayered astrocyte sheets. Since the 3D tissue construct is composed only by neurons and astrocytes, they can communicate physiologically without obstruction of a scaffold. The medical benefits of scaffold-free tissue generation provide new opportunities for the development of human cell-based tissue models required to better understand the mechanisms of neurodegenerative diseases. Therefore, this new tissue engineering approach may be useful to establish a technology for regenerative medicine and drug discovery using the patient's own neurons. PMID:27331769

  8. Characterization of muscarinic receptor subtypes in human tissues

    SciTech Connect

    Giraldo, E.; Martos, F.; Gomez, A.; Garcia, A.; Vigano, M.A.; Ladinsky, H.; Sanchez de La Cuesta, F.

    1988-01-01

    The affinities of selective, pirenzepine and AF-DX 116, and classical, N-methylscopolamine and atropine, muscarinic cholinergic receptor antagonists were investigated in displacement binding experiments with (/sup 3/H)Pirenzepine and (/sup 3/H)N-methylscopolamine in membranes from human autoptic tissues (forebrain, cerebellum, atria, ventricle and submaxillary salivary glands). Affinity estimates of N-methylscopolamine and atropine indicated a non-selective profile. Pirenzepine showed differentiation between the M/sub 1/ neuronal receptor of the forebrain and the receptors in other tissues while AF-DX 116 clearly discriminated between muscarinic receptors of heart and glands. The results in human tissues confirm the previously described selectivity profiles of pirenzepine and AF-DX 116 in rat tissues. These findings thus reveal the presence also in man of three distinct muscarinic receptor subtypes: the neuronal M/sub 1/, the cardiac M/sub 2/ and the glandular M/sub 3/.

  9. Engineered human broncho-epithelial tissue-like assemblies

    NASA Technical Reports Server (NTRS)

    Goodwin, Thomas J. (Inventor)

    2012-01-01

    Three-dimensional human broncho-epithelial tissue-like assemblies (TLAs) are produced in a rotating wall vessel (RWV) with microcarriers by coculturing mesenchymal bronchial-tracheal cells (BTC) and bronchial epithelium cells (BEC). These TLAs display structural characteristics and express markers of in vivo respiratory epithelia. TLAs are useful for screening compounds active in lung tissues such as antiviral compounds, cystic fibrosis treatments, allergens, and cytotoxic compounds.

  10. Infrared absorption spectra of human malignant tumor tissues

    NASA Astrophysics Data System (ADS)

    Skornyakov, I. V.; Tolstorozhev, G. B.; Butra, V. A.

    2008-05-01

    We used infrared spectroscopy methods to study the molecular structure of tissues from human organs removed during surgery. The IR spectra of the surgical material from breast, thyroid, and lung are compared with data from histological examination. We show that in malignant neoplasms, a change occurs in the hydrogen bonds of protein macromolecules found in the tissue of the studied organs. We identify the spectral signs of malignant pathology.

  11. Laser ablation of human atherosclerotic plaque without adjacent tissue injury

    NASA Technical Reports Server (NTRS)

    Grundfest, W. S.; Litvack, F.; Forrester, J. S.; Goldenberg, T.; Swan, H. J. C.

    1985-01-01

    Seventy samples of human cadaver atherosclerotic aorta were irradiated in vitro using a 308 nm xenon chloride excimer laser. Energy per pulse, pulse duration and frequency were varied. For comparison, 60 segments were also irradiated with an argon ion and an Nd:YAG laser operated in the continuous mode. Tissue was fixed in formalin, sectioned and examined microscopically. The Nd:YAG and argon ion-irradiated tissue exhibited a central crater with irregular edges and concentric zones of thermal and blast injury. In contrast, the excimer laser-irradiated tissue had narrow deep incisions with minimal or no thermal injury. These preliminary experiments indicate that the excimer laser vaporizes tissue in a manner different from that of the continuous wave Nd:YAG or argon ion laser. The sharp incision margins and minimal damage to adjacent normal tissue suggest that the excimer laser is more desirable for general surgical and intravascular uses than are the conventionally used medical lasers.

  12. Solubility of Freon 22 in human blood and lung tissue.

    PubMed

    Varene, N; Choukroun, M L; Marthan, R; Varene, P

    1989-05-01

    The solubility of Freon 22 in human blood and lung tissue was determined using the chromatographic method of Wagner et al. (J. Appl. Physiol. 36: 600-605, 1974). In normal human blood, the mean Bunsen coefficient of solubility (alpha B) was 0.804 cm3 STPD.cm-3.ATA-1 at 37 degrees C. It increased with hematocrit (Hct) according to the equation alpha B = 0.274 Hct + 0.691. Tissue homogenates were prepared from macroscopically normal lung pieces obtained at thoracotomy from eight patients undergoing resection for lung carcinoma. The Bunsen solubility coefficients were 0.537 +/- 0.068 and 0.635 +/- 0.091 in washed and unwashed lung, respectively. These values can be used in the determination of both cardiac output and pulmonary tissue volume in humans by use of the rebreathing technique.

  13. Solubility of Freon 22 in human blood and lung tissue

    SciTech Connect

    Varene, N.; Choukroun, M.L.; Marthan, R.; Varene, P.

    1989-05-01

    The solubility of Freon 22 in human blood and lung tissue was determined using the chromatographic method of Wagner et al. In normal human blood, the mean Bunsen coefficient of solubility (alpha B) was 0.804 cm3 STPD.cm-3.ATA-1 at 37 degrees C. It increased with hematocrit (Hct) according to the equation alpha B = 0.274 Hct + 0.691. Tissue homogenates were prepared from macroscopically normal lung pieces obtained at thoracotomy from eight patients undergoing resection for lung carcinoma. The Bunsen solubility coefficients were 0.537 +/- 0.068 and 0.635 +/- 0.091 in washed and unwashed lung, respectively. These values can be used in the determination of both cardiac output and pulmonary tissue volume in humans by use of the rebreathing technique.

  14. The Role of Plasma Kallikrein-Kinin Pathway in the Development of Diabetic Retinopathy: Pathophysiology and Therapeutic Approaches.

    PubMed

    Abdulaal, Marwan; Haddad, Nour Maya N; Sun, Jennifer K; Silva, Paolo S

    2016-01-01

    Diabetic retinal disease is characterized by a series of retinal microvascular changes and increases in retinal vascular permeability that lead to development of diabetic retinopathy (DR) and diabetic macular edema (DME), respectively. Current treatment strategies for DR and DME are mostly limited to vascular endothelial growth factor (VEGF) inhibitors and laser photocoagulation. These treatment modalities are not universally effective in all patients, and potential side effects persist in a significant portion of patients. The plasma kallikrein-kinin system (KKS) is one of the pathways that has been identified in the vitreous in proliferative DR and DME. Preclinical studies have shown that the activation of intraocular KKS induces retinal vascular permeability, vasodilation, and retinal thickening. Proteomic analysis from vitreous of eyes with DME has shown that KKS and VEGF pathways are potentially independent biologic pathways. Furthermore, proteins associated with DME in the vitreous were significantly more correlated with the KKS pathway compared to VEGF pathway. Preclinical experiments on diabetic animals showed that inhibition of KKS components was found to be an effective approach to decrease retinal vascular permeability. An initial phase I human trial of a novel plasma kallikrein inhibitor for the treatment of DME is currently ongoing to test the safety of this approach and serves as an initial step in the translation of basic science discovery into an innovative clinical intervention. PMID:26959125

  15. Glomus tissue in the vicinity of the human carotid sinus.

    PubMed Central

    Garfia, A

    1980-01-01

    Three of 60 cadavers have shown, in the adventitia or in the adipose tissue from the human carotid sinus region, small islands of tissue richly and typically vascularized and with nerve endings contacting cells like the tissue of the principal carotid body. In two of the cases such 'miniglomera' were single but in the third there were several all on the same side. A modified en bloc silver nitrate reduction stain was used to demonstrate the microvascular arrangements and the nerve endings by light microscopy of serial tangential sections of the carotid bifurcation. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 PMID:7364653

  16. Engineering human cells and tissues through pluripotent stem cells.

    PubMed

    Jones, Jeffrey R; Zhang, Su-Chun

    2016-08-01

    The utility of human pluripotent stem cells (hPSCs) depends on their ability to produce functional cells and tissues of the body. Two strategies have been developed: directed differentiation of enriched populations of cells that match a regional and functional profile and spontaneous generation of three-dimensional organoids that resemble tissues in the body. Genomic editing of hPSCs and their differentiated cells broadens the use of the hPSC paradigm in studying human cellular function and disease as well as developing therapeutics.

  17. Discordance of DNA methylation variance between two accessible human tissues.

    PubMed

    Jiang, Ruiwei; Jones, Meaghan J; Chen, Edith; Neumann, Sarah M; Fraser, Hunter B; Miller, Gregory E; Kobor, Michael S

    2015-01-01

    Population epigenetic studies have been seeking to identify differences in DNA methylation between specific exposures, demographic factors, or diseases in accessible tissues, but relatively little is known about how inter-individual variability differs between these tissues. This study presents an analysis of DNA methylation differences between matched peripheral blood mononuclear cells (PMBCs) and buccal epithelial cells (BECs), the two most accessible tissues for population studies, in 998 promoter-located CpG sites. Specifically we compared probe-wise DNA methylation variance, and how this variance related to demographic factors across the two tissues. PBMCs had overall higher DNA methylation than BECs, and the two tissues tended to differ most at genomic regions of low CpG density. Furthermore, although both tissues showed appreciable probe-wise variability, the specific regions and magnitude of variability differed strongly between tissues. Lastly, through exploratory association analysis, we found indication of differential association of BEC and PBMC with demographic variables. The work presented here offers insight into variability of DNA methylation between individuals and across tissues and helps guide decisions on the suitability of buccal epithelial or peripheral mononuclear cells for the biological questions explored by epigenetic studies in human populations.

  18. Cortisol in human tissues at different stages of life.

    PubMed

    Costa, A; Benedetto, C; Fabris, C; Giraudi, G F; Testori, O; Bertino, E; Marozio, L; Varvello, G; Arisio, R; Ariano, M; Emanuel, A

    1996-01-01

    Aim of the work was to measure the cortisol level in human tissues at different stages of life, by means of radioimmunoassay and by chromatography. Viable samples of 13 different tissues were obtained during surgical intervention from 30 to 70 years old patients of either sex. Mean tissue cortisol concentration was 78 +/- 35 ng/g, ranging from 20 +/- 10 ng/g in the thyroid to 124 +/- 76 ng/g in the kidney. Similar values were measured in the corresponding tissues from not decayed corpses, so that paired values could be mediated. However the pancreas, and corrupted autopsy tissues, gave nil or exceedingly high cortisol concentration values; in some cases, opposite extreme values were measured in different organs of the same body. Cortisol concentration was also measured in 11 sound different tissues of spontaneously aborted or stillbirth fetuses, between 16 and 36 weeks of gestation. Mean value was 63 +/- 27 ng/g, ranging from 30 +/- 25 ng/g in the liver to 104 +/- 52 ng/g in the lungs. Also in fetuses nil or exceedingly high cortisol values occurred in altered tissues. One hundred and fourteen samples of limbs and carcasses of 7 to 12 gestational weeks embryos, obtained from voluntary abortions, were also examined: 20% gave nil result, in the remaining mean cortisol concentration was 32 ng/g. In 33 samples of embryos' mixed viscera, RIA and chromatography gave unreliable exceedingly high values. The nil and the exceedingly high values measured in the altered autoptic tissue specimens were inconsistent with the cortisol blood level measured in the patients, as were those measured in embryonic tissues with the acknowledged blood and adrenals cortisol levels at that stage of life. Thus cortisol may be measured by RIA and by chromatography in sound tissues, while the values obtained in the pancreas, in corrupted tissues, and in embryonal viscera do not represent the hormonal milieu, but are likely artifacts due to impeachment of the diagnostic system. PMID:8884541

  19. Formation of tissue factor activity following incubation of recombinant human tissue factor apoprotein with plasma lipoproteins

    SciTech Connect

    Sakai, T.; Kisiel, W. )

    1990-11-01

    Incubation of recombinant human tissue factor apoprotein (Apo-TF) with human plasma decreased the recalcified clotting time of this plasma in a time-and dose-dependent manner suggesting relipidation of the Apo-TF by plasma lipoproteins. Incubation of Apo-TF with purified preparations of human very low density, low density and high density lipoproteins resulted in tissue factor activity in a clotting assay. The order of effectiveness was VLDL greater than LDL much greater than HDL. Tissue factor activity generated by incubation of a fixed amount of Apo-TF with plasma lipoproteins was lipoprotein concentration-dependent and saturable. The association of Apo-TF with lipoprotein particles was supported by gel filtration studies in which {sup 125}I-Apo-TF coeluted with the plasma lipoprotein in the void volume of a Superose 6 column in the presence and absence of calcium ions. In addition, void-volume Apo-TF-lipoprotein fractions exhibited tissue factor activity. These results suggest that the factor VIII-bypassing activity of bovine Apo-TF observed in a canine hemophilic model may be due, in part, to its association with plasma lipoproteins and expression of functional tissue factor activity.

  20. Human monocytes can produce tissue-type plasminogen activator

    PubMed Central

    1989-01-01

    Evidence has previously been presented that monocytes and macrophages produce urokinase-type plasminogen activator. We have shown for the first time that human monocytes, when stimulated appropriately in vitro, can produce tissue type-plasminogen activator (t-PA) of 70 kD. Detection of t-PA mRNA was consistent with the biochemical and immunological characterization of t-PA produced by human monocytes. PMID:2494295

  1. Catalogue of human tissue optical properties at terahertz frequencies.

    PubMed

    Fitzgerald, A J; Berry, E; Zinov'ev, N N; Homer-Vanniasinkam, S; Miles, R E; Chamberlain, J M; Smith, M A

    2003-06-01

    Recently published studies suggest thatterahertz pulsed imaging will have applications inmedicine and biology, but there iscurrently very little information about the opticalproperties of human tissue at terahertzfrequencies. Such information would be useful forpredicting the feasibility of proposedapplications, optimising acquisition protocols,providing information about variability ofhealthy tissue and supplying data for studies of theinteraction mechanisms. Research ethicscommittee approval was obtained, andmeasurements made from samples of freshlyexcised human tissue, using a broadbandterahertz pulsed imaging system comprisingfrequencies approximately 0.5 to 2.5 THz.Refractive index and linear absorptioncoefficient were found. Reproducibility wasdetermined using blood from one volunteer,which was drawn and measured on consecutivedays. Skin, adipose tissue, striatedmuscle, vein and nerve were measured (to date, from oneindividual). Water had a higher refractiveindex (2.04 ± 0.07) than any tissue.The linear absorption coefficient was higher formuscle than adipose tissue, as expectedfrom the higher hydration of muscle. As these samples camefrom a single subject, there is currentlyinsufficient statistical power to draw firmconclusions, but results suggest that in vivo clinical imaging will be feasible in certainapplications. PMID:23345827

  2. Translational neuropharmacology: the use of human isolated gastrointestinal tissues.

    PubMed

    Sanger, G J; Broad, J; Kung, V; Knowles, C H

    2013-01-01

    Translational sciences increasingly emphasize the measurement of functions in native human tissues. However, such studies must confront variations in patient age, gender, genetic background and disease. Here, these are discussed with reference to neuromuscular and neurosecretory functions of the human gastrointestinal (GI) tract. Tissues are obtained after informed consent, in collaboration with surgeons (surgical techniques help minimize variables) and pathologists. Given the difficulties of directly recording from human myenteric neurones (embedded between muscle layers), enteric motor nerve functions are studied by measuring muscle contractions/relaxations evoked by electrical stimulation of intrinsic nerves; responses are regionally dependent, often involving cholinergic and nitrergic phenotypes. Enteric sensory functions can be studied by evoking the peristaltic reflex, involving enteric sensory and motor nerves, but this has rarely been achieved. As submucosal neurones are more accessible (after removing the mucosa), direct neuronal recordings are possible. Neurosecretory functions are studied by measuring changes in short-circuit current across the mucosa. For all experiments, basic questions must be addressed. Because tissues are from patients, what are the controls and the influence of disease? How long does it take before function fully recovers? What is the impact of age- and gender-related differences? What is the optimal sample size? Addressing these and other questions minimizes variability and raises the scientific credibility of human tissue research. Such studies also reduce animal use. Further, the many differences between animal and human GI functions also means that human tissue research must question the ethical validity of using strains of animals with unproved translational significance. PMID:22946540

  3. Collagen in Human Tissues: Structure, Function, and Biomedical Implications from a Tissue Engineering Perspective

    NASA Astrophysics Data System (ADS)

    Balasubramanian, Preethi; Prabhakaran, Molamma P.; Sireesha, Merum; Ramakrishna, Seeram

    The extracellular matrix is a complex biological structure encoded with various proteins, among which the collagen family is the most significant and abundant of all, contributing 30-35% of the whole-body protein. "Collagen" is a generic term for proteins that forms a triple-helical structure with three polypeptide chains, and around 29 types of collagen have been identified up to now. Although most of the members of the collagen family form such supramolecular structures, extensive diversity exists between each type of collagen. The diversity is not only based on the molecular assembly and supramolecular structures of collagen types but is also observed within its tissue distribution, function, and pathology. Collagens possess complex hierarchical structures and are present in various forms such as collagen fibrils (1.5-3.5 nm wide), collagen fibers (50-70 nm wide), and collagen bundles (150-250 nm wide), with distinct properties characteristic of each tissue providing elasticity to skin, softness of the cartilage, stiffness of the bone and tendon, transparency of the cornea, opaqueness of the sclera, etc. There exists an exclusive relation between the structural features of collagen in human tissues (such as the collagen composition, collagen fibril length and diameter, collagen distribution, and collagen fiber orientation) and its tissue-specific mechanical properties. In bone, a transverse collagen fiber orientation prevails in regions of higher compressive stress whereas longitudinally oriented collagen fibers correlate to higher tensile stress. The immense versatility of collagen compels a thorough understanding of the collagen types and this review discusses the major types of collagen found in different human tissues, highlighting their tissue-specific uniqueness based on their structure and mechanical function. The changes in collagen during a specific tissue damage or injury are discussed further, focusing on the many tissue engineering applications for

  4. Tissue distribution of human acetylcholinesterase and butyrylcholinesterase messenger RNA

    SciTech Connect

    Jbilo, O.; Barteles, C.F.; Chatonnet, A.; Toutant, J.P.; Lockridge, O.

    1994-12-31

    Tissue distribution of human acetyicholinesterase and butyryicholinesterase messenger RNA. 1 Cholinesterase inhibitors occur naturally in the calabar bean (eserine), green potatoes (solanine), insect-resistant crab apples, the coca plant (cocaine) and snake venom (fasciculin). There are also synthetic cholinesterase inhibitors, for example man-made insecticides. These inhibitors inactivate acetyicholinesterase and butyrylcholinesterase as well as other targets. From a study of the tissue distribution of acetylcholinesterase and butyrylcholinesterase mRNA by Northern blot analysis, we have found the highest levels of butyrylcholinesterase mRNA in the liver and lungs, tissues known as the principal detoxication sites of the human body. These results indicate that butyrylcholinesterase may be a first line of defense against poisons that are eaten or inhaled.

  5. Arrhenius parameters for primary thermal injury in human tonsillar tissue

    NASA Astrophysics Data System (ADS)

    McMillan, Kathleen; Radabaugh, Rebecca; Coad, James E.

    2011-03-01

    Clinical implementation of a thermal therapy requires the ability to predict tissue injury following exposures to specific thermal histories. As part of an effort to develop a nonexcisional alternative to tonsillectomy, the degree of primary hyperthermic tissue injury in human tonsil was characterized. Fifteen fresh pediatric hypertrophic tonsillectomy specimens were sectioned and treated in a NIST-calibrated saline bath at temperatures of 40 to 70°C with hold times of one to seven minutes. The treated tissues were subsequently nitroblue tetrazolium (NBT) stained to assess for thermal respiratory enzyme inactivation as a marker of cellular injury/death. The NBT stains were quantitatively image analyzed and used to calculate Arrhenius parameters for primary thermal injury in human tonsils.

  6. Plant-Derived Human Collagen Scaffolds for Skin Tissue Engineering

    PubMed Central

    Willard, James J.; Drexler, Jason W.; Das, Amitava; Roy, Sashwati; Shilo, Shani; Shoseyov, Oded

    2013-01-01

    Tissue engineering scaffolds are commonly formed using proteins extracted from animal tissues, such as bovine hide. Risks associated with the use of these materials include hypersensitivity and pathogenic contamination. Human-derived proteins lower the risk of hypersensitivity, but possess the risk of disease transmission. Methods engineering recombinant human proteins using plant material provide an alternate source of these materials without the risk of disease transmission or concerns regarding variability. To investigate the utility of plant-derived human collagen (PDHC) in the development of engineered skin (ES), PDHC and bovine hide collagen were formed into tissue engineering scaffolds using electrospinning or freeze-drying. Both raw materials were easily formed into two common scaffold types, electrospun nonwoven scaffolds and lyophilized sponges, with similar architectures. The processing time, however, was significantly lower with PDHC. PDHC scaffolds supported primary human cell attachment and proliferation at an equivalent or higher level than the bovine material. Interleukin-1 beta production was significantly lower when activated THP-1 macrophages where exposed to PDHC electrospun scaffolds compared to bovine collagen. Both materials promoted proper maturation and differentiation of ES. These data suggest that PDHC may provide a novel source of raw material for tissue engineering with low risk of allergic response or disease transmission. PMID:23298216

  7. Optical spectroscopy for quantitative sensing in human pancreatic tissues

    NASA Astrophysics Data System (ADS)

    Wilson, Robert H.; Chandra, Malavika; Lloyd, William; Chen, Leng-Chun; Scheiman, James; Simeone, Diane; McKenna, Barbara; Mycek, Mary-Ann

    2011-07-01

    Pancreatic adenocarcinoma has a five-year survival rate of only 6%, largely because current diagnostic methods cannot reliably detect the disease in its early stages. Reflectance and fluorescence spectroscopies have the potential to provide quantitative, minimally-invasive means of distinguishing pancreatic adenocarcinoma from normal pancreatic tissue and chronic pancreatitis. The first collection of wavelength-resolved reflectance and fluorescence spectra and time-resolved fluorescence decay curves from human pancreatic tissues was acquired with clinically-compatible instrumentation. Mathematical models of reflectance and fluorescence extracted parameters related to tissue morphology and biochemistry that were statistically significant for distinguishing between pancreatic tissue types. These results suggest that optical spectroscopy has the potential to detect pancreatic disease in a clinical setting.

  8. Studies on kallikrein in the duct systems of the salivary glands of the cat

    PubMed Central

    Shnitka, †T. K.; Maranda, B.; Rodrigues, J. A. A.; Schachter, M.; Weinberg, J.

    1978-01-01

    By correlating immunofluorescence light microscopy with electron microscope studies and with kallikrein concentrations under various conditions, we have made the following observations and conclusions about kallikrein in the submandibular and other salivary glands. 1. In the submandibular gland, specific immunofluorescence to kallikrein was observed in the luminal region of the striated ducts particularly, but also in the outer epithelial cells of the stratified epithelial collecting ducts. Sympathetic nerve stimulation resulted in a reduction in intensity of specific fluorescence and in its increased localization towards the lumen. The nearly complete elimination of kallikrein from the gland by duct obstruction for four days resulted in complete disappearance of specific fluorescence in the gland. Prolonged parasympathetic nerve stimulation at frequencies which did not reduce the kallikrein concentration of the gland failed to alter the specific immunofluorescence despite copious secretion of saliva. Our results failed to reveal evidence of secretion of kallikrein either into or towards the interstitium of the gland. The luminal layer of stratified epithelial cells in the collecting ducts contained small secretory granules closely resembling those in the striated ducts. Our results are not conclusive, but do suggest that kallikrein is located in these granules whence it is secreted into the lumen of the duct. 2. The parotid gland was found to contain much lower concentrations of kallikrein than the submandibular gland. This finding was associated with the presence of far fewer striated ducts in the parotid gland. Otherwise, specific fluorescence and the response to sympathetic nerve stimulation was like that of the submandibular gland. Small secretory granules in the striated and collecting ducts resembled those of the submandibular gland. 3. The sublingual gland, like the parotid, had a low concentration of kallikrein and very few striated ducts. These ducts were

  9. Assessment of permeation of lipoproteins in human carotid tissue

    NASA Astrophysics Data System (ADS)

    Ghosn, Mohamad G.; Syed, Saba H.; Leba, Michael; Morrisett, Joel D.; Tuchin, Valery V.; Larin, Kirill V.

    2010-02-01

    Cardiovascular disease is among the leading causes of death in the United States. Specifically, atherosclerosis is an increasingly devastating contributor to the tally and has been found to be a byproduct of arterial permeability irregularities in regards to lipoprotein penetration. To further explore arterial physiology and molecular transport, the imaging technique of Optical Coherence Tomography (OCT) was employed. With OCT, the permeation of glucose (MW = 180 Da), low density lipoprotein (LDL; MW = 2.1 × 106 Da), and high density lipoprotein (HDL; MW = 2.5 × 105 Da) in human carotid tissue was studied to determine the effect of different molecular characteristics on permeation in atherosclerotic tissues. The permeability rates calculated from the diffusion of the molecular agents into the abnormal carotid tissue samples is compared to those of normal, healthy tissue. The results show that in the abnormal tissue, the permeation of agents correlate to the size constraints. The larger molecules of LDL diffuse the slowest, while the smallest molecules of glucose diffuse the fastest. However, in normal tissue, LDL permeates at a faster rate than the other two agents, implying the existence of a transport mechanism that facilitates the passage of LDL molecules. These results highlight the capability of OCT as a sensitive and specific imaging technique as well as provide significant information to the understanding of atherosclerosis and its effect on tissue properties.

  10. Human cancers overexpress genes that are specific to a variety of normal human tissues

    PubMed Central

    Lotem, Joseph; Netanely, Dvir; Domany, Eytan; Sachs, Leo

    2005-01-01

    We have analyzed gene expression data from three different kinds of samples: normal human tissues, human cancer cell lines, and leukemic cells from lymphoid and myeloid leukemia pediatric patients. We have searched for genes that are overexpressed in human cancer and also show specific patterns of tissue-dependent expression in normal tissues. Using the expression data of the normal tissues, we identified 4,346 genes with a high variability of expression and clustered these genes according to their relative expression level. Of 91 stable clusters obtained, 24 clusters included genes preferentially expressed either only in hematopoietic tissues or in hematopoietic and one to two other tissues; 28 clusters included genes preferentially expressed in various nonhematopoietic tissues such as neuronal, testis, liver, kidney, muscle, lung, pancreas, and placenta. Analysis of the expression levels of these two groups of genes in the human cancer cell lines and leukemias identified genes that were highly expressed in cancer cells but not in their normal counterparts and, thus, were overexpressed in the cancers. The different cancer cell lines and leukemias varied in the number and identity of these overexpressed genes. The results indicate that many genes that are overexpressed in human cancer cells are specific to a variety of normal tissues, including normal tissues other than those from which the cancer originated. It is suggested that this general property of cancer cells plays a major role in determining the behavior of the cancers, including their metastatic potential. PMID:16339305

  11. Gene expression in human ovarian tissue after xenografting.

    PubMed

    Van Langendonckt, A; Romeu, L; Ambroise, J; Amorim, C; Bearzatto, B; Gala, J L; Donnez, J; Dolmans, M M

    2014-06-01

    Cryobanking and transplantation of ovarian tissue is a promising approach to restore fertility in cancer patients. However, ischemic stress following avascular ovarian cortex grafting is known to induce stromal tissue fibrosis and alterations in follicular development. The aim of the study was to analyze the impact of freeze-thawing and grafting procedures on gene expression in human ovarian tissue. Frozen-thawed ovarian tissue from 14 patients was xenografted for 7 days to nude mice and one ungrafted fragment was used as a control. Immediately after recovery, grafts were processed for RNA extraction and histological analysis. Their expression profile was screened by whole-genome oligonucleotide array (n = 4) and validated by reverse-transcriptase polymerase chain analysis (n = 10). After data filtering, the Limma package was used to build a linear regression model for each gene and to compute its fold change between tissues on Days 0 and 7. After adjusting the P-value by the Sidak method, 84 of the transcripts were significantly altered after 7 days of grafting, including matrix metalloproteinase-9 and -14 and angiogenic factors such as placental growth factor and C-X-C chemokine receptor type 4 (CXCR4). Major biological processes were related to tissue remodeling, including secretory processes, cellular adhesion and response to chemical and hormonal stimuli. Angiopoietin signaling, the interleukin-8 pathway and peroxisome proliferator-activated receptor activation were shown to be differentially regulated. On Day 7, overexpression was confirmed by PCR for interleukin-8, transforming growth factor-beta 1, matrix metalloproteinase-14 and CXCR4, compared with ungrafted controls. In conclusion, new as well as known genes involved in tissue restructuring and angiogenesis were identified and found to play a key role during the first days after human ovarian tissue transplantation. This will facilitate the development of strategies to optimize grafting techniques. PMID

  12. Reprogramming of the human intestinal epigenome by surgical tissue transposition

    PubMed Central

    Lay, Fides D.; Triche, Timothy J.; Tsai, Yvonne C.; Su, Sheng-Fang; Martin, Sue Ellen; Daneshmand, Siamak; Skinner, Eila C.; Liang, Gangning; Chihara, Yoshitomo; Jones, Peter A.

    2014-01-01

    Extracellular cues play critical roles in the establishment of the epigenome during development and may also contribute to epigenetic perturbations found in disease states. The direct role of the local tissue environment on the post-development human epigenome, however, remains unclear due to limitations in studies of human subjects. Here, we use an isogenic human ileal neobladder surgical model and compare global DNA methylation levels of intestinal epithelial cells pre- and post-neobladder construction using the Infinium HumanMethylation450 BeadChip. Our study is the first to quantify the effect of environmental cues on the human epigenome and show that the local tissue environment directly modulates DNA methylation patterns in normal differentiated cells in vivo. In the neobladder, the intestinal epithelial cells lose their tissue-specific epigenetic landscape in a time-dependent manner following the tissue’s exposure to a bladder environment. We find that de novo methylation of many intestine-specific enhancers occurs at the rate of 0.41% per month (P < 0.01, Pearson = 0.71), while demethylation of primarily non-intestine-specific transcribed regions occurs at the rate of −0.37% per month (P < 0.01, Pearson = −0.57). The dynamic resetting of the DNA methylome in the neobladder not only implicates local environmental cues in the shaping and maintenance of the epigenome but also illustrates an unexpected cross-talk between the epigenome and the cellular environment. PMID:24515120

  13. Tissue-engineered models of human tumors for cancer research

    PubMed Central

    Villasante, Aranzazu; Vunjak-Novakovic, Gordana

    2015-01-01

    Introduction Drug toxicity often goes undetected until clinical trials, which are the most costly and dangerous phase of drug development. Both the cultures of human cells and animal studies have limitations that cannot be overcome by incremental improvements in drug-testing protocols. A new generation of bioengineered tumors is now emerging in response to these limitations, with potential to transform drug screening by providing predictive models of tumors within their tissue context, for studies of drug safety and efficacy. An area that could greatly benefit from these models is cancer research. Areas covered In this review, the authors first describe the engineered tumor systems, using Ewing's sarcoma as an example of human tumor that cannot be predictably studied in cell culture and animal models. Then, they discuss the importance of the tissue context for cancer progression and outline the biomimetic principles for engineering human tumors. Finally, they discuss the utility of bioengineered tumor models for cancer research and address the challenges in modeling human tumors for use in drug discovery and testing. Expert opinion While tissue models are just emerging as a new tool for cancer drug discovery, they are already demonstrating potential for recapitulating, in vitro, the native behavior of human tumors. Still, numerous challenges need to be addressed before we can have platforms with a predictive power appropriate for the pharmaceutical industry. Some of the key needs include the incorporation of the vascular compartment, immune system components, and mechanical signals that regulate tumor development and function. PMID:25662589

  14. Enabling research with human embryonic and fetal tissue resources

    PubMed Central

    Gerrelli, Dianne; Lisgo, Steven; Copp, Andrew J.; Lindsay, Susan

    2015-01-01

    Summary Congenital anomalies are a significant burden on human health. Understanding the developmental origins of such anomalies is key to developing potential therapies. The Human Developmental Biology Resource (HDBR), based in London and Newcastle UK, was established to provide embryonic and fetal material for a variety of human studies ranging from single gene expression analysis to large scale genomic/transcriptomic studies. Increasingly HDBR material is enabling the derivation of stem cell lines and contributing towards developments in tissue engineering. Use of the HDBR and other fetal tissue resources discussed here will contribute to the long term aims of understanding the causation and pathogenesis of congenital anomalies, and developing new methods for their treatment and prevention. PMID:26395135

  15. Profiling RNA editing in human tissues: towards the inosinome Atlas

    PubMed Central

    Picardi, Ernesto; Manzari, Caterina; Mastropasqua, Francesca; Aiello, Italia; D’Erchia, Anna Maria; Pesole, Graziano

    2015-01-01

    Adenine to Inosine RNA editing is a widespread co- and post-transcriptional mechanism mediated by ADAR enzymes acting on double stranded RNA. It has a plethora of biological effects, appears to be particularly pervasive in humans with respect to other mammals, and is implicated in a number of diverse human pathologies. Here we present the first human inosinome atlas comprising 3,041,422 A-to-I events identified in six tissues from three healthy individuals. Matched directional total-RNA-Seq and whole genome sequence datasets were generated and analysed within a dedicated computational framework, also capable of detecting hyper-edited reads. Inosinome profiles are tissue specific and edited gene sets consistently show enrichment of genes involved in neurological disorders and cancer. Overall frequency of editing also varies, but is strongly correlated with ADAR expression levels. The inosinome database is available at: http://srv00.ibbe.cnr.it/editing/. PMID:26449202

  16. Somatic expression of LINE-1 elements in human tissues.

    PubMed

    Belancio, Victoria P; Roy-Engel, Astrid M; Pochampally, Radhika R; Deininger, Prescott

    2010-07-01

    LINE-1 expression damages host DNA via insertions and endonuclease-dependent DNA double-strand breaks (DSBs) that are highly toxic and mutagenic. The predominant tissue of LINE-1 expression has been considered to be the germ line. We show that both full-length and processed L1 transcripts are widespread in human somatic tissues and transformed cells, with significant variation in both L1 expression and L1 mRNA processing. This is the first demonstration that RNA processing is a major regulator of L1 activity. Many tissues also produce translatable spliced transcript (SpORF2). An Alu retrotransposition assay, COMET assays and 53BP1 foci staining show that the SpORF2 product can support functional ORF2 protein expression and can induce DNA damage in normal cells. Tests of the senescence-associated beta-galactosidase expression suggest that expression of exogenous full-length L1, or the SpORF2 mRNA alone in human fibroblasts and adult stem cells triggers a senescence-like phenotype, which is one of the reported responses to DNA damage. In contrast to previous assumptions that L1 expression is germ line specific, the increased spectrum of tissues exposed to L1-associated damage suggests a role for L1 as an endogenous mutagen in somatic tissues. These findings have potential consequences for the whole organism in the form of cancer and mammalian aging.

  17. The human tri-peptide GHK and tissue remodeling.

    PubMed

    Pickart, Loren

    2008-01-01

    Tissue remodeling follows the initial phase of wound healing and stops inflammatory and scar-forming processes, then restores the normal tissue morphology. The human peptide Gly-(L-His)-(L-Lys) or GHK, has a copper 2+ (Cu(2+)) affinity similar to the copper transport site on albumin and forms GHK-Cu, a complex with Cu(2+). These two molecules activate a plethora of remodeling related processes: (1) chemoattraction of repair cells such as macrophages, mast cells, capillary cells; (2) anti-inflammatory actions (suppression of free radicals, thromboxane formation, release of oxidizing iron, transforming growth factor beta-1, tumor necrosis factor alpha and protein glycation while increasing superoxide dismutase, vessel vasodilation, blocking ultraviolet damage to skin keratinocytes and improving fibroblast recovery after X-ray treatments); (3) increases protein synthesis of collagen, elastin, metalloproteinases, anti-proteases, vascular endothelial growth factor, fibroblast growth factor 2, nerve growth factor, neutrotropins 3 and 4, and erythropoietin; (4) increases the proliferation of fibroblasts and keratinocytes; nerve outgrowth, angiogenesis, and hair follicle size. GHK-Cu stimulates wound healing in numerous models and in humans. Controlled studies on aged skin demonstrated that it tightens skin, improves elasticity and firmness, reduces fine lines, wrinkles, photodamage and hyperpigmentation. GHK-Cu also improves hair transplant success, protects hepatic tissue from tetrachloromethane poisoning, blocks stomach ulcer development, and heals intestinal ulcers and bone tissue. These results are beginning to define the complex biochemical processes that regulate tissue remodeling. PMID:18644225

  18. Identification of rheological properties of human body surface tissue.

    PubMed

    Benevicius, Vincas; Gaidys, Rimvydas; Ostasevicius, Vytautas; Marozas, Vaidotas

    2014-04-11

    According to World Health Organization obesity is one of the greatest public health challenges of the 21st century. It has tripled since the 1980s and the numbers of those affected continue to rise at an alarming rate, especially among children. There are number of devices that act as a prevention measure to boost person's motivation for physical activity and its levels. The placement of these devices is not restricted thus the measurement errors that appear because of the body rheology, clothes, etc. cannot be eliminated. The main objective of this work is to introduce a tool that can be applied directly to process measured accelerations so human body surface tissue induced errors can be reduced. Both the modeling and experimental techniques are proposed to identify body tissue rheological properties and prelate them to body mass index. Multi-level computational model composed from measurement device model and human body surface tissue rheological model is developed. Human body surface tissue induced inaccuracies can increase the magnitude of measured accelerations up to 34% when accelerations of the magnitude of up to 27 m/s(2) are measured. Although the timeframe of those disruptions are short - up to 0.2 s - they still result in increased overall measurement error.

  19. FT-Raman spectroscopy study of human breast tissue

    NASA Astrophysics Data System (ADS)

    Bitar Carter, Renata A.; Martin, Airton A.; Netto, Mario M.; Soares, Fernando A.

    2004-07-01

    Optical spectroscopy has been extensively studied as a potential in vivo diagnostic tool to provide information about the chemical and morphologic structure of tissue. Raman Spectroscpy is an inelastic scattering process that can provide a wealth of spectral features that can be related to the specific molecular structure of the sample. This article reports results of an in vitro study of the FT-Raman human breast tissue spectra. An Nd:YAG laser at 1064nm was used as the excitation source in the FT-Raman Spectrometer. The neoplastic human breast samples, both Fibroadenoma and ICD, were obtained during therapeutical routine medical procedures required by the primary disease, and the non-diseased human tissue was obtained in plastic surgery. No sample preparation was needed for the FT-Raman spectra collection. The FT-Raman spectra were recorded from normal, benign (Fibroadenomas) and malignant (IDC-Intraductal Carcinoma) samples, adding up 51 different areas. The main spectral differences of a typical FT-Raman spectra of a Normal (Non-diseased), Fibroadenoma, and Infiltrating Ductal Carcinoma (IDC) breast tissue at the interval of 600 to 1800cm-1, which may differentiate diagnostically the sample, were found in the bands of 1230 to 1295cm-1, 1440 to 1460 cm-1 and 1650 to 1680 cm-1, assigned to the vibrational bands of the carbohydrate-amide III, proteins and lipids, and carbohydrate-amide I, respectively.

  20. Injury Response of Resected Human Brain Tissue In Vitro.

    PubMed

    Verwer, Ronald W H; Sluiter, Arja A; Balesar, Rawien A; Baaijen, Johannes C; de Witt Hamer, Philip C; Speijer, Dave; Li, Yichen; Swaab, Dick F

    2015-07-01

    Brain injury affects a significant number of people each year. Organotypic cultures from resected normal neocortical tissue provide unique opportunities to study the cellular and neuropathological consequences of severe injury of adult human brain tissue in vitro. The in vitro injuries caused by resection (interruption of the circulation) and aggravated by the preparation of slices (severed neuronal and glial processes and blood vessels) reflect the reaction of human brain tissue to severe injury. We investigated this process using immunocytochemical markers, reverse transcriptase quantitative polymerase chain reaction and Western blot analysis. Essential features were rapid shrinkage of neurons, loss of neuronal marker expression and proliferation of reactive cells that expressed Nestin and Vimentin. Also, microglia generally responded strongly, whereas the response of glial fibrillary acidic protein-positive astrocytes appeared to be more variable. Importantly, some reactive cells also expressed both microglia and astrocytic markers, thus confounding their origin. Comparison with post-mortem human brain tissue obtained at rapid autopsies suggested that the reactive process is not a consequence of epilepsy.

  1. Occurrence of human bocaviruses and parvovirus 4 in solid tissues.

    PubMed

    Norja, Päivi; Hedman, Lea; Kantola, Kalle; Kemppainen, Kaisa; Suvilehto, Jari; Pitkäranta, Anne; Aaltonen, Leena-Maija; Seppänen, Mikko; Hedman, Klaus; Söderlund-Venermo, Maria

    2012-08-01

    Human bocaviruses 1-4 (HBoV1-4) and parvovirus 4 (PARV4) are recently discovered human parvoviruses. HBoV1 is associated with respiratory infections of young children, while HBoV2-4 are enteric viruses. The clinical manifestations of PARV4 remain unknown. The objective of this study was to determine whether the DNAs of HBoV1-4 and PARV4 persist in human tissues long after primary infection. Biopsies of tonsillar tissue, skin, and synovia were examined for HBoV1-4 DNA and PARV4 DNA by PCR. Serum samples from the tissue donors were assayed for HBoV1 and PARV4 IgG and IgM antibodies. To obtain species-specific seroprevalences for HBoV1 and for HBoV2/3 combined, the sera were analyzed after virus-like particle (VLP) competition. While HBoV1 DNA was detected exclusively in the tonsillar tissues of 16/438 individuals (3.7%), all of them ≤8 years of age. HBoV2-4 and PARV4 DNAs were absent from all tissue types. HBoV1 IgG seroprevalence was 94.9%. No subject had HBoV1 or PARV4 IgM, nor did they have PARV4 IgG. The results indicate that HBoV1 DNA occurred in a small proportion of tonsils of young children after recent primary HBoV1 infection, but did not persist long in the other tissue types studied, unlike parvovirus B19 DNA. The results obtained by the PARV4 assays are in line with previous results on PARV4 epidemiology.

  2. Tissue-engineered microenvironment systems for modeling human vasculature.

    PubMed

    Tourovskaia, Anna; Fauver, Mark; Kramer, Gregory; Simonson, Sara; Neumann, Thomas

    2014-09-01

    The high attrition rate of drug candidates late in the development process has led to an increasing demand for test assays that predict clinical outcome better than conventional 2D cell culture systems and animal models. Government agencies, the military, and the pharmaceutical industry have started initiatives for the development of novel in-vitro systems that recapitulate functional units of human tissues and organs. There is growing evidence that 3D cell arrangement, co-culture of different cell types, and physico-chemical cues lead to improved predictive power. A key element of all tissue microenvironments is the vasculature. Beyond transporting blood the microvasculature assumes important organ-specific functions. It is also involved in pathologic conditions, such as inflammation, tumor growth, metastasis, and degenerative diseases. To provide a tool for modeling this important feature of human tissue microenvironments, we developed a microfluidic chip for creating tissue-engineered microenvironment systems (TEMS) composed of tubular cell structures. Our chip design encompasses a small chamber that is filled with an extracellular matrix (ECM) surrounding one or more tubular channels. Endothelial cells (ECs) seeded into the channels adhere to the ECM walls and grow into perfusable tubular tissue structures that are fluidically connected to upstream and downstream fluid channels in the chip. Using these chips we created models of angiogenesis, the blood-brain barrier (BBB), and tumor-cell extravasation. Our angiogenesis model recapitulates true angiogenesis, in which sprouting occurs from a "parent" vessel in response to a gradient of growth factors. Our BBB model is composed of a microvessel generated from brain-specific ECs within an ECM populated with astrocytes and pericytes. Our tumor-cell extravasation model can be utilized to visualize and measure tumor-cell migration through vessel walls into the surrounding matrix. The described technology can be used

  3. Effects of laser interaction with living human tissues

    NASA Astrophysics Data System (ADS)

    Molchanova, O. E.; Protasov, E. A.; Protasov, D. E.; Smirnova, A. V.

    2016-09-01

    With the help of a highly sensitive laser device with the wavelength λ = 0.808 pm, which is optimal for deep penetration of the radiation into biological tissues, the effects associated with the appearance of uncontrolled human infrasonic vibrations of different frequencies were investigated. It was established that the observed fluctuations are associated with the vascular system which is characterized by its own respiratory movements, occurring synchronously with the movements of the respiratory muscles, the operation of the heart muscle, and the effect of compression ischemia. The effect of “enlightenment” of a tissue is observed with stopping of blood flow in vessels by applying a tourniquet on the wrist.

  4. Detection of renal tissue and urinary tract proteins in the human urine after space flight.

    PubMed

    Pastushkova, Lyudmila Kh; Kireev, Kirill S; Kononikhin, Alexey S; Tiys, Evgeny S; Popov, Igor A; Starodubtseva, Natalia L; Dobrokhotov, Igor V; Ivanisenko, Vladimir A; Larina, Irina M; Kolchanov, Nicolay A; Nikolaev, Evgeny N

    2013-01-01

    The urine protein composition samples of ten Russian cosmonauts (male, aged of 35 up to 51) performed long flight missions and varied from 169 up to 199 days on the International Space Station (ISS) were analyzed. As a control group, urine samples of six back-up cosmonauts were analyzed. We used proteomic techniques to obtain data and contemporary bioinformatics approaches to perform the analysis. From the total number of identified proteins (238) in our data set, 129 were associated with a known tissue origin. Preflight samples contained 92 tissue-specific proteins, samples obtained on Day 1 after landing had 90 such proteins, while Day 7 samples offered 95 tissue-specific proteins. Analysis showed that consistently present proteins in urine (under physiological conditions and after space flight) are cubilin, epidermal growth factor, kallikrein-1, kininogen-1, megalin, osteopontin, vitamin K-dependent protein Z, uromodulin. Variably present proteins consists of: Na(+)/K(+) ATPase subunit gamma, β-defensin-1, dipeptidyl peptidase 4, maltasa-glucoamilasa, cadherin-like protein, neutral endopeptidase and vascular cell adhesion protein 1. And only three renal proteins were related to the space flight factors. They were not found in the pre-flight samples and in the back-up cosmonaut urine, but were found in the urine samples after space flight: AFAM (afamin), AMPE (aminopeptidase A) and AQP2 (aquaporin-2). This data related with physiological readaptation of water-salt balance. The proteomic analysis of urine samples in different phases of space missions with bioinformation approach to protein identification provides new data relative to biomechemical mechanism of kidney functioning after space flight.

  5. Detection of Renal Tissue and Urinary Tract Proteins in the Human Urine after Space Flight

    PubMed Central

    Pastushkova, Lyudmila Kh.; Kireev, Kirill S.; Kononikhin, Alexey S.; Tiys, Evgeny S.; Popov, Igor A.; Starodubtseva, Natalia L.; Dobrokhotov, Igor V.; Ivanisenko, Vladimir A.; Larina, Irina M.; Kolchanov, Nicolay A.; Nikolaev, Evgeny N.

    2013-01-01

    The urine protein composition samples of ten Russian cosmonauts (male, aged of 35 up to 51) performed long flight missions and varied from 169 up to 199 days on the International Space Station (ISS) were analyzed. As a control group, urine samples of six back-up cosmonauts were analyzed. We used proteomic techniques to obtain data and contemporary bioinformatics approaches to perform the analysis. From the total number of identified proteins (238) in our data set, 129 were associated with a known tissue origin. Preflight samples contained 92 tissue-specific proteins, samples obtained on Day 1 after landing had 90 such proteins, while Day 7 samples offered 95 tissue-specific proteins. Analysis showed that consistently present proteins in urine (under physiological conditions and after space flight) are cubilin, epidermal growth factor, kallikrein-1, kininogen-1, megalin, osteopontin, vitamin K-dependent protein Z, uromodulin. Variably present proteins consists of: Na(+)/K(+) ATPase subunit gamma, β-defensin-1, dipeptidyl peptidase 4, maltasa-glucoamilasa, cadherin-like protein, neutral endopeptidase and vascular cell adhesion protein 1. And only three renal proteins were related to the space flight factors. They were not found in the pre-flight samples and in the back-up cosmonaut urine, but were found in the urine samples after space flight: AFAM (afamin), AMPE (aminopeptidase A) and AQP2 (aquaporin-2). This data related with physiological readaptation of water-salt balance. The proteomic analysis of urine samples in different phases of space missions with bioinformation approach to protein identification provides new data relative to biomechemical mechanism of kidney functioning after space flight. PMID:23967230

  6. Engineering of human hepatic tissue with functional vascular networks.

    PubMed

    Takebe, Takanori; Koike, Naoto; Sekine, Keisuke; Fujiwara, Ryoji; Amiya, Takeru; Zheng, Yun-Wen; Taniguchi, Hideki

    2014-01-01

    Although absolute organ shortage highlights the needs of alternative organ sources for regenerative medicine, the generation of a three-dimensional (3D) and complex vital organ, such as well-vascularized liver, remains a challenge. To this end, tissue engineering holds great promise; however, this approach is significantly limited by the failure of early vascularization in vivo after implantation. Here, we established a stable 3D in vitro pre-vascularization platform to generate human hepatic tissue after implantation in vivo. Human fetal liver cells (hFLCs) were mixed with human umbilical vein endothelial cells (HUVECs) and mesenchymal stem cells (hMSCs) and were implanted into a collagen/fibronectin matrix composite that was used as a 3-D carrier. After a couple of days, the fluorescent HUVECs developed premature vascular networks in vitro, which were stabilized by hMSCs. The establishment of functional vessels inside the pre-vascularized constructs was proven using dextran infusion studies after implantation under a transparency cranial window. Furthermore, dynamic morphological changes during embryonic liver cell maturation were intravitaly quantified with high-resolution confocal microscope analysis. The engineered human hepatic tissue demonstrated multiple liver-specific features, both structural and functional. Our new techniques discussed here can be implemented in future clinical uses and industrial uses, such as drug testing. PMID:24451152

  7. Engineering of human hepatic tissue with functional vascular networks

    PubMed Central

    Takebe, Takanori; Koike, Naoto; Sekine, Keisuke; Fujiwara, Ryoji; Amiya, Takeru; Zheng, Yun-Wen; Taniguchi, Hideki

    2014-01-01

    Although absolute organ shortage highlights the needs of alternative organ sources for regenerative medicine, the generation of a three-dimensional (3D) and complex vital organ, such as well-vascularized liver, remains a challenge. To this end, tissue engineering holds great promise; however, this approach is significantly limited by the failure of early vascularization in vivo after implantation. Here, we established a stable 3D in vitro pre-vascularization platform to generate human hepatic tissue after implantation in vivo. Human fetal liver cells (hFLCs) were mixed with human umbilical vein endothelial cells (HUVECs) and mesenchymal stem cells (hMSCs) and were implanted into a collagen/fibronectin matrix composite that was used as a 3-D carrier. After a couple of days, the fluorescent HUVECs developed premature vascular networks in vitro, which were stabilized by hMSCs. The establishment of functional vessels inside the pre-vascularized constructs was proven using dextran infusion studies after implantation under a transparency cranial window. Furthermore, dynamic morphological changes during embryonic liver cell maturation were intravitaly quantified with high-resolution confocal microscope analysis. The engineered human hepatic tissue demonstrated multiple liver-specific features, both structural and functional. Our new techniques discussed here can be implemented in future clinical uses and industrial uses, such as drug testing. PMID:24451152

  8. Induction of tissue transglutaminase in human peripheral blood monocytes

    PubMed Central

    1984-01-01

    The levels and activity of tissue transglutaminase were studied in human peripheral blood monocytes during differentiation into macrophages in vitro. The enzyme was present at low levels in freshly isolated monocytes (less than 20 ng/mg cell protein) but increased 50- fold during 10 d of adherent culture in autologous serum, reaching levels of 0.1% of total cellular protein. The rate of appearance of tissue transglutaminase in monocytes was accelerated by low levels of lipopolysaccharide. The half-life of disappearance of transglutaminase from human monocytes was 11 and 7 h in 2-d-old and 10-d-old cells, respectively. Treatment of 1-day-old monocytes with actinomycin D for 24 h blocked the increase in transglutaminase levels. These results indicated that the induction of gene transcription and protein synthesis was responsible for the increased transglutaminase levels and activity observed with cultured human monocytes. The induction of tissue transglutaminase may be a component in the in vivo differentiation of human monocytes into macrophages. PMID:6141210

  9. Engineering of human hepatic tissue with functional vascular networks.

    PubMed

    Takebe, Takanori; Koike, Naoto; Sekine, Keisuke; Fujiwara, Ryoji; Amiya, Takeru; Zheng, Yun-Wen; Taniguchi, Hideki

    2014-01-01

    Although absolute organ shortage highlights the needs of alternative organ sources for regenerative medicine, the generation of a three-dimensional (3D) and complex vital organ, such as well-vascularized liver, remains a challenge. To this end, tissue engineering holds great promise; however, this approach is significantly limited by the failure of early vascularization in vivo after implantation. Here, we established a stable 3D in vitro pre-vascularization platform to generate human hepatic tissue after implantation in vivo. Human fetal liver cells (hFLCs) were mixed with human umbilical vein endothelial cells (HUVECs) and mesenchymal stem cells (hMSCs) and were implanted into a collagen/fibronectin matrix composite that was used as a 3-D carrier. After a couple of days, the fluorescent HUVECs developed premature vascular networks in vitro, which were stabilized by hMSCs. The establishment of functional vessels inside the pre-vascularized constructs was proven using dextran infusion studies after implantation under a transparency cranial window. Furthermore, dynamic morphological changes during embryonic liver cell maturation were intravitaly quantified with high-resolution confocal microscope analysis. The engineered human hepatic tissue demonstrated multiple liver-specific features, both structural and functional. Our new techniques discussed here can be implemented in future clinical uses and industrial uses, such as drug testing.

  10. Maresin conjugates in tissue regeneration biosynthesis enzymes in human macrophages

    PubMed Central

    Dalli, Jesmond; Vlasakov, Iliyan; Riley, Ian R.; Rodriguez, Ana R.; Spur, Bernd W.; Chiang, Nan; Serhan, Charles N.

    2016-01-01

    Macrophages are central in coordinating immune responses, tissue repair, and regeneration, with different subtypes being associated with inflammation-initiating and proresolving actions. We recently identified a family of macrophage-derived proresolving and tissue regenerative molecules coined maresin conjugates in tissue regeneration (MCTR). Herein, using lipid mediator profiling we identified MCTR in human serum, lymph nodes, and plasma and investigated MCTR biosynthetic pathways in human macrophages. With human recombinant enzymes, primary cells, and enantiomerically pure compounds we found that the synthetic maresin epoxide intermediate 13S,14S-eMaR (13S,14S-epoxy- 4Z,7Z,9E,11E,16Z,19Z-docosahexaenoic acid) was converted to MCTR1 (13R-glutathionyl, 14S-hydroxy-4Z,7Z,9E,11E,13R,14S,16Z,19Z-docosahexaenoic acid) by LTC4S and GSTM4. Incubation of human macrophages with LTC4S inhibitors blocked LTC4 and increased resolvins and lipoxins. The conversion of MCTR1 to MCTR2 (13R-cysteinylglycinyl, 14S-hydroxy-4Z,7Z,9E,11E,13R,14S,16Z,19Z-docosahexaenoic acid) was catalyzed by γ-glutamyl transferase (GGT) in human macrophages. Biosynthesis of MCTR3 was mediated by dipeptidases that cleaved the cysteinyl-glycinyl bond of MCTR2 to give 13R-cysteinyl, 14S-hydroxy-4Z,7Z,9E,11E,13R,14S,16Z,19Z-docosahexaenoic acid. Of note, both GSTM4 and GGT enzymes displayed higher affinity to 13S,14S-eMaR and MCTR1 compared with their classic substrates in the cysteinyl leukotriene metabolome. Together these results establish the MCTR biosynthetic pathway and provide mechanisms in tissue repair and regeneration. PMID:27791009

  11. Soft tissues store and return mechanical energy in human running.

    PubMed

    Riddick, R C; Kuo, A D

    2016-02-01

    During human running, softer parts of the body may deform under load and dissipate mechanical energy. Although tissues such as the heel pad have been characterized individually, the aggregate work performed by all soft tissues during running is unknown. We therefore estimated the work performed by soft tissues (N=8 healthy adults) at running speeds ranging 2-5 m s(-1), computed as the difference between joint work performed on rigid segments, and whole-body estimates of work performed on the (non-rigid) body center of mass (COM) and peripheral to the COM. Soft tissues performed aggregate negative work, with magnitude increasing linearly with speed. The amount was about -19 J per stance phase at a nominal 3 m s(-1), accounting for more than 25% of stance phase negative work performed by the entire body. Fluctuations in soft tissue mechanical power over time resembled a damped oscillation starting at ground contact, with peak negative power comparable to that for the knee joint (about -500 W). Even the positive work from soft tissue rebound was significant, about 13 J per stance phase (about 17% of the positive work of the entire body). Assuming that the net dissipative work is offset by an equal amount of active, positive muscle work performed at 25% efficiency, soft tissue dissipation could account for about 29% of the net metabolic expenditure for running at 5 m s(-1). During running, soft tissue deformations dissipate mechanical energy that must be offset by active muscle work at non-negligible metabolic cost. PMID:26806689

  12. Monitoring changes in tissue optical properties following interstitial photothermal therapy of ex vivo human prostate tissue

    NASA Astrophysics Data System (ADS)

    Weersink, Robert A.; He, Jie; Veilleux, Israel; Trachtenberg, John; Wilson, Brian C.

    2013-03-01

    We are developing a method of monitoring treatment progression of interstitial photothermal therapy of focal prostate cancer using transrectal diffuse optical tomography (TRDOT) combined with transrectal 3D ultrasound (3D-TRUS). Measurements of prostate tissue optical properties were made on ex vivo human prostate samples prior to and post coagulation. Interstitial photothermal treatments were delivered to the ex vivo samples and monitored using an interstitial probe near the treatment fiber. After treatment, bulk optical properties were measured on native and coagulated zones of tissue. Changes in optical properties across the boundary between native and coagulated tissues were spatially mapped using a small diffuse reflectance probe. The optical property estimates and spatial information obtained using each method was compared.

  13. An Introduction to The Royan Human Ovarian Tissue Bank.

    PubMed

    Abtahi, Naeimeh Sadat; Ebrahimi, Bita; Fathi, Rouhollah; Khodaverdi, Sepideh; Mehdizadeh Kashi, Abolfazl; Valojerdi, Mojtaba Rezazadeh

    2016-01-01

    From December 2000 until 2010, the researchers at Royan Institute conducted a wide range of investigations on ovarian tissue cryopreservation with the intent to provide fertility pres- ervation to cancer patients that were considered to be candidates for these services. In 2010, Royan Institute established the Royan Human Ovarian Tissue Bank as a subgroup of the Embryology Department. Since its inception, approximately 180 patients between the ages of 747 years have undergone consultations. Ovarian samples were cryopreserved from 47 patients (age: 7-35 years) diagnosed with cervical adenocarcinoma (n=9); breast carcinoma (n=7), Ewing's sarcoma (n=7), opposite side ovarian tumor (n=7), endometrial adenocarci- noma (n=4), malignant colon tumors (n=3), as well as Hodgkin's lymphoma, major thalas- semia and acute lymphoblastic leukemia (n=1-2 patients for each disease). Additionally, two patients requested ovarian tissue transplantation after completion of their treatments. PMID:27441061

  14. An Introduction to The Royan Human Ovarian Tissue Bank

    PubMed Central

    Abtahi, Naeimeh Sadat; Ebrahimi, Bita; Fathi, Rouhollah; Khodaverdi, Sepideh; Mehdizadeh Kashi, Abolfazl; Valojerdi, Mojtaba Rezazadeh

    2016-01-01

    From December 2000 until 2010, the researchers at Royan Institute conducted a wide range of investigations on ovarian tissue cryopreservation with the intent to provide fertility pres- ervation to cancer patients that were considered to be candidates for these services. In 2010, Royan Institute established the Royan Human Ovarian Tissue Bank as a subgroup of the Embryology Department. Since its inception, approximately 180 patients between the ages of 747 years have undergone consultations. Ovarian samples were cryopreserved from 47 patients (age: 7-35 years) diagnosed with cervical adenocarcinoma (n=9); breast carcinoma (n=7), Ewing’s sarcoma (n=7), opposite side ovarian tumor (n=7), endometrial adenocarci- noma (n=4), malignant colon tumors (n=3), as well as Hodgkin’s lymphoma, major thalas- semia and acute lymphoblastic leukemia (n=1-2 patients for each disease). Additionally, two patients requested ovarian tissue transplantation after completion of their treatments. PMID:27441061

  15. Fallout sup 3 H in human tissue at Akita, Japan

    SciTech Connect

    Hisamatsu, S.; Takizawa, Y.; Itoh, M.; Ueno, K.; Katsumata, T.; Sakanoue, M. )

    1989-10-01

    The {sup 3}H concentration in Japanese human tissue samples is reported in this paper. Four brain, 10 liver, and nine lung samples from 11 cases were collected from Akita Prefecture in northern Japan from January to July 1986. The median of free-water {sup 3}H concentration was similar in these tissues and agreed well with the concentrations in the diet, including tap water. The median specific activity ratio of tissue-bound {sup 3}H to free-water {sup 3}H was 1.1 and was slightly lower than that in the diet. The specific activity ratio was also lower than that reported in the United States and significantly lower than in Italy.

  16. A continuous fiber distribution material model for human cervical tissue.

    PubMed

    Myers, Kristin M; Hendon, Christine P; Gan, Yu; Yao, Wang; Yoshida, Kyoko; Fernandez, Michael; Vink, Joy; Wapner, Ronald J

    2015-06-25

    The uterine cervix during pregnancy is the vital mechanical barrier which resists compressive and tensile loads generated from a growing fetus. Premature cervical remodeling and softening is hypothesized to result in the shortening of the cervix, which is known to increase a woman׳s risk of preterm birth. To understand the role of cervical material properties in preventing preterm birth, we derive a cervical material model based on previous mechanical, biochemical and histological experiments conducted on nonpregnant and pregnant human hysterectomy cervical tissue samples. In this study we present a three-dimensional fiber composite model that captures the equilibrium material behavior of the tissue in tension and compression. Cervical tissue is modeled as a fibrous composite material, where a single family of preferentially aligned and continuously distributed collagen fibers are embedded in a compressible neo-Hookean ground substance. The total stress in the collagen solid network is calculated by integrating the fiber stresses. The shape of the fiber distribution is described by an ellipsoid where semi-principal axis lengths are fit to optical coherence tomography measurements. The composite material model is fit to averaged mechanical testing data from uni-axial compression and tension experiments, and averaged material parameters are reported for nonpregnant and term pregnant human cervical tissue. The model is then evaluated by investigating the stress and strain state of a uniform thick-walled cylinder under a compressive stress with collagen fibers preferentially aligned in the circumferential direction. This material modeling framework for the equilibrium behavior of human cervical tissue serves as a basis to determine the role of preferentially-aligned cervical collagen fibers in preventing cervical deformation during pregnancy.

  17. Levels of chlordane, oxychlordane, and nonachlor in human adipose tissues

    SciTech Connect

    Hirai, Yukio; Tomokuni, Katsumaro )

    1991-08-01

    Chlordane was used as a termiticide for more than twenty years in Japan. Chlordane is stable in the environment such as sediment and its bioaccumulation in some species of bacteria, freshwater invertebrates, and marine fish is large. Many researches were done to elucidate the levels of chlordane and/or its metabolite oxychlordane in human adipose tissues. A comprehensive review concerning chlordane was recently provided by USEPA. On the other hand, Japan authorities banned the use of chlordane in September 1986. In the last paper, the authors reported that both water and sediment of the rivers around Saga city were slightly contaminated with chlordane. In the present study, they investigated the levels of chlordane, oxychlordane and nonachlor in human adipose tissues.

  18. Engineering human neo-tendon tissue in vitro with human dermal fibroblasts under static mechanical strain.

    PubMed

    Deng, Dan; Liu, Wei; Xu, Feng; Yang, Yang; Zhou, Guangdong; Zhang, Wen Jie; Cui, Lei; Cao, Yilin

    2009-12-01

    Proper cell source is one of the key issues for tendon engineering. Our previous study showed that dermal fibroblasts could be used to successfully engineer tendon in vivo and tenocytes could engineer neo-tendon in vitro with static strain. This study further investigated the possibility of engineering human neo-tendon tissue in vitro using dermal fibroblasts. Human dermal fibroblasts were seeded on polyglycolic acid (PGA) fibers pre-fixed on a U-shape as a mechanical loading group, or simply cultured in a dish as a tension-free group. In addition, human tenocytes were also seeded on PGA fibers with tension as a comparison to human dermal fibroblasts. The results showed that human neo-tendon tissue could be generated using dermal fibroblasts during in vitro culture under static strain and the tissue structure became more mature with the increase of culture time. Longitudinally aligned collagen fibers and spindle shape cells were observed histologically and collagen fibril diameter and tensile strength increased with time and reached a peak at 14 weeks. In contrast, the dermal fibroblast-PGA constructs failed to form neo-tendon, but formed disorganized fibrous tissue in tension-free condition with significantly weaker strength and poor collagen fiber formation. Interestingly, neo-tendon tissues generated with human dermal fibroblasts were indistinguishable from the counterpart engineered with human tenocytes, which supports the viewpoint that human dermal fibroblasts is likely to replace tenocytes for future tendon graft development in vitro with dynamic mechanical loading in a bioreactor system.

  19. Two types of brown adipose tissue in humans.

    PubMed

    Lidell, Martin E; Betz, Matthias J; Enerbäck, Sven

    2014-01-01

    During the last years the existence of metabolically active brown adipose tissue in adult humans has been widely accepted by the research community. Its unique ability to dissipate chemical energy stored in triglycerides as heat makes it an attractive target for new drugs against obesity and its related diseases. Hence the tissue is now subject to intense research, the hypothesis being that an expansion and/or activation of the tissue is associated with a healthy metabolic phenotype. Animal studies provide evidence for the existence of at least two types of brown adipocytes. Apart from the classical brown adipocyte that is found primarily in the interscapular region where it constitutes a thermogenic organ, a second type of brown adipocyte, the so-called beige adipocyte, can appear within white adipose tissue depots. The fact that the two cell types develop from different precursors suggests that they might be recruited and stimulated by different cues and therefore represent two distinct targets for therapeutic intervention. The aim of this commentary is to discuss recent work addressing the question whether also humans possess two types of brown adipocytes and to highlight some issues when looking for molecular markers for such cells.

  20. Expression of the Endocannabinoid Receptors in Human Fascial Tissue

    PubMed Central

    Fede, C.; Albertin, G.; Petrelli, L.; Sfriso, M.M.; Biz, C.; Caro, R. De; Stecco, C.

    2016-01-01

    Cannabinoid receptors have been localized in the central and peripheral nervous system as well as on cells of the immune system, but recent studies on animal tissue gave evidence for the presence of cannabinoid receptors in different types of tissues. Their presence was supposed also in myofascial tissue, suggesting that the endocannabinoid system may help resolve myofascial trigger points and relieve symptoms of fibromyalgia. However, until now the expression of CB1 (cannabinoid receptor 1) and CB2 (cannabinoid receptor 2) in fasciae has not yet been established. Small samples of fascia were collected from volunteers patients during orthopedic surgery. For each sample were done a cell isolation, immunohistochemical investigation (CB1 and CB2 antibodies) and real time RT-PCR to detect the expression of CB1 and CB2. Both cannabinoid receptors are expressed in human fascia and in human fascial fibroblasts culture cells, although to a lesser extent than the control gene. We can assume that the expression of mRNA and protein of CB1 and CB2 receptors in fascial tissue are concentrated into the fibroblasts. This is the first demonstration that the fibroblasts of the muscular fasciae express CB1 and CB2. The presence of these receptors could help to provide a description of cannabinoid receptors distribution and to better explain the role of fasciae as pain generator and the efficacy of some fascial treatments. Indeed the endocannabinoid receptors of fascial fibroblasts can contribute to modulate the fascial fibrosis and inflammation. PMID:27349320

  1. Human omental and subcutaneous adipose tissue exhibit specific lipidomic signatures.

    PubMed

    Jové, Mariona; Moreno-Navarrete, José María; Pamplona, Reinald; Ricart, Wifredo; Portero-Otín, Manuel; Fernández-Real, José Manuel

    2014-03-01

    Despite their differential effects on human metabolic pathophysiology, the differences in omental and subcutaneous lipidomes are largely unknown. To explore this field, liquid chromatography coupled with mass spectrometry was used for lipidome analyses of adipose tissue samples (visceral and subcutaneous) selected from a group of obese subjects (n=38). Transcriptomics and in vitro studies in adipocytes were used to confirm the pathways affected by location. The analyses revealed the existence of obesity-related specific lipidome signatures in each of these locations, attributed to selective enrichment of specific triglycerides, glycerophospholipids, and sphingolipids, because these were not observed in adipose tissues from nonobese individuals. The changes were compatible with subcutaneous enrichment in pathways involved in adipogenesis, triacylglyceride synthesis, and lipid droplet formation, as well as increased α-oxidation. Marked differences between omental and subcutaneous depots in obese individuals were seen in the association of lipid species with metabolic traits (body mass index and insulin sensitivity). Targeted studies also revealed increased cholesterol (Δ56%) and cholesterol epoxide (Δ34%) concentrations in omental adipose tissue. In view of the effects of cholesterol epoxide, which induced enhanced expression of adipocyte differentiation and α-oxidation genes in human omental adipocytes, a novel role for cholesterol epoxide as a signaling molecule for differentiation is proposed. In summary, in obesity, adipose tissue exhibits a location-specific differential lipid profile that may contribute to explaining part of its distinct pathogenic role.

  2. Effect of various concentrations of caffeine, pentoxifylline, and kallikrein on hyperactivation of frozen bovine semen.

    PubMed

    Barakat, Ibrahim A H; Danfour, Mohamed A; Galewan, Fatma A M; Dkhil, Mohamed A

    2015-01-01

    Caffeine, pentoxifylline, and kallikrein are substances that affect the efficiency of sperms in the fertilization process; however, they have not been adequately studied. The present study aimed to examine the influence of caffeine, kallikrein, and pentoxifylline on sperm motility in bovine as well as investigate their optimum concentrations for increasing the movement of sperms in bovine. Frozen bovine sperms were thawed in universal IVF medium supplemented with 1, 5, and 10 mM caffeine or pentoxifylline or 1, 4, and 8 U/mL kallikrein and were then incubated for 30 min. Treated semen parameters were analyzed using a computer assisted semen analyzer (CASA). Data analysis showed that the mean values concerning progression and motility of sperm increased in caffeine and pentoxifylline treatments when compared with the kallikrein group. The obtained results revealed that kallikrein is not necessary for the improvement of bovine sperm motility. Additionally, our results revealed that 5 mM from caffeine was the best concentration added to the medium, followed by 1 or 5 mM from pentoxifylline. Therefore, it is concluded from the present study that caffeine has hyperactivation efficacy at 5 mM concentration compared to other treatments.

  3. Effect of Various Concentrations of Caffeine, Pentoxifylline, and Kallikrein on Hyperactivation of Frozen Bovine Semen

    PubMed Central

    Barakat, Ibrahim A. H.; Danfour, Mohamed A.; Galewan, Fatma A. M.; Dkhil, Mohamed A.

    2015-01-01

    Caffeine, pentoxifylline, and kallikrein are substances that affect the efficiency of sperms in the fertilization process; however, they have not been adequately studied. The present study aimed to examine the influence of caffeine, kallikrein, and pentoxifylline on sperm motility in bovine as well as investigate their optimum concentrations for increasing the movement of sperms in bovine. Frozen bovine sperms were thawed in universal IVF medium supplemented with 1, 5, and 10 mM caffeine or pentoxifylline or 1, 4, and 8 U/mL kallikrein and were then incubated for 30 min. Treated semen parameters were analyzed using a computer assisted semen analyzer (CASA). Data analysis showed that the mean values concerning progression and motility of sperm increased in caffeine and pentoxifylline treatments when compared with the kallikrein group. The obtained results revealed that kallikrein is not necessary for the improvement of bovine sperm motility. Additionally, our results revealed that 5 mM from caffeine was the best concentration added to the medium, followed by 1 or 5 mM from pentoxifylline. Therefore, it is concluded from the present study that caffeine has hyperactivation efficacy at 5 mM concentration compared to other treatments. PMID:25950005

  4. Cervical Tissue Engineering Using Silk Scaffolds and Human Cervical Cells

    PubMed Central

    Sanchez, Cristina C.; Rice, William L.; Socrate, Simona; Kaplan, David L.

    2010-01-01

    Spontaneous preterm birth is a frequent complication of pregnancy and a common cause of morbidity in childhood. Obstetricians suspect abnormalities of the cervix are implicated in a significant number of preterm births. The cervix is composed of fibrous connective tissue and undergoes significant remodeling in preparation for birth. We hypothesized that a tissue engineering strategy could be used to develop three-dimensional cervical-like tissue constructs that would be suitable for investigating cervical remodeling. Cervical cells were isolated from two premenopausal women undergoing hysterectomy for a benign gynecological condition, and the cells were seeded on porous silk scaffolds in the presence or absence of dynamic culture and with 10% or 20% serum. Morphological, biochemical, and mechanical properties were measured during the 8-week culture period. Cervical cells proliferated in three-dimensions and synthesized an extracellular matrix with biochemical constituents and morphology similar to native tissue. Compared to static culture, dynamic culture was associated with significantly increased collagen deposition (p < 0.05), sulfated glycosaminoglycan synthesis (p < 0.05), and mechanical stiffness (p < 0.05). Serum concentration did not affect measured variables. Relevant human tissue-engineered cervical-like constructs constitute a novel model system for a range of fundamental and applied studies related to cervical remodeling. PMID:20121593

  5. Streamlined bioreactor-based production of human cartilage tissues.

    PubMed

    Tonnarelli, B; Santoro, R; Adelaide Asnaghi, M; Wendt, D

    2016-05-27

    Engineered tissue grafts have been manufactured using methods based predominantly on traditional labour-intensive manual benchtop techniques. These methods impart significant regulatory and economic challenges, hindering the successful translation of engineered tissue products to the clinic. Alternatively, bioreactor-based production systems have the potential to overcome such limitations. In this work, we present an innovative manufacturing approach to engineer cartilage tissue within a single bioreactor system, starting from freshly isolated human primary chondrocytes, through the generation of cartilaginous tissue grafts. The limited number of primary chondrocytes that can be isolated from a small clinically-sized cartilage biopsy could be seeded and extensively expanded directly within a 3D scaffold in our perfusion bioreactor (5.4 ± 0.9 doublings in 2 weeks), bypassing conventional 2D expansion in flasks. Chondrocytes expanded in 3D scaffolds better maintained a chondrogenic phenotype than chondrocytes expanded on plastic flasks (collagen type II mRNA, 18-fold; Sox-9, 11-fold). After this "3D expansion" phase, bioreactor culture conditions were changed to subsequently support chondrogenic differentiation for two weeks. Engineered tissues based on 3D-expanded chondrocytes were more cartilaginous than tissues generated from chondrocytes previously expanded in flasks. We then demonstrated that this streamlined bioreactor-based process could be adapted to effectively generate up-scaled cartilage grafts in a size with clinical relevance (50 mm diameter). Streamlined and robust tissue engineering processes, as the one described here, may be key for the future manufacturing of grafts for clinical applications, as they facilitate the establishment of compact and closed bioreactor-based production systems, with minimal automation requirements, lower operating costs, and increased compliance to regulatory guidelines.

  6. Streamlined bioreactor-based production of human cartilage tissues.

    PubMed

    Tonnarelli, B; Santoro, R; Adelaide Asnaghi, M; Wendt, D

    2016-01-01

    Engineered tissue grafts have been manufactured using methods based predominantly on traditional labour-intensive manual benchtop techniques. These methods impart significant regulatory and economic challenges, hindering the successful translation of engineered tissue products to the clinic. Alternatively, bioreactor-based production systems have the potential to overcome such limitations. In this work, we present an innovative manufacturing approach to engineer cartilage tissue within a single bioreactor system, starting from freshly isolated human primary chondrocytes, through the generation of cartilaginous tissue grafts. The limited number of primary chondrocytes that can be isolated from a small clinically-sized cartilage biopsy could be seeded and extensively expanded directly within a 3D scaffold in our perfusion bioreactor (5.4 ± 0.9 doublings in 2 weeks), bypassing conventional 2D expansion in flasks. Chondrocytes expanded in 3D scaffolds better maintained a chondrogenic phenotype than chondrocytes expanded on plastic flasks (collagen type II mRNA, 18-fold; Sox-9, 11-fold). After this "3D expansion" phase, bioreactor culture conditions were changed to subsequently support chondrogenic differentiation for two weeks. Engineered tissues based on 3D-expanded chondrocytes were more cartilaginous than tissues generated from chondrocytes previously expanded in flasks. We then demonstrated that this streamlined bioreactor-based process could be adapted to effectively generate up-scaled cartilage grafts in a size with clinical relevance (50 mm diameter). Streamlined and robust tissue engineering processes, as the one described here, may be key for the future manufacturing of grafts for clinical applications, as they facilitate the establishment of compact and closed bioreactor-based production systems, with minimal automation requirements, lower operating costs, and increased compliance to regulatory guidelines. PMID:27232665

  7. Insulin is ubiquitous in extrapancreatic tissues of rats and humans.

    PubMed Central

    Rosenzweig, J L; Havrankova, J; Lesniak, M A; Brownstein, M; Roth, J

    1980-01-01

    Insulin has been detected, at levels higher than those in plasma, in a broad range of extrapancreatic tissues in both rats and humans. Rat liver insulin was shown to be indistinguishable from genuine insulin by radioimmunoassay, Sephadex chromatography, bioassay, and antibody neutralization. Liver insulin (like brain insulin) was unchanged in ob/ob mice, in rats treated with streptozotocin, or in fasted rats, despite marked alterations in pancreatic secretion of insulin and in liver content of insulin receptors. Insulin was found in cultured human IM-9 lymphocytes and cultured fibroblasts at concentrations greater than 100 times the levels in the media. IM-9 lymphocyte insulin also was shown to be indistinguishable from genuine insulin, by the same criteria used for liver insulin. The insulin concentration in cultured human cells was unaffected by depletion of insulin from the culture medium or by addition of beef insulin to the medium. The data suggest that a part, if not all, of the extrapancreatic tissue insulin is independent of plasma insulin and may be synthesized by the tissues themselves. PMID:6987656

  8. Rat and human membrane dipeptidase: tissue distribution and developmental changes.

    PubMed

    Kera, Y; Liu, Z; Matsumoto, T; Sorimachi, Y; Nagasaki, H; Yamada, R H

    1999-05-01

    Distribution and developmental changes in membrane dipeptidase activity were examined in rat and human tissues. The activity to hydrolyze glycyl-D-alanine in rat and human tissues was completely or almost completely inhibited by 5 mM cilastatin, suggesting that the activity was due to membrane dipeptidase and that the contribution of leucine aminopeptidase to the activity was minor. In 8-week-old rats, the activity was high in lung, kidney, pancreas and testis, and in each pooled sample of ileal mucosa, duodenal mucosa, jejunal mucosa and adrenal mucosa. A low activity was found in spleen, liver, serum and heart. The activity in lung, kidney, adrenal and intestinal mucosa increased up to the age of 5 or 8 weeks, while that in pancreas, testis and spleen reached a maximal level at around 3 weeks and declined thereafter. The distribution profile of the enzyme in postmortem tissues of adult humans was similar to that in rat, except for an extremely low activity in lung. The enzyme was also found in serum and urine from healthy volunteers. In urine, the activity was significantly correlated to the creatinine content. No clear dependence of the activity on gender or age was observed in urine and serum.

  9. Human mediastinal adipose tissue displays certain characteristics of brown fat

    PubMed Central

    Cheung, L; Gertow, J; Werngren, O; Folkersen, L; Petrovic, N; Nedergaard, J; Franco-Cereceda, A; Eriksson, P; Fisher, R M

    2013-01-01

    Background: The amount of intra-thoracic fat, of which mediastinal adipose tissue comprises the major depot, is related to various cardiometabolic risk factors. Autopsy and imaging studies indicate that the mediastinal depot in adult humans could contain brown adipose tissue (BAT). To gain a better understanding of this intra-thoracic fat depot, we examined possible BAT characteristics of human mediastinal in comparison with subcutaneous adipose tissue. Materials and methods: Adipose tissue biopsies from thoracic subcutaneous and mediastinal depots were obtained during open-heart surgery from 33 subjects (26 male, 63.7±13.8 years, body mass index 29.3±5.1 kg m−2). Microarray analysis was performed on 10 patients and genes of interest confirmed by quantitative PCR (qPCR) in samples from another group of 23 patients. Adipocyte size was determined and uncoupling protein 1 (UCP1) protein expression investigated with immunohistochemistry. Results: The microarray data showed that a number of BAT-specific genes had significantly higher expression in the mediastinal depot than in the subcutaneous depot. Higher expression of UCP1 (24-fold, P<0.001) and PPARGC1A (1.7-fold, P=0.0047), and lower expression of SHOX2 (0.12-fold, P<0.001) and HOXC8 (0.14-fold, P<0.001) in the mediastinal depot was confirmed by qPCR. Gene set enrichment analysis identified two gene sets related to mitochondria, which were significantly more highly expressed in the mediastinal than in the subcutaneous depot (P<0.01). No significant changes in UCP1 gene expression were observed in the subcutaneous or mediastinal depots following lowering of body temperature during surgery. UCP1 messenger RNA levels in the mediastinal depot were lower than those in murine BAT and white adipose tissue. In some mediastinal adipose tissue biopsies, a small number of multilocular adipocytes that stained positively for UCP1 were observed. Adipocytes were significantly smaller in the mediastinal than the

  10. Elastic, permeability and swelling properties of human intervertebral disc tissues: A benchmark for tissue engineering.

    PubMed

    Cortes, Daniel H; Jacobs, Nathan T; DeLucca, John F; Elliott, Dawn M

    2014-06-27

    The aim of functional tissue engineering is to repair and replace tissues that have a biomechanical function, i.e., connective orthopaedic tissues. To do this, it is necessary to have accurate benchmarks for the elastic, permeability, and swelling (i.e., biphasic-swelling) properties of native tissues. However, in the case of the intervertebral disc, the biphasic-swelling properties of individual tissues reported in the literature exhibit great variation and even span several orders of magnitude. This variation is probably caused by differences in the testing protocols and the constitutive models used to analyze the data. Therefore, the objective of this study was to measure the human lumbar disc annulus fibrosus (AF), nucleus pulposus (NP), and cartilaginous endplates (CEP) biphasic-swelling properties using a consistent experimental protocol and analyses. The testing protocol was composed of a swelling period followed by multiple confined compression ramps. To analyze the confined compression data, the tissues were modeled using a biphasic-swelling model, which augments the standard biphasic model through the addition of a deformation-dependent osmotic pressure term. This model allows considering the swelling deformations and the contribution of osmotic pressure in the analysis of the experimental data. The swelling stretch was not different between the disc regions (AF: 1.28±0.16; NP: 1.73±0.74; CEP: 1.29±0.26), with a total average of 1.42. The aggregate modulus (Ha) of the extra-fibrillar matrix was higher in the CEP (390kPa) compared to the NP (100kPa) or AF (30kPa). The permeability was very different across tissue regions, with the AF permeability (64 E(-16)m(4)/Ns) higher than the NP and CEP (~5.5 E(-16)m(4)/Ns). Additionally, a normalized time-constant (3000s) for the stress relaxation was similar for all the disc tissues. The properties measured in this study are important as benchmarks for tissue engineering and for modeling the disc's mechanical

  11. The kallikrein-kinin system in diabetic nephropathy

    PubMed Central

    Tomita, Hirofumi; Sanford, Ryan B.; Smithies, Oliver; Kakoki, Masao

    2012-01-01

    Diabetic nephropathy is the major cause of end-stage renal disease worldwide. Although the renin-angiotensin system has been implicated in the pathogenesis of diabetic nephropathy, angiotensin I-converting enzyme (ACE) inhibitors have a beneficial effect on diabetic nephropathy independently of their effects on blood pressure and plasma angiotensin II levels. This suggests that the kallikrein-kinin system (KKS) is also involved in the disease. To study the role of the KKS in diabetic nephropathy, mice lacking either the bradykinin B1 receptor (B1R) or the bradykinin B2 receptor (B2R) have been commonly used. However, because absence of either receptor causes enhanced expression of the other, it is difficult to determine the precise functions of each receptor. This difficulty has recently been overcome by comparing mice lacking both receptors with mice lacking each receptor. Deletion of both B1R and B2R reduces nitric oxide (NO) production and aggravates renal diabetic phenotypes, relevant to either lack of B1R or B2R, demonstrating that both B1R and B2R exert protective effects on diabetic nephropathy presumably via NO. Here, we review previous epidemiological and experimental studies, and discuss novel insights regarding the therapeutic implications of the importance of the KKS in averting diabetic nephropathy. PMID:22318421

  12. Kinins— The Kallikrein-Kinin System and Oxidative Stress

    PubMed Central

    Kayashima, Yukako; Smithies, Oliver; Kakoki, Masao

    2012-01-01

    Purpose of review The Kallikrein-kinin system (KKS) constitutes a complex multi-enzyme cascade that produces several bioactive kinin peptides and their derivatives including bradykinin. In addition to the classical notion of the KKS as a potent vasodilator and a mediator of inflammatory responses, recent studies suggest a link between the KKS and oxidative stress. A number of established mouse model with altered levels of KKS components opened the way to evaluate precise functions of the KKS. Here we review recent findings on the role of the KKS in cardiovascular diseases and chronic kidney diseases, and discuss potential benefits of KKS activation in these diseases. Recent findings Deletion of both B1R and B2R in a diabetic mouse model exacerbates its renal phenotypes, suggesting that the KKS exerts protective effects on diabetic nephropathy by suppressing oxidative stress, presumably via nitric oxide (NO) and prostaglandins (PGs). Summary Accumulating evidence has highlighted the importance of the KKS as a protective system against oxidative stress and organ damage in the heart and kidney. The activation of the KKS by ACE inhibitors and vasopeptidase inhibitors is likely to be beneficial in senescence-associated cardiovascular diseases and chronic kidney diseases. PMID:22048723

  13. Maturing human pluripotent stem cell-derived cardiomyocytes in human engineered cardiac tissues.

    PubMed

    Feric, Nicole T; Radisic, Milica

    2016-01-15

    Engineering functional human cardiac tissue that mimics the native adult morphological and functional phenotype has been a long held objective. In the last 5 years, the field of cardiac tissue engineering has transitioned from cardiac tissues derived from various animal species to the production of the first generation of human engineered cardiac tissues (hECTs), due to recent advances in human stem cell biology. Despite this progress, the hECTs generated to date remain immature relative to the native adult myocardium. In this review, we focus on the maturation challenge in the context of hECTs, the present state of the art, and future perspectives in terms of regenerative medicine, drug discovery, preclinical safety testing and pathophysiological studies.

  14. Effect of prepuberal gonadectomy upon aldosterone levels in female and male SHR: interaction between blood pressure and kallikrein kinin system.

    PubMed

    Oddo, Elisabet M; de Luca Sarobe, Verónica; Krmar, Rafael; Periz, Gabriela A; Herrera, Horacio; Martín, Rodolfo S; Ibarra, Fernando R; Arrizurieta, Elvira E

    2006-02-01

    It has been suggested that an abnormal activity of the hypothalamic-pituitary-adrenal-gonadal axis may be implicated in the pathogenesis of spontaneously hypertensive rats (SHR) blood pressure hypertension. However, it is widely known that the kallikrein-kinin system plays a role in blood pressure regulation in this strain, because an inverse relation between blood pressure and urinary kallikrein excretion has been reported. It was of our interest to study how early suppression of sexual hormones affected blood pressure regulation in SHR and urinary kallikrein excretion and to elucidate the involved mechanisms. For these purpose, SH and Wistar-Kyoto (WKY) rats blood pressure, renal function, and hormonal profile were studied after prepuberal gonadectomy starting at 4 weeks of age throughout until the 12th week of age. Results were compared with those of untreated SH and WKY rats of either sex. The response to blocking agents against aldosterone and kallikrein-kinin system also were evaluated. Systolic blood pressure increased progressively in male and female SHR 12 weeks of age. Systolic blood pressure was higher in male than in female SHR, but urinary kallikrein was lower in male SHR. Prepuberal gonadectomy induced a significant decrease in systolic blood pressure in male and in female SHR at 12 weeks of age, accompanied by an increase in urinary kallikrein in male and in female SHR. Plasma aldosterone increased markedly in female and male SHR after gonadectomy. No concurrent changes in plasma renin activity or corticosterone levels were observed. The aldosterone receptor antagonist and the kallikrein inhibitor treatment blunted the blood pressure lowering effect of gonadectomy and diminished urinary kallikrein excretion. Results support the existence of a sexual dimorphism related to hypertension and urinary kallikrein and suggest an interaction among the kallikrein-kinin system, sexual hormones, and mineralocorticoids in the neonatal programming of hypertension.

  15. 3D Extracellular Matrix from Sectioned Human Tissues

    PubMed Central

    Campbell, Catherine B; Cukierman, Edna; Artym, Vira V

    2014-01-01

    corneal endothelial cell lines produce an ECM mimicking an in vivo subendothelium, and the EHS tumor cell line produces a matrix that can be extracted to produce Matrigel, which simulates basement membrane molecular complexity including laminin, collagen IV and nidogen (Beacham, et al., 2007; Friedl and Brocker, 2000). To simulate a physiological environment even more closely, 3D matrices derived from mouse tissue slices from which cells were extracted have reportedly provided successful ECM replicas for studying in vivo cellular behavior (Cukierman, et al., 2001). Because of the important roles of the extracellular microenvironment on normal and tumor cells, we have developed protocols to produce cell-free (decellularized) 3D matrices from cryostat sections of normal and tumor human tissues. These extracted matrices can be used as a 3D tissue culture environment to analyze effects of various 3D matrices on normal and tumor cell responses and behavior. Using human pancreas and breast tissue samples, we have successfully prepared cell-free 3D ECM models, used them as cell culture substrates for a human breast cancer cell line, MDA-MB-231, and then performed immunofluorescence staining to characterize intracellular structures. A frequently observed difference between normal and tumor tissue-derived ECM environments involves the amount of deposited fibrillar collagen (Provenzano, 2008). Tumor tissues from both breast and pancreas often contain substantially more collagen than normal adjacent tissue, and this protocol preserves this difference in cell-free 3D matrices from these tissues (Vidi, et al., 2013). This 3D culture system we describe using cell-free 3D matrix provides an approach to studying cellular behavior and migratory mechanisms associated with cancer. The basic protocol describes methods for successfully extracting cells and cellular debris from human tissue cryostat sections to obtain a clean, cell-free 3D ECM for plating cell lines (Figure 1). Cellular

  16. Comparative Analysis of Human Tissue Interactomes Reveals Factors Leading to Tissue-Specific Manifestation of Hereditary Diseases

    PubMed Central

    Barshir, Ruth; Shwartz, Omer; Smoly, Ilan Y.; Yeger-Lotem, Esti

    2014-01-01

    An open question in human genetics is what underlies the tissue-specific manifestation of hereditary diseases, which are caused by genomic aberrations that are present in cells across the human body. Here we analyzed this phenomenon for over 300 hereditary diseases by using comparative network analysis. We created an extensive resource of protein expression and interactions in 16 main human tissues, by integrating recent data of gene and protein expression across tissues with data of protein-protein interactions (PPIs). The resulting tissue interaction networks (interactomes) shared a large fraction of their proteins and PPIs, and only a small fraction of them were tissue-specific. Applying this resource to hereditary diseases, we first show that most of the disease-causing genes are widely expressed across tissues, yet, enigmatically, cause disease phenotypes in few tissues only. Upon testing for factors that could lead to tissue-specific vulnerability, we find that disease-causing genes tend to have elevated transcript levels and increased number of tissue-specific PPIs in their disease tissues compared to unaffected tissues. We demonstrate through several examples that these tissue-specific PPIs can highlight disease mechanisms, and thus, owing to their small number, provide a powerful filter for interrogating disease etiologies. As two thirds of the hereditary diseases are associated with these factors, comparative tissue analysis offers a meaningful and efficient framework for enhancing the understanding of the molecular basis of hereditary diseases. PMID:24921629

  17. Comparative analysis of human tissue interactomes reveals factors leading to tissue-specific manifestation of hereditary diseases.

    PubMed

    Barshir, Ruth; Shwartz, Omer; Smoly, Ilan Y; Yeger-Lotem, Esti

    2014-06-01

    An open question in human genetics is what underlies the tissue-specific manifestation of hereditary diseases, which are caused by genomic aberrations that are present in cells across the human body. Here we analyzed this phenomenon for over 300 hereditary diseases by using comparative network analysis. We created an extensive resource of protein expression and interactions in 16 main human tissues, by integrating recent data of gene and protein expression across tissues with data of protein-protein interactions (PPIs). The resulting tissue interaction networks (interactomes) shared a large fraction of their proteins and PPIs, and only a small fraction of them were tissue-specific. Applying this resource to hereditary diseases, we first show that most of the disease-causing genes are widely expressed across tissues, yet, enigmatically, cause disease phenotypes in few tissues only. Upon testing for factors that could lead to tissue-specific vulnerability, we find that disease-causing genes tend to have elevated transcript levels and increased number of tissue-specific PPIs in their disease tissues compared to unaffected tissues. We demonstrate through several examples that these tissue-specific PPIs can highlight disease mechanisms, and thus, owing to their small number, provide a powerful filter for interrogating disease etiologies. As two thirds of the hereditary diseases are associated with these factors, comparative tissue analysis offers a meaningful and efficient framework for enhancing the understanding of the molecular basis of hereditary diseases. PMID:24921629

  18. RNA Extraction from Animal and Human's Cancerous Tissues: Does Tissue Matter?

    PubMed Central

    Samadani, Ali Akbar; Nikbakhsh, Novin; Fattahi, Sadegh; Pourbagher, Roghayeh; Aghajanpour Mir, Seyyed Mohsen; Mousavi Kani, Narges; Abedian, Zeinab; Akhavan-Niaki, Haleh

    2015-01-01

    The reliability of gene expression profiling, based technologies and methods to find transcriptional differences representative of the original samples is influenced by the quality of the extracted RNA. Hence, RNA extraction is the first step to investigate the gene expression and its function. Consequently, the quality of extracted RNA is really significant. Correspondingly, this research was accomplished to optimize the RNA extraction methods and compare the amounts of tissue or quality of tissue. Relatively, the cancerous tissue of human stomach in fresh and frozen conditions and also the mouse fresh tissue were studied. Some factors like the amount of samples, efficacy differences of diverse extraction buffers (TriPure, Trizol) and also the efficacy of b-mercaptoethanol were compared and investigated. The results indicated that the less amount (1-2 mg) compared to other amounts (2-5 mg, 5-15 mg) yielded the best quality and the RNA bands (5S, 18S, 28S) were observed perfectly. Relatively, comparing and measuring some kinds of buffers (Trizol, TriPure) indicated no difference in RNA extraction quality. The last investigated factor was the effect of b- mercaptoethanol which was used along with TriPure to remove the RNAse. Conclusively, no effective impression was observed. PMID:25815283

  19. Evidence for two types of brown adipose tissue in humans.

    PubMed

    Lidell, Martin E; Betz, Matthias J; Dahlqvist Leinhard, Olof; Heglind, Mikael; Elander, Louise; Slawik, Marc; Mussack, Thomas; Nilsson, Daniel; Romu, Thobias; Nuutila, Pirjo; Virtanen, Kirsi A; Beuschlein, Felix; Persson, Anders; Borga, Magnus; Enerbäck, Sven

    2013-05-01

    The previously observed supraclavicular depot of brown adipose tissue (BAT) in adult humans was commonly believed to be the equivalent of the interscapular thermogenic organ of small mammals. This view was recently disputed on the basis of the demonstration that this depot consists of beige (also called brite) brown adipocytes, a newly identified type of brown adipocyte that is distinct from the classical brown adipocytes that make up the interscapular thermogenic organs of other mammals. A combination of high-resolution imaging techniques and histological and biochemical analyses showed evidence for an anatomically distinguishable interscapular BAT (iBAT) depot in human infants that consists of classical brown adipocytes, a cell type that has so far not been shown to exist in humans. On the basis of these findings, we conclude that infants, similarly to rodents, have the bona fide iBAT thermogenic organ consisting of classical brown adipocytes that is essential for the survival of small mammals in a cold environment.

  20. Multipotent progenitor cells isolated from adult human pancreatic tissue.

    PubMed

    Todorov, I; Nair, I; Ferreri, K; Rawson, J; Kuroda, A; Pascual, M; Omori, K; Valiente, L; Orr, C; Al-Abdullah, I; Riggs, A; Kandeel, F; Mullen, Y

    2005-10-01

    The supply of islet cells is a limiting factor for the widespread application of islet transplantation of type-1 diabetes. Islets constitute 1% to 2% of pancreatic tissue, leaving approximately 98% as discard after islet isolation and purification. In this report we present our data on the isolation of multipotent progenitor cells from discarded adult human pancreatic tissue. The collected cells from discarded nonislet fractions, after enzymatic digestion and gradient purification of islets, were dissociated for suspension culture in a serum-free medium. The cell clusters grown to a size of 100 to 150 mum contained cells staining for stage-specific embryonic antigens, but not insulin or C-peptide. To direct cell differentiation toward islets, clusters were recultured in a pancreatic differentiation medium. Insulin and C-peptide-positive cells by immunocytochemistry appeared within a week, reaching over 10% of the cell population. Glucagon and somatostatin-positive cells were also detected. The cell clusters were found to secrete insulin in response to glucose stimulation. Cells from the same clusters also had the capacity for differentiation into neural cells, as documented by staining for neural and glial cell markers when cultured as monolayers in media containing neurotrophic factors. These data suggest that multipotent pancreatic progenitor cells exist within the human pancreatic tissue that is typically discarded during islet isolation procedures. These adult progenitor cells can be successfully differentiated into insulin-producing cells, and thus they have the potential for treatment of type-1 diabetes mellitus. PMID:16298614

  1. Dynamic biaxial tissue properties of the human cadaver aorta.

    PubMed

    Shah, Chirag S; Hardy, Warren N; Mason, Matthew J; Yang, King H; Van Ee, Chris A; Morgan, Richard; Digges, Kennerly

    2006-11-01

    This study focuses on the biaxial mechanical properties of planar aorta tissue at strain rates likely to be experienced during automotive crashes. It also examines the structural response of the whole aorta to longitudinal tension. Twenty-six tissue-level tests were conducted using twelve thoracic aortas harvested from human cadavers. Cruciate samples were excised from the ascending, peri-isthmic, and descending regions. The samples were subjected to equibiaxial stretch at two nominal speed levels using a new biaxial tissue-testing device. Inertia-compensated loads were measured to facilitate calculation of true stress. High-speed videography and regional correlation analysis were used to track ink dots marked on the center of each sample to obtain strain. In a series of component-level tests, the response of the intact thoracic aorta to longitudinal stretch was obtained using seven aorta specimens. The aorta fails within the peri-isthmic region. The aorta fails in the transverse direction, and the intima fails before the media or adventitia. The aorta tissue exhibits nonlinear behavior. The aorta as complete structure can transect completely from 92 N axial load and 0.221 axial strain. Complete transection can be accompanied by intimal tears. These results have application to finite element modeling and the better understanding of traumatic rupture of the aorta. PMID:17311166

  2. An immunohistochemical study of Na+/I- symporter in human thyroid tissues and salivary gland tissues.

    PubMed

    Jhiang, S M; Cho, J Y; Ryu, K Y; DeYoung, B R; Smanik, P A; McGaughy, V R; Fischer, A H; Mazzaferri, E L

    1998-10-01

    The human Na+/I- symporter (hNIS) is the plasma membrane protein that mediates active iodide uptake into several tissues, such as the thyroid and salivary glands. To study the distribution and cellular localization of the hNIS protein, we have generated a polyclonal antibody that could detect the hNIS protein by immunohistochemical staining on tissue sections. In normal thyroids, hNIS expression is heterogeneous, and it is only detected in sporadic thyrocytes of a given follicle. The hNIS protein was not detected in thyroid carcinomas, yet it was detected in the majority of thyrocytes in Graves' thyroids. In salivary glands, hNIS protein was not detected in acinar cells, but it was detected in ductal cells. The hNIS proteins are clustered in the basal and lateral membranes in cells stained positive for hNIS.

  3. Computational model of soft tissues in the human upper airway.

    PubMed

    Pelteret, J-P V; Reddy, B D

    2012-01-01

    This paper presents a three-dimensional finite element model of the tongue and surrounding soft tissues with potential application to the study of sleep apnoea and of linguistics and speech therapy. The anatomical data was obtained from the Visible Human Project, and the underlying histological data was also extracted and incorporated into the model. Hyperelastic constitutive models were used to describe the material behaviour, and material incompressibility was accounted for. An active Hill three-element muscle model was used to represent the muscular tissue of the tongue. The neural stimulus for each muscle group was determined through the use of a genetic algorithm-based neural control model. The fundamental behaviour of the tongue under gravitational and breathing-induced loading is investigated. It is demonstrated that, when a time-dependent loading is applied to the tongue, the neural model is able to control the position of the tongue and produce a physiologically realistic response for the genioglossus. PMID:25830209

  4. Computational model of soft tissues in the human upper airway.

    PubMed

    Pelteret, J-P V; Reddy, B D

    2012-01-01

    This paper presents a three-dimensional finite element model of the tongue and surrounding soft tissues with potential application to the study of sleep apnoea and of linguistics and speech therapy. The anatomical data was obtained from the Visible Human Project, and the underlying histological data was also extracted and incorporated into the model. Hyperelastic constitutive models were used to describe the material behaviour, and material incompressibility was accounted for. An active Hill three-element muscle model was used to represent the muscular tissue of the tongue. The neural stimulus for each muscle group was determined through the use of a genetic algorithm-based neural control model. The fundamental behaviour of the tongue under gravitational and breathing-induced loading is investigated. It is demonstrated that, when a time-dependent loading is applied to the tongue, the neural model is able to control the position of the tongue and produce a physiologically realistic response for the genioglossus.

  5. 75 FR 34146 - Proposed Collection; Comment Request Resource for the Collection and Evaluation of Human Tissues...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-16

    ... Collection and Evaluation of Human Tissues and Cells From Donors With an Epidemiology Profile (NCI) SUMMARY... Collection: Title: Resource for the Collection and Evaluation of Human Tissues and Cells From Donors With...

  6. Diversity of lipid mediators in human adipose tissue depots

    PubMed Central

    Clària, Joan; Nguyen, Binh T.; Madenci, Arin L.; Ozaki, C. Keith

    2013-01-01

    Adipose tissue is a heterogeneous organ with remarkable variations in fat cell metabolism depending on the anatomical location. However, the pattern and distribution of bioactive lipid mediators between different fat depots and their relationships in complex diseases have not been investigated. Using LC-MS/MS-based metabolo-lipidomics, here we report that human subcutaneous (SC) adipose tissues possess a range of specialized proresolving mediators (SPM) including resolvin (Rv) D1, RvD2, protectin (PD) 1, lipoxin (LX) A4, and the monohydroxy biosynthetic pathway markers of RvD1 and PD1 (17-HDHA), RvE1 (18-HEPE), and maresin 1 (14-HDHA). The “classic” eicosanoids prostaglandin (PG) E2, PGD2, PGF2α, leukotriene (LT) B4, 5-hydroxyeicosatetraenoic acid (5-HETE), 12-HETE, and 15-HETE were also identified in SC fat. SC fat from patients with peripheral vascular disease (PVD) exhibited a marked deficit in PD1 and 17-HDHA levels. Compared with SC, perivascular adipose tissue displayed higher SPM levels, suggesting an enhanced resolution capacity in this fat depot. In addition, augmented levels of eicosanoids and SPM were observed in SC fat surrounding foot wounds. Notably, the profile of SC PGF2α differed significantly when patients were grouped by body mass index (BMI). In the case of peri-wound SC fat, BMI negatively correlated with PGE2. In this tissue, proresolving mediators RvD2 and LXA4 were identified in lower levels than the proinflammatory LTB4. Collectively, these findings demonstrate a diverse distribution of bioactive lipid mediators depending on the localization of human fat depots and uncover a specific SPM pattern closely associated with PVD. PMID:23364264

  7. Magnetic Resonance Imaging of Human Tissue-Engineered Adipose Substitutes.

    PubMed

    Proulx, Maryse; Aubin, Kim; Lagueux, Jean; Audet, Pierre; Auger, Michèle; Fortin, Marc-André; Fradette, Julie

    2015-07-01

    Adipose tissue (AT) substitutes are being developed to answer the strong demand in reconstructive surgery. To facilitate the validation of their functional performance in vivo, and to avoid resorting to excessive number of animals, it is crucial at this stage to develop biomedical imaging methodologies, enabling the follow-up of reconstructed AT substitutes. Until now, biomedical imaging of AT substitutes has scarcely been reported in the literature. Therefore, the optimal parameters enabling good resolution, appropriate contrast, and graft delineation, as well as blood perfusion validation, must be studied and reported. In this study, human adipose substitutes produced from adipose-derived stem/stromal cells using the self-assembly approach of tissue engineering were implanted into athymic mice. The fate of the reconstructed AT substitutes implanted in vivo was successfully followed by magnetic resonance imaging (MRI), which is the imaging modality of choice for visualizing soft ATs. T1-weighted images allowed clear delineation of the grafts, followed by volume integration. The magnetic resonance (MR) signal of reconstructed AT was studied in vitro by proton nuclear magnetic resonance ((1)H-NMR). This confirmed the presence of a strong triglyceride peak of short longitudinal proton relaxation time (T1) values (200 ± 53 ms) in reconstructed AT substitutes (total T1=813 ± 76 ms), which establishes a clear signal difference between adjacent muscle, connective tissue, and native fat (total T1 ~300 ms). Graft volume retention was followed up to 6 weeks after implantation, revealing a gradual resorption rate averaging at 44% of initial substitute's volume. In addition, vascular perfusion measured by dynamic contrast-enhanced-MRI confirmed the graft's vascularization postimplantation (14 and 21 days after grafting). Histological analysis of the grafted tissues revealed the persistence of numerous adipocytes without evidence of cysts or tissue necrosis. This study

  8. Nonproteolytic neuroprotection by human recombinant tissue plasminogen activator.

    PubMed

    Kim, Y H; Park, J H; Hong, S H; Koh, J Y

    1999-04-23

    Human recombinant tissue plasminogen activator (tPA) may benefit ischemic stroke patients by dissolving clots. However, independent of thrombolysis, tPA may also have deleterious effects on neurons by promoting excitotoxicity. Zinc neurotoxicity has been shown to be an additional key mechanism in brain injuries. Hence, if tPA affects zinc neurotoxicity, this may provide additional insights into its effect on neuronal death. Independent of its proteolytic action, tPA markedly attenuated zinc-induced cell death in cortical culture, and, when injected into cerebrospinal fluid, also reduced kainate seizure-induced hippocampal neuronal death in adult rats.

  9. 21 CFR 1270.43 - Retention, recall, and destruction of human tissue.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Retention, recall, and destruction of human tissue. 1270.43 Section 1270.43 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN... HUMAN TISSUE INTENDED FOR TRANSPLANTATION Inspection of Tissue Establishments § 1270.43...

  10. 21 CFR 1270.43 - Retention, recall, and destruction of human tissue.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Retention, recall, and destruction of human tissue. 1270.43 Section 1270.43 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN... HUMAN TISSUE INTENDED FOR TRANSPLANTATION Inspection of Tissue Establishments § 1270.43...

  11. 21 CFR 1270.43 - Retention, recall, and destruction of human tissue.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Retention, recall, and destruction of human tissue. 1270.43 Section 1270.43 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN... HUMAN TISSUE INTENDED FOR TRANSPLANTATION Inspection of Tissue Establishments § 1270.43...

  12. 21 CFR 1270.43 - Retention, recall, and destruction of human tissue.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Retention, recall, and destruction of human tissue. 1270.43 Section 1270.43 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN... HUMAN TISSUE INTENDED FOR TRANSPLANTATION Inspection of Tissue Establishments § 1270.43...

  13. 21 CFR 1270.43 - Retention, recall, and destruction of human tissue.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Retention, recall, and destruction of human tissue. 1270.43 Section 1270.43 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN... HUMAN TISSUE INTENDED FOR TRANSPLANTATION Inspection of Tissue Establishments § 1270.43...

  14. Identification of Tissue-Specific Protein-Coding and Noncoding Transcripts across 14 Human Tissues Using RNA-seq

    PubMed Central

    Zhu, Jinhang; Chen, Geng; Zhu, Sibo; Li, Suqing; Wen, Zhuo; Bin Li; Zheng, Yuanting; Shi, Leming

    2016-01-01

    Many diseases and adverse drug reactions exhibit tissue specificity. To better understand the tissue-specific expression characteristics of transcripts in different human tissues, we deeply sequenced RNA samples from 14 different human tissues. After filtering many lowly expressed transcripts, 24,729 protein-coding transcripts and 1,653 noncoding transcripts were identified. By analyzing highly expressed tissue-specific protein-coding transcripts (TSCTs) and noncoding transcripts (TSNTs), we found that testis expressed the highest numbers of TSCTs and TSNTs. Brain, monocytes, ovary, and heart expressed more TSCTs than the rest tissues, whereas brain, placenta, heart, and monocytes expressed more TSNTs than other tissues. Co-expression network constructed based on the TSCTs and TSNTs showed that each hub TSNT was co-expressed with several TSCTs, allowing functional annotation of TSNTs. Important biological processes and KEGG pathways highly related to the specific functions or diseases of each tissue were enriched with the corresponding TSCTs. These TSCTs and TSNTs may participate in the tissue-specific physiological or pathological processes. Our study provided a unique data set and systematic analysis of expression characteristics and functions of both TSCTs and TSNTs based on 14 distinct human tissues, and could facilitate future investigation of the mechanisms behind tissue-specific diseases and adverse drug reactions. PMID:27329541

  15. Identification of Tissue-Specific Protein-Coding and Noncoding Transcripts across 14 Human Tissues Using RNA-seq.

    PubMed

    Zhu, Jinhang; Chen, Geng; Zhu, Sibo; Li, Suqing; Wen, Zhuo; Bin Li; Zheng, Yuanting; Shi, Leming

    2016-06-22

    Many diseases and adverse drug reactions exhibit tissue specificity. To better understand the tissue-specific expression characteristics of transcripts in different human tissues, we deeply sequenced RNA samples from 14 different human tissues. After filtering many lowly expressed transcripts, 24,729 protein-coding transcripts and 1,653 noncoding transcripts were identified. By analyzing highly expressed tissue-specific protein-coding transcripts (TSCTs) and noncoding transcripts (TSNTs), we found that testis expressed the highest numbers of TSCTs and TSNTs. Brain, monocytes, ovary, and heart expressed more TSCTs than the rest tissues, whereas brain, placenta, heart, and monocytes expressed more TSNTs than other tissues. Co-expression network constructed based on the TSCTs and TSNTs showed that each hub TSNT was co-expressed with several TSCTs, allowing functional annotation of TSNTs. Important biological processes and KEGG pathways highly related to the specific functions or diseases of each tissue were enriched with the corresponding TSCTs. These TSCTs and TSNTs may participate in the tissue-specific physiological or pathological processes. Our study provided a unique data set and systematic analysis of expression characteristics and functions of both TSCTs and TSNTs based on 14 distinct human tissues, and could facilitate future investigation of the mechanisms behind tissue-specific diseases and adverse drug reactions.

  16. Characterization of kallikrein-related peptidase 4 glycosylations

    PubMed Central

    Yamakoshi, Yasuo; Yamakoshi, Fumiko; Hu, Jan C-C.; Simmer, James P.

    2012-01-01

    Kallikrein-related peptidase 4 (KLK4) is a glycosylated serine protease that functions in the maturation (hardening) of dental enamel. Pig and mouse KLK4 contain three potential N-glycosylation sites. We isolated KLK4 from developing pig and mouse molars and characterized their N-glycosylations. N-glycans were enzymatically released by digestion with N-glycosidase F and fluorescently labeled with 2-aminobenzoic acid. Normal-phase high-performance liquid chromatography (NP-HPLC) revealed N-glycans with no, or with one, two, or three sialic acid attachments in pig KLK4 and with no, or with one or two sialic acid attachments in mouse KLK4. The labeled N-glycans were digested with sialidase to generate the asialo N-glycan cores that were fractionated by reverse-phase HPLC, and their retention times were compared with similarly labeled glycan standards. The purified cores were characterized by mass spectrometric and monosaccharide composition analyses. We determined that pig and mouse KLK4 have NA2 and NA2F biantennary N-glycan cores. The pig triantennary core is NA3. The mouse triantennary core is NA3 with a fucose connected by an α1–6 linkage, indicating that it is attached to the first N-acetyglucosamine (NA3F). We conclude that pig KLK4 has NA2, NA2F, and NA3 N-glycan cores with no, or with one, two, or three sialic acids. Mouse KLK4 has NA2, NA2F, and NA3F N-glycan cores with no, or with one or two sialic acids. PMID:22243251

  17. Characterization of kallikrein-related peptidase 4 glycosylations.

    PubMed

    Yamakoshi, Yasuo; Yamakoshi, Fumiko; Hu, Jan C-C; Simmer, James P

    2011-12-01

    Kallikrein-related peptidase 4 (KLK4) is a glycosylated serine protease that functions in the maturation (hardening) of dental enamel. Pig and mouse KLK4 contain three potential N-glycosylation sites. We isolated KLK4 from developing pig and mouse molars and characterized their N-glycosylations. N-glycans were enzymatically released by digestion with N-glycosidase F and fluorescently labeled with 2-aminobenzoic acid. Normal-phase high-performance liquid chromatography (NP-HPLC) revealed N-glycans with no, or with one, two, or three sialic acid attachments in pig KLK4 and with no, or with one or two sialic acid attachments in mouse KLK4. The labeled N-glycans were digested with sialidase to generate the asialo N-glycan cores that were fractionated by reverse-phase HPLC, and their retention times were compared with similarly labeled glycan standards. The purified cores were characterized by mass spectrometric and monosaccharide composition analyses. We determined that pig and mouse KLK4 have NA2 and NA2F biantennary N-glycan cores. The pig triantennary core is NA3. The mouse triantennary core is NA3 with a fucose connected by an α1-6 linkage, indicating that it is attached to the first N-acetyglucosamine (NA3F). We conclude that pig KLK4 has NA2, NA2F, and NA3 N-glycan cores with no, or with one, two, or three sialic acids. Mouse KLK4 has NA2, NA2F, and NA3F N-glycan cores with no, or with one or two sialic acids.

  18. Marketing of human organs and tissues is justified and necessary.

    PubMed

    Kevorkian, J

    1989-01-01

    The bioethical guidelines now banning commerce in human body parts to be used for transplantation manifest unrealistic and arbitrary inflexibility which perpetuates and worsens the deficit in organ supply. Instead of relying on traditionally revered but now outmoded and even irrelevant bioethical maxims, formulators of the guidelines should have concentrated on a more meaningful situational adaptation to contemporary real-life circumstances. Many unexpectedly relevant and important nuances of concepts such as property, ownership, and altruism must now be taken into account. Hypothetical examples explore the morality of a universal ban by fiat and the associated problems of organ supply and demand, of cost and affordability, and of fair equity. It is difficult to justify purely altruistic organ donation today, when the health care professions and industries are frantically pursuing commercial profits. It is concluded that the ban should be scrapped in favor of a well-organized, open, and legally regulated commercial market for human organs and tissues.

  19. Fracture of human femur tissue monitored by acoustic emission sensors.

    PubMed

    Aggelis, Dimitrios G; Strantza, Maria; Louis, Olivia; Boulpaep, Frans; Polyzos, Demosthenes; van Hemelrijck, Danny

    2015-01-01

    The study describes the acoustic emission (AE) activity during human femur tissue fracture. The specimens were fractured in a bending-torsion loading pattern with concurrent monitoring by two AE sensors. The number of recorded signals correlates well with the applied load providing the onset of micro-fracture at approximately one sixth of the maximum load. Furthermore, waveform frequency content and rise time are related to the different modes of fracture (bending of femur neck or torsion of diaphysis). The importance of the study lies mainly in two disciplines. One is that, although femurs are typically subjects of surgical repair in humans, detailed monitoring of the fracture with AE will enrich the understanding of the process in ways that cannot be achieved using only the mechanical data. Additionally, from the point of view of monitoring techniques, applying sensors used for engineering materials and interpreting the obtained data pose additional difficulties due to the uniqueness of the bone structure.

  20. Fracture of Human Femur Tissue Monitored by Acoustic Emission Sensors

    PubMed Central

    Aggelis, Dimitrios. G.; Strantza, Maria; Louis, Olivia; Boulpaep, Frans; Polyzos, Demosthenes; van Hemelrijck, Danny

    2015-01-01

    The study describes the acoustic emission (AE) activity during human femur tissue fracture. The specimens were fractured in a bending-torsion loading pattern with concurrent monitoring by two AE sensors. The number of recorded signals correlates well with the applied load providing the onset of micro-fracture at approximately one sixth of the maximum load. Furthermore, waveform frequency content and rise time are related to the different modes of fracture (bending of femur neck or torsion of diaphysis). The importance of the study lies mainly in two disciplines. One is that, although femurs are typically subjects of surgical repair in humans, detailed monitoring of the fracture with AE will enrich the understanding of the process in ways that cannot be achieved using only the mechanical data. Additionally, from the point of view of monitoring techniques, applying sensors used for engineering materials and interpreting the obtained data pose additional difficulties due to the uniqueness of the bone structure. PMID:25763648

  1. Fracture of human femur tissue monitored by acoustic emission sensors.

    PubMed

    Aggelis, Dimitrios G; Strantza, Maria; Louis, Olivia; Boulpaep, Frans; Polyzos, Demosthenes; van Hemelrijck, Danny

    2015-01-01

    The study describes the acoustic emission (AE) activity during human femur tissue fracture. The specimens were fractured in a bending-torsion loading pattern with concurrent monitoring by two AE sensors. The number of recorded signals correlates well with the applied load providing the onset of micro-fracture at approximately one sixth of the maximum load. Furthermore, waveform frequency content and rise time are related to the different modes of fracture (bending of femur neck or torsion of diaphysis). The importance of the study lies mainly in two disciplines. One is that, although femurs are typically subjects of surgical repair in humans, detailed monitoring of the fracture with AE will enrich the understanding of the process in ways that cannot be achieved using only the mechanical data. Additionally, from the point of view of monitoring techniques, applying sensors used for engineering materials and interpreting the obtained data pose additional difficulties due to the uniqueness of the bone structure. PMID:25763648

  2. Rolling the human amnion to engineer laminated vascular tissues.

    PubMed

    Amensag, Salma; McFetridge, Peter S

    2012-11-01

    The prevalence of cardiovascular disease and the limited availability of suitable autologous transplant vessels for coronary and peripheral bypass surgeries is a significant clinical problem. A great deal of progress has been made over recent years to develop biodegradable materials with the potential to remodel and regenerate vascular tissues. However, the creation of functional biological scaffolds capable of withstanding vascular stress within a clinically relevant time frame has proved to be a challenging proposition. As an alternative approach, we report the use of a multilaminate rolling approach using the human amnion to generate a tubular construct for blood vessel regeneration. The human amniotic membrane was decellularized by agitation in 0.03% (w/v) sodium dodecyl sulfate to generate an immune compliant material. The adhesion of human umbilical vein endothelial cells (EC) and human vascular smooth muscle cells (SMC) was assessed to determine initial binding and biocompatibility (monocultures). Extended cultures were either assessed as flat membranes, or rolled to form concentric multilayered conduits. Results showed positive EC adhesion and a progressive repopulation by SMC. Functional changes in SMC gene expression and the constructs' bulk mechanical properties were concomitant with vessel remodeling as assessed over a 40-day culture period. A significant advantage with this approach is the ability to rapidly produce a cell-dense construct with an extracellular matrix similar in architecture and composition to natural vessels. The capacity to control physical parameters such as vessel diameter, wall thickness, shape, and length are critical to match vessel compliance and tailor vessel specifications to distinct anatomical locations. As such, this approach opens new avenues in a range of tissue regenerative applications that may have a much wider clinical impact.

  3. Human papillomavirus detection in paraffin-embedded colorectal cancer tissues.

    PubMed

    Tanzi, Elisabetta; Bianchi, Silvia; Frati, Elena R; Amicizia, Daniela; Martinelli, Marianna; Bragazzi, Nicola L; Brisigotti, Maria Pia; Colzani, Daniela; Fasoli, Ester; Zehender, Gianguglielmo; Panatto, Donatella; Gasparini, Roberto

    2015-01-01

    Human papillomavirus (HPV) has a well-recognized aetiological role in the development of cervical cancer and other anogenital tumours. Recently, an association between colorectal cancer and HPV infection has been suggested, although this is still controversial. This study aimed at detecting and characterizing HPV infection in 57 paired biopsies from colorectal cancers and adjacent intact tissues using a degenerate PCR approach. All amplified fragments were genotyped by means of sequencing. Overall, HPV prevalence was 12.3 %. In particular, 15.8 % of tumour tissues and 8.8 % of non-cancerous tissue samples were HPV DNA-positive. Of these samples, 85.7 % were genotyped successfully, with 41.7 % of sequences identifying four genotypes of the HR (high oncogenic risk) clade Group 1; the remaining 58.3 % of HPV-genotyped specimens had an unclassified β-HPV. Examining additional cases and analysing whole genomes will help to outline the significance of these findings.

  4. Mechanical stimulation improves tissue-engineered human skeletal muscle

    NASA Technical Reports Server (NTRS)

    Powell, Courtney A.; Smiley, Beth L.; Mills, John; Vandenburgh, Herman H.

    2002-01-01

    Human bioartificial muscles (HBAMs) are tissue engineered by suspending muscle cells in collagen/MATRIGEL, casting in a silicone mold containing end attachment sites, and allowing the cells to differentiate for 8 to 16 days. The resulting HBAMs are representative of skeletal muscle in that they contain parallel arrays of postmitotic myofibers; however, they differ in many other morphological characteristics. To engineer improved HBAMs, i.e., more in vivo-like, we developed Mechanical Cell Stimulator (MCS) hardware to apply in vivo-like forces directly to the engineered tissue. A sensitive force transducer attached to the HBAM measured real-time, internally generated, as well as externally applied, forces. The muscle cells generated increasing internal forces during formation which were inhibitable with a cytoskeleton depolymerizer. Repetitive stretch/relaxation for 8 days increased the HBAM elasticity two- to threefold, mean myofiber diameter 12%, and myofiber area percent 40%. This system allows engineering of improved skeletal muscle analogs as well as a nondestructive method to determine passive force and viscoelastic properties of the resulting tissue.

  5. Biomonitoring of nickel and chromium in human pulmonary tissue.

    PubMed

    Raithel, H J; Schaller, K H; Kraus, T; Lehnert, G

    1993-01-01

    Nickel (Ni) and chromium (Cr) and some of its compounds may be able to induce cancer in the lungs as well as in the nose and paranasal sinuses after occupational exposure. Latency periods amount to 20 years and more. Therefore objective exposure data are not available in the most cases and expert evaluation of the causal connection is often difficult. Recent investigations have shown, that Ni and Cr can cumulate in human lung tissue after occupational exposure. For the evaluation of "normal" Ni- and Cr-values a total of 495 human lung tissue samples of 30 occupationally non-exposed persons were analysed by AAS including ZEEMAN-compensation after wet oxidative digestion. Additional samples of 10 deceased persons who have been occupationally exposed to nickel in previous times by nickel-refining and welding, especially flame spraying have been investigated. The median Ni- and Cr- concentrations in the lungs of the non-exposed persons ranged between 20-40 resp. 133-277 ng/g (wet weight). In nickel refinery workers Ni- concentrations were found which exceeded the normal range about 1,000. In welders, especially flame sprayers, also values more than 100 times higher could be analysed for Ni and Cr. Partially these concentrations were found years after the end of the inhalative exposure. PMID:8406925

  6. The significance of using pooled human serum in human articular cartilage tissue engineering.

    PubMed

    Azmi, B; Aminuddin, B S; Sharaf, I; Samsudin, O C; Munirah, S; Chua, K H; Ruszymah, B H I

    2004-05-01

    Animal serum is commonly used in chondrocytes culture expansion to promote cell proliferation and shorten the time lag before new tissue reconstruction is possible. However, animal serum is not suitable for regeneration of clinical tissue because it has potential risk of viral and prion related disease transmission particularly mad cow disease and foreign protein contamination that can stimulate immune reaction leading to graft rejection. In this context, human serum as homologous supplement has a greater potential as growth promoting agents for human chondrocytes culture. PMID:15468795

  7. Tissue microchimerism is increased during pregnancy: a human autopsy study.

    PubMed

    Rijnink, Emilie C; Penning, Marlies E; Wolterbeek, Ron; Wilhelmus, Suzanne; Zandbergen, Malu; van Duinen, Sjoerd G; Schutte, Joke; Bruijn, Jan A; Bajema, Ingeborg M

    2015-11-01

    Microchimerism is the occurrence of small populations of cells with a different genetic background within an individual. Tissue microchimerism is considered to be primarily pregnancy-derived and is often studied relative to female-dominant autoimmune diseases, pregnancy complications, malignancies, response to injury, and transplantation outcomes. A particular distribution pattern of chimeric cells across various organs was recently described in a model of murine pregnancies. Our aim was to determine the frequency and distribution of tissue microchimerism across organs during and after pregnancy in humans. We performed in situ hybridization of the Y chromosome on paraffin-embedded autopsy samples of kidneys, livers, spleens, lungs, hearts and brains that were collected from 26 women who died while pregnant or within 1 month after delivery of a son. Frequencies of chimeric cells in various tissues were compared with those of a control group of non-pregnant women who had delivered sons. Tissue microchimerism occurred significantly more frequently in the lungs, spleens, livers, kidneys and hearts of pregnant women compared with non-pregnant women (all P < 0.01). We showed that some of the chimeric cells were CD3+ or CD34+. After correction for cell density, the lung was most chimeric (470 Y chromosome-positive nuclei per million nuclei scored), followed by the spleen (208 Y+/10(6) nuclei), liver (192 Y+/10(6) nuclei), kidney (135 Y+/10(6) nuclei), brain (85 Y+/10(6) nuclei) and heart (40 Y+/10(6) nuclei). Data from this unique study group of women who died while pregnant or shortly after delivery provide information about the number and physiologic distribution of chimeric cells in organs of pregnant women. We demonstrate that during pregnancy, a boost of chimeric cells is observed in women, with a distribution across organs, that parallels findings in mouse models. PMID:26307194

  8. Quantification of human body fat tissue percentage by MRI.

    PubMed

    Müller, Hans-Peter; Raudies, Florian; Unrath, Alexander; Neumann, Heiko; Ludolph, Albert C; Kassubek, Jan

    2011-01-01

    The MRI-based evaluation of the quantity and regional distribution of adipose tissue is one objective measure in the investigation of obesity. The aim of this article was to report a comprehensive and automatic analytical method for the determination of the volumes of subcutaneous fat tissue (SFT) and visceral fat tissue (VFT) in either the whole human body or selected slices or regions of interest. Using an MRI protocol in an examination position that was convenient for volunteers and patients with severe diseases, 22 healthy subjects were examined. The software platform was able to merge MRI scans of several body regions acquired in separate acquisitions. Through a cascade of image processing steps, SFT and VFT volumes were calculated. Whole-body SFT and VFT distributions, as well as fat distributions of defined body slices, were analysed in detail. Complete three-dimensional datasets were analysed in a reproducible manner with as few operator-dependent interventions as possible. In order to determine the SFT volume, the ARTIS (Adapted Rendering for Tissue Intensity Segmentation) algorithm was introduced. The advantage of the ARTIS algorithm was the delineation of SFT volumes in regions in which standard region grow techniques fail. Using the ARTIS algorithm, an automatic SFT volume detection was feasible. MRI data analysis was able to determine SFT and VFT volume percentages using new analytical strategies. With the techniques described, it was possible to detect changes in SFT and VFT percentages of the whole body and selected regions. The techniques presented in this study are likely to be of use in obesity-related investigations, as well as in the examination of longitudinal changes in weight during various medical conditions.

  9. Human adipose tissue expresses intrinsic circadian rhythm in insulin sensitivity.

    PubMed

    Carrasco-Benso, Maria P; Rivero-Gutierrez, Belen; Lopez-Minguez, Jesus; Anzola, Andrea; Diez-Noguera, Antoni; Madrid, Juan A; Lujan, Juan A; Martínez-Augustin, Olga; Scheer, Frank A J L; Garaulet, Marta

    2016-09-01

    In humans, insulin sensitivity varies according to time of day, with decreased values in the evening and at night. Mechanisms responsible for the diurnal variation in insulin sensitivity are unclear. We investigated whether human adipose tissue (AT) expresses intrinsic circadian rhythms in insulin sensitivity that could contribute to this phenomenon. Subcutaneous and visceral AT biopsies were obtained from extremely obese participants (body mass index, 41.8 ± 6.3 kg/m(2); 46 ± 11 y) during gastric-bypass surgery. To assess the rhythm in insulin signaling, AKT phosphorylation was determined every 4 h over 24 h in vitro in response to different insulin concentrations (0, 1, 10, and 100 nM). Data revealed that subcutaneous AT exhibited robust circadian rhythms in insulin signaling (P < 0.00001). Insulin sensitivity reached its maximum (acrophase) around noon, being 54% higher than during midnight (P = 0.009). The amplitude of the rhythm was positively correlated with in vivo sleep duration (r = 0.53; P = 0.023) and negatively correlated with in vivo bedtime (r = -0.54; P = 0.020). No circadian rhythms were detected in visceral AT (P = 0.643). Here, we demonstrate the relevance of the time of the day for how sensitive AT is to the effects of insulin. Subcutaneous AT shows an endogenous circadian rhythm in insulin sensitivity that could provide an underlying mechanism for the daily rhythm in systemic insulin sensitivity.-Carrasco-Benso, M. P., Rivero-Gutierrez, B., Lopez-Minguez, J., Anzola, A., Diez-Noguera, A., Madrid, J. A., Lujan, J. A., Martínez-Augustin, O., Scheer, F. A. J. L., Garaulet, M. Human adipose tissue expresses intrinsic circadian rhythm in insulin sensitivity.

  10. Mouse DNA contamination in human tissue tested for XMRV

    PubMed Central

    2010-01-01

    Background We used a PCR-based approach to study the prevalence of genetic sequences related to a gammaretrovirus, xenotropic murine leukemia virus-related virus, XMRV, in human prostate cancer. This virus has been identified in the US in prostate cancer patients and in those with chronic fatigue syndrome. However, with the exception of two patients in Germany, XMRV has not been identified in prostate cancer tissue in Europe. Most putative associations of new or old human retroviruses with diseases have turned out to be due to contamination. We have looked for XMRV sequences in DNA extracted from formalin-fixed paraffin- embedded prostate tissues. To control for contamination, PCR assays to detect either mouse mitochondrial DNA (mtDNA) or intracisternal A particle (IAP) long terminal repeat DNA were run on all samples, owing to their very high copy number in mouse cells. Results In general agreement with the US prevalence, XMRV-like sequences were found in 4.8% of prostate cancers. However, these were also positive, as were 21.5% of XMRV-negative cases, for IAP sequences, and many, but not all were positive for mtDNA sequences. Conclusions These results show that contamination with mouse DNA is widespread and detectable by the highly sensitive IAP assay, but not always with less sensitive assays, such as murine mtDNA PCR. This study highlights the ubiquitous presence of mouse DNA in laboratory specimens and offers a means of rigorous validation for future studies of murine retroviruses in human disease. PMID:21171966

  11. Computational analysis of tissue-specific combinatorial gene regulation: predicting interaction between transcription factors in human tissues

    PubMed Central

    Yu, Xueping; Lin, Jimmy; Zack, Donald J.; Qian, Jiang

    2006-01-01

    Tissue-specific gene expression is generally regulated by more than a single transcription factor (TF). Multiple TFs work in concert to achieve tissue specificity. In order to explore these complex TF interaction networks, we performed a large-scale analysis of TF interactions for 30 human tissues. We first identified tissue-specific genes for 30 tissues based on gene expression databases. We then evaluated the relationships between TFs using the relative position and co-occurrence of their binding sites in the promoters of tissue-specific genes. The predicted TF–TF interactions were validated by both known protein–protein interactions and co-expression of their target genes. We found that our predictions are enriched in known protein–protein interactions (>80 times that of random expectation). In addition, we found that the target genes show the highest co-expression in the tissue of interest. Our findings demonstrate that non-tissue specific TFs play a large role in regulation of tissue-specific genes. Furthermore, they show that individual TFs can contribute to tissue specificity in different tissues by interacting with distinct TF partners. Lastly, we identified several tissue-specific TF clusters that may play important roles in tissue-specific gene regulation. PMID:16982645

  12. Hemoglobin enhances tissue factor expression on human malignant cells.

    PubMed

    Siddiqui, F A; Amirkhosravi, A; Amaya, M; Meyer, T; Biggerstaff, J; Desai, H; Francis, J L

    2001-04-01

    Tissue Factor (TF) is a transmembrane glycoprotein that complexes with factor VII/activated factor VII to initiate blood coagulation. TF may be expressed on the surface of various cells including monocytes and endothelial cells. Over-expression of TF in human tumor cell lines promotes metastasis. We recently showed that hemoglobin (Hb) forms a specific complex with TF purified from human malignant melanoma cells and enhances its procoagulant activity (PCA). To further study this interaction, we examined the effect of Hb on the expression of TF on human malignant (TF+) cells and KG1 myeloid leukemia (TF-) cells. Human melanoma A375 and J82 bladder carcinoma cells, which express TF at moderate and relatively high levels, respectively, were incubated with varying concentrations (0-1.5 mg/ml) of Hb. After washing, cells were analyzed for Hb binding and TF expression using flow cytometry and confocal microscopy. Hb bound to the cells in a concentration-dependent manner, and increased both TF expression and PCA. The human A375 malignant melanoma cells incubated with Hb (1 mg/ml) expressed up to six times more TF antigen than cells without Hb. This increase in TF expression and PCA of intact cells incubated with Hb was significantly inhibited by cycloheximide at a concentration of 10 microg/ml (P < 0.01). An increase in total cellular TF antigen content was demonstrated by specific immunoassay. In contrast, Hb (5 mg/ml) did not induce TF expression and PCA on KG1 cells as determined by flow cytometry and TF (FXAA) activity. We conclude that Hb specifically binds to TF-bearing malignant cells and increases their PCA. This effect seems to be at least partly due to de novo synthesis of TF and increased surface expression. However, the exact mechanism by which Hb binds and upregulates TF expression remains to be determined.

  13. Hemoglobin enhances tissue factor expression on human malignant cells.

    PubMed

    Siddiqui, F A; Amirkhosravi, A; Amaya, M; Meyer, T; Biggerstaff, J; Desai, H; Francis, J L

    2001-04-01

    Tissue Factor (TF) is a transmembrane glycoprotein that complexes with factor VII/activated factor VII to initiate blood coagulation. TF may be expressed on the surface of various cells including monocytes and endothelial cells. Over-expression of TF in human tumor cell lines promotes metastasis. We recently showed that hemoglobin (Hb) forms a specific complex with TF purified from human malignant melanoma cells and enhances its procoagulant activity (PCA). To further study this interaction, we examined the effect of Hb on the expression of TF on human malignant (TF+) cells and KG1 myeloid leukemia (TF-) cells. Human melanoma A375 and J82 bladder carcinoma cells, which express TF at moderate and relatively high levels, respectively, were incubated with varying concentrations (0-1.5 mg/ml) of Hb. After washing, cells were analyzed for Hb binding and TF expression using flow cytometry and confocal microscopy. Hb bound to the cells in a concentration-dependent manner, and increased both TF expression and PCA. The human A375 malignant melanoma cells incubated with Hb (1 mg/ml) expressed up to six times more TF antigen than cells without Hb. This increase in TF expression and PCA of intact cells incubated with Hb was significantly inhibited by cycloheximide at a concentration of 10 microg/ml (P < 0.01). An increase in total cellular TF antigen content was demonstrated by specific immunoassay. In contrast, Hb (5 mg/ml) did not induce TF expression and PCA on KG1 cells as determined by flow cytometry and TF (FXAA) activity. We conclude that Hb specifically binds to TF-bearing malignant cells and increases their PCA. This effect seems to be at least partly due to de novo synthesis of TF and increased surface expression. However, the exact mechanism by which Hb binds and upregulates TF expression remains to be determined. PMID:11414630

  14. Inhibition of Plasma Kallikrein by a Highly Specific Active Site Blocking Antibody

    PubMed Central

    Kenniston, Jon A.; Faucette, Ryan R.; Martik, Diana; Comeau, Stephen R.; Lindberg, Allison P.; Kopacz, Kris J.; Conley, Gregory P.; Chen, Jie; Viswanathan, Malini; Kastrapeli, Niksa; Cosic, Janja; Mason, Shauna; DiLeo, Mike; Abendroth, Jan; Kuzmic, Petr; Ladner, Robert C.; Edwards, Thomas E.; TenHoor, Christopher; Adelman, Burt A.; Nixon, Andrew E.; Sexton, Daniel J.

    2014-01-01

    Plasma kallikrein (pKal) proteolytically cleaves high molecular weight kininogen to generate the potent vasodilator and the pro-inflammatory peptide, bradykinin. pKal activity is tightly regulated in healthy individuals by the serpin C1-inhibitor, but individuals with hereditary angioedema (HAE) are deficient in C1-inhibitor and consequently exhibit excessive bradykinin generation that in turn causes debilitating and potentially fatal swelling attacks. To develop a potential therapeutic agent for HAE and other pKal-mediated disorders, we used phage display to discover a fully human IgG1 monoclonal antibody (DX-2930) against pKal. In vitro experiments demonstrated that DX-2930 potently inhibits active pKal (Ki = 0.120 ± 0.005 nm) but does not target either the zymogen (prekallikrein) or any other serine protease tested. These findings are supported by a 2.1-Å resolution crystal structure of pKal complexed to a DX-2930 Fab construct, which establishes that the pKal active site is fully occluded by the antibody. DX-2930 injected subcutaneously into cynomolgus monkeys exhibited a long half-life (t½ ∼12.5 days) and blocked high molecular weight kininogen proteolysis in activated plasma in a dose- and time-dependent manner. Furthermore, subcutaneous DX-2930 reduced carrageenan-induced paw edema in rats. A potent and long acting inhibitor of pKal activity could be an effective treatment option for pKal-mediated diseases, such as HAE. PMID:24970892

  15. Disulphide bond assignment in human tissue inhibitor of metalloproteinases (TIMP).

    PubMed Central

    Williamson, R A; Marston, F A; Angal, S; Koklitis, P; Panico, M; Morris, H R; Carne, A F; Smith, B J; Harris, T J; Freedman, R B

    1990-01-01

    Disulphide bonds in human recombinant tissue inhibitor of metalloproteinases (TIMP) were assigned by resolving proteolytic digests of TIMP on reverse-phase h.p.l.c. and sequencing those peaks judged to contain disulphide bonds by virtue of a change in retention time on reduction. This procedure allowed the direct assignment of Cys-145-Cys-166 and the isolation of two other peptides containing two disulphide bonds each. Further peptide cleavage in conjunction with fast-atom-bombardment m.s. analysis permitted the assignments Cys-1-Cys-70, Cys-3-Cys-99, Cys-13-Cys-124 and Cys-127-Cys-174 from these peptides. The sixth bond Cys-132-Cys-137 was assigned by inference, as the native protein has no detectable free thiol groups. Images Fig. 1. PMID:2163605

  16. Engineering bone tissue substitutes from human induced pluripotent stem cells

    PubMed Central

    de Peppo, Giuseppe Maria; Marcos-Campos, Iván; Kahler, David John; Alsalman, Dana; Shang, Linshan; Vunjak-Novakovic, Gordana; Marolt, Darja

    2013-01-01

    Congenital defects, trauma, and disease can compromise the integrity and functionality of the skeletal system to the extent requiring implantation of bone grafts. Engineering of viable bone substitutes that can be personalized to meet specific clinical needs represents a promising therapeutic alternative. The aim of our study was to evaluate the utility of human-induced pluripotent stem cells (hiPSCs) for bone tissue engineering. We first induced three hiPSC lines with different tissue and reprogramming backgrounds into the mesenchymal lineages and used a combination of differentiation assays, surface antigen profiling, and global gene expression analysis to identify the lines exhibiting strong osteogenic differentiation potential. We then engineered functional bone substitutes by culturing hiPSC-derived mesenchymal progenitors on osteoconductive scaffolds in perfusion bioreactors and confirmed their phenotype stability in a subcutaneous implantation model for 12 wk. Molecular analysis confirmed that the maturation of bone substitutes in perfusion bioreactors results in global repression of cell proliferation and an increased expression of lineage-specific genes. These results pave the way for growing patient-specific bone substitutes for reconstructive treatments of the skeletal system and for constructing qualified experimental models of development and disease. PMID:23653480

  17. Magnesium degradation products: effects on tissue and human metabolism.

    PubMed

    Seitz, J-M; Eifler, R; Bach, Fr-W; Maier, H J

    2014-10-01

    Owing to their mechanical properties, metallic materials present a promising solution in the field of resorbable implants. The magnesium metabolism in humans differs depending on its introduction. The natural, oral administration of magnesium via, for example, food, essentially leads to an intracellular enrichment of Mg(2+) . In contrast, introducing magnesium-rich substances or implants into the tissue results in a different decomposition behavior. Here, exposing magnesium to artificial body electrolytes resulted in the formation of the following products: magnesium hydroxide, magnesium oxide, and magnesium chloride, as well as calcium and magnesium apatites. Moreover, it can be assumed that Mg(2+) , OH(-) ions, and gaseous hydrogen are also present and result from the reaction for magnesium in an aqueous environment. With the aid of physiological metabolic processes, the organism succeeds in either excreting the above mentioned products or integrating them into the natural metabolic process. Only a burst release of these products is to be considered a problem. A multitude of general tissue effects and responses from the Mg's degradation products is considered within this review, which is not targeting specific implant classes. Furthermore, common alloying elements of magnesium and their hazardous potential in vivo are taken into account.

  18. Engineering bone tissue substitutes from human induced pluripotent stem cells.

    PubMed

    de Peppo, Giuseppe Maria; Marcos-Campos, Iván; Kahler, David John; Alsalman, Dana; Shang, Linshan; Vunjak-Novakovic, Gordana; Marolt, Darja

    2013-05-21

    Congenital defects, trauma, and disease can compromise the integrity and functionality of the skeletal system to the extent requiring implantation of bone grafts. Engineering of viable bone substitutes that can be personalized to meet specific clinical needs represents a promising therapeutic alternative. The aim of our study was to evaluate the utility of human-induced pluripotent stem cells (hiPSCs) for bone tissue engineering. We first induced three hiPSC lines with different tissue and reprogramming backgrounds into the mesenchymal lineages and used a combination of differentiation assays, surface antigen profiling, and global gene expression analysis to identify the lines exhibiting strong osteogenic differentiation potential. We then engineered functional bone substitutes by culturing hiPSC-derived mesenchymal progenitors on osteoconductive scaffolds in perfusion bioreactors and confirmed their phenotype stability in a subcutaneous implantation model for 12 wk. Molecular analysis confirmed that the maturation of bone substitutes in perfusion bioreactors results in global repression of cell proliferation and an increased expression of lineage-specific genes. These results pave the way for growing patient-specific bone substitutes for reconstructive treatments of the skeletal system and for constructing qualified experimental models of development and disease.

  19. Characterization of proopiomelanocortin transcripts in human nonpituitary tissues

    SciTech Connect

    Lacaze-Masmonteil, T.; De Keyzer, Y.; Luton, J.P.; Kahn, A.; Bertagna, X.

    1987-10-01

    Proopiomelanocortin (POMC), the precursor to adrenocorticotropic hormone and other related peptides, was originally identified in the corticotropic cell. Recent evidence shows that POMC products are also normally present in a variety of nonpituitary tissues. To investigate this phenomenon in humans the authors looked for the presence and characteristics of POMC transcripts in various adult tissues. Blot hybridization analysis of normal adrenal, thymus, and testis RNAs revealed a small RNA species approximately 400 nucleotides shorter than the 1200-nucleotide pituitary species. Primer extension and S1 nuclease mapping studies showed that this small RNA lacked exon 1 and exon 2 of the gene, and it corresponded to a set of at least six molecules starting 41 to 162 nucleotides downstream from the 5' end of exon 3. These RNAs appear to result from heterogeneous transcription initiation sites presumably under the control of GC box promoter sequences located in the 3' end of intron 2. They cannot encode a complete POMC molecule, and the only truncated POMC molecules that could be translated would lack a signal peptide necessary for membrane translocation and precursor processing. The use of highly sensitive S1 nuclease mapping techniques with uniformly labeled single-stranded DNA probes allowed the detection of a small but definite amount of the normal, 1200-nucleotide, mRNA species. It is suggested that it is this POMC mRNA that is responsible for the local production of all the POMC peptides.

  20. Second harmonic generation imaging of dermal collagen component in human keloid tissue

    NASA Astrophysics Data System (ADS)

    Yu, H. B.; Chen, S.; Zhu, X. Q.; Yang, H. Q.; Chen, J. X.

    2011-01-01

    In this paper, we report second harmonic generation (SHG) imaging of human keloid tissue. High resolution SHG images of collagen component were obtained in the superficial, medial and deep dermis of human keloid tissue, respectively. Our results show that this method has a capability to observe the structure of collagen component in human keloid tissue, which will help to better understand the formation process of human keloid scar at the molecular level.

  1. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins.

    PubMed

    Fukusumi, Hayato; Shofuda, Tomoko; Bamba, Yohei; Yamamoto, Atsuyo; Kanematsu, Daisuke; Handa, Yukako; Okita, Keisuke; Nakamura, Masaya; Yamanaka, Shinya; Okano, Hideyuki; Kanemura, Yonehiro

    2016-01-01

    Human neural progenitor cells (hNPCs) have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC) clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB) formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi). Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes.

  2. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins.

    PubMed

    Fukusumi, Hayato; Shofuda, Tomoko; Bamba, Yohei; Yamamoto, Atsuyo; Kanematsu, Daisuke; Handa, Yukako; Okita, Keisuke; Nakamura, Masaya; Yamanaka, Shinya; Okano, Hideyuki; Kanemura, Yonehiro

    2016-01-01

    Human neural progenitor cells (hNPCs) have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC) clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB) formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi). Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes. PMID:27212953

  3. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins

    PubMed Central

    Fukusumi, Hayato; Shofuda, Tomoko; Bamba, Yohei; Yamamoto, Atsuyo; Kanematsu, Daisuke; Handa, Yukako; Okita, Keisuke; Nakamura, Masaya; Yamanaka, Shinya; Okano, Hideyuki; Kanemura, Yonehiro

    2016-01-01

    Human neural progenitor cells (hNPCs) have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC) clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB) formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi). Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes. PMID:27212953

  4. Procoagulant tissue factor-exposing vesicles in human seminal fluid.

    PubMed

    Franz, C; Böing, A N; Hau, C M; Montag, M; Strowitzki, T; Nieuwland, R; Toth, B

    2013-06-01

    Recent studies indicate that various types of vesicles, like microparticles (MP) and exosomes, are present in blood, saliva, bone marrow, urine and synovial fluid. These vesicles, which are released upon activation or shear stress, are thought to play a role in coagulation, neovascularisation, inflammation and intercellular signalling. Seminal fluid is a cell-, sperm- and protein-rich suspension. Although seminal fluid is known to contain vesicles like prostasomes, MP and exosomes have never been characterised. Therefore, the aim of our study was to analyse and characterise vesicles in seminal fluid in male partners of patients undergoing controlled ovarian stimulation for IVF/ICSI. MP from seminal fluid of patients during routine IVF/ICSI procedures were detected and analysed with flow cytometry (FACS) and transmission electron microscopy (TEM), using antibodies against tissue factor (TF), CD10, CD13, CD26 and annexin V. The coagulant properties of vesicles were studied using a fibrin generation test. MP were detected in human seminal fluid by both flow cytometry and TEM. Seminal fluid-derived MP expressed CD10, CD13, CD26 and TF, which was highly procoagulant and a powerful trigger of the extrinsic pathway of coagulation. The extent to which the procoagulant activity of MP in seminal fluid contributes to the implantation process itself and therefore affects human reproduction needs to be further elucidated.

  5. Metabolic control analysis of respiration in human cancer tissue.

    PubMed

    Kaambre, Tuuli; Chekulayev, Vladimir; Shevchuk, Igor; Tepp, Kersti; Timohhina, Natalja; Varikmaa, Minna; Bagur, Rafaela; Klepinin, Aleksandr; Anmann, Tiia; Koit, Andre; Kaldma, Andrus; Guzun, Rita; Valvere, Vahur; Saks, Valdur

    2013-01-01

    Bioenergetic profiling of cancer cells is of great potential because it can bring forward new and effective therapeutic strategies along with early diagnosis. Metabolic Control Analysis (MCA) is a methodology that enables quantification of the flux control exerted by different enzymatic steps in a metabolic network thus assessing their contribution to the system's function. Our main goal is to demonstrate the applicability of MCA for in situ studies of energy metabolism in human breast and colorectal cancer cells as well as in normal tissues. We seek to determine the metabolic conditions leading to energy flux redirection in cancer cells. A main result obtained is that the adenine nucleotide translocator exhibits the highest control of respiration in human breast cancer thus becoming a prospective therapeutic target. Additionally, we present evidence suggesting the existence of mitochondrial respiratory supercomplexes that may represent a way by which cancer cells avoid apoptosis. The data obtained show that MCA applied in situ can be insightful in cancer cell energetic research.

  6. A chromatin immunoprecipitation (ChIP) protocol for use in whole human adipose tissue.

    PubMed

    Haim, Yulia; Tarnovscki, Tanya; Bashari, Dana; Rudich, Assaf

    2013-11-01

    Chromatin immunoprecipitation (ChIP) has become a central method when studying in vivo protein-DNA interactions, with the major challenge being the hope to capture "authentic" interactions. While ChIP protocols have been optimized for use with specific cell types and tissues including adipose tissue-derived cells, a working ChIP protocol addressing the challenges imposed by fresh whole human adipose tissue has not been described. Utilizing human paired omental and subcutaneous adipose tissue obtained during elective abdominal surgeries, we have carefully identified and optimized individual steps in the ChIP protocol employed directly on fresh tissue fragments. We describe a complete working protocol for using ChIP on whole adipose tissue fragments. Specific steps required adaptation of the ChIP protocol to human whole adipose tissue. In particular, a cross-linking step was performed directly on fresh small tissue fragments. Nuclei were isolated before releasing chromatin, allowing better management of fat content; a sonication protocol to obtain fragmented chromatin was optimized. We also demonstrate the high sensitivity of immunoprecipitated chromatin from adipose tissue to freezing. In conclusion, we describe the development of a ChIP protocol optimized for use in studying whole human adipose tissue, providing solutions for the unique challenges imposed by this tissue. Unraveling protein-DNA interaction in whole human adipose tissue will likely contribute to elucidating molecular pathways contributing to common human diseases such as obesity and type 2 diabetes.

  7. Characterization and comparison of adipose tissue-derived cells from human subcutaneous and omental adipose tissues.

    PubMed

    Toyoda, Mito; Matsubara, Yoshinori; Lin, Konghua; Sugimachi, Keizou; Furue, Masutaka

    2009-10-01

    Different fat depots contribute differently to disease and function. These differences may be due to the regional variation in cell types and inherent properties of fat cell progenitors. To address the differences of cell types in the adipose tissue from different depots, the phenotypes of freshly isolated adipose tissue-derived cells (ATDCs) from subcutaneous (SC) and omental (OM) adipose tissues were compared using flow cytometry. Our results showed that CD31(-)CD34(+)CD45(-)CD90(-)CD105(-)CD146(+) population, containing vascular smooth muscle cells and pericytes, was specifically defined in the SC adipose tissue while no such population was observed in OM adipose tissue. On the other hand, CD31(-)CD34(+)CD45(-)CD90(-)CD105(-)CD146(-) population, which is an undefined cell population, were found solely in OM adipose tissue. Overall, the SC adipose tissue contained more ATDCs than OM adipose tissue, while OM adipose tissue contained more blood-derived cells. Regarding to the inherent properties of fat cell progenitors from the two depots, adipose-derived stem cells (ADSCs) from SC had higher capacity to differentiate into both adipogenic and osteogenic lineages than those from OM, regardless of that the proliferation rates of ADSCs from both depots were similar. The higher differentiation capacity of ADSCs from SC adipose tissue suggests that SC tissue is more suitable cell source for regenerative medicine than OM adipose tissue.

  8. Impact of Statins on Gene Expression in Human Lung Tissues

    PubMed Central

    Lane, Jérôme; van Eeden, Stephan F.; Obeidat, Ma’en; Sin, Don D.; Tebbutt, Scott J.; Timens, Wim; Postma, Dirkje S.; Laviolette, Michel; Paré, Peter D.; Bossé, Yohan

    2015-01-01

    Statins are 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors that alter the synthesis of cholesterol. Some studies have shown a significant association of statins with improved respiratory health outcomes of patients with asthma, chronic obstructive pulmonary disease and lung cancer. Here we hypothesize that statins impact gene expression in human lungs and may reveal the pleiotropic effects of statins that are taking place directly in lung tissues. Human lung tissues were obtained from patients who underwent lung resection or transplantation. Gene expression was measured on a custom Affymetrix array in a discovery cohort (n = 408) and two replication sets (n = 341 and 282). Gene expression was evaluated by linear regression between statin users and non-users, adjusting for age, gender, smoking status, and other covariables. The results of each cohort were combined in a meta-analysis and biological pathways were studied using Gene Set Enrichment Analysis. The discovery set included 141 statin users. The lung mRNA expression levels of eighteen and three genes were up-regulated and down-regulated in statin users (FDR < 0.05), respectively. Twelve of the up-regulated genes were replicated in the first replication set, but none in the second (p-value < 0.05). Combining the discovery and replication sets into a meta-analysis improved the significance of the 12 up-regulated genes, which includes genes encoding enzymes and membrane proteins involved in cholesterol biosynthesis. Canonical biological pathways altered by statins in the lung include cholesterol, steroid, and terpenoid backbone biosynthesis. No genes encoding inflammatory, proteases, pro-fibrotic or growth factors were altered by statins, suggesting that the direct effect of statin in the lung do not go beyond its antilipidemic action. Although more studies are needed with specific lung cell types and different classes and doses of statins, the improved health outcomes and survival observed in statin

  9. The landscape of genomic imprinting across diverse adult human tissues

    PubMed Central

    Baran, Yael; Subramaniam, Meena; Biton, Anne; Tukiainen, Taru; Tsang, Emily K.; Rivas, Manuel A.; Pirinen, Matti; Gutierrez-Arcelus, Maria; Smith, Kevin S.; Kukurba, Kim R.; Zhang, Rui; Eng, Celeste; Torgerson, Dara G.; Urbanek, Cydney; Li, Jin Billy; Rodriguez-Santana, Jose R.; Burchard, Esteban G.; Seibold, Max A.; MacArthur, Daniel G.; Montgomery, Stephen B.; Zaitlen, Noah A.; Lappalainen, Tuuli

    2015-01-01

    Genomic imprinting is an important regulatory mechanism that silences one of the parental copies of a gene. To systematically characterize this phenomenon, we analyze tissue specificity of imprinting from allelic expression data in 1582 primary tissue samples from 178 individuals from the Genotype-Tissue Expression (GTEx) project. We characterize imprinting in 42 genes, including both novel and previously identified genes. Tissue specificity of imprinting is widespread, and gender-specific effects are revealed in a small number of genes in muscle with stronger imprinting in males. IGF2 shows maternal expression in the brain instead of the canonical paternal expression elsewhere. Imprinting appears to have only a subtle impact on tissue-specific expression levels, with genes lacking a systematic expression difference between tissues with imprinted and biallelic expression. In summary, our systematic characterization of imprinting in adult tissues highlights variation in imprinting between genes, individuals, and tissues. PMID:25953952

  10. Polybrominated diphenyl ethers: human tissue levels and toxicology.

    PubMed

    Gill, Udai; Chu, Ih; Ryan, John J; Feeley, Mark

    2004-01-01

    with behavioral alterations in neonatal mice. When considering the outlier value for PBDE-99 at 229 ng/g, this would result in an estimated PBDE-99 body burden of 46 microg/kg, or a MOS of only 9. However, no toxicokinetics data are available for humans, and the actual margin of safety may be much smaller if based on levels in critical target organs or tissues. PMID:15369322

  11. Viability of human composite tissue model for experimental study of burns.

    PubMed

    Qu, Miao; Kruse, Stephan; Pitsch, Heinz; Pallua, Norbert; Nourbakhsh, Mahtab

    2016-08-01

    Experimental studies of burns are primarily performed with animal models that have important anatomical and physiological differences relative to human systems. The aim of this study was to develop a human experimental burn model using composite tissue obtained from bariatric surgery. We established a new protocol to maintain viable sections of human cutaneous and subcutaneous (sub/cutaneous) tissue in vitro. Under the conditions selected, multiparametric flow cytometry and histological analysis confirmed the viability and integrity of the human sub/cutaneous tissue for at least 5 days. Furthermore, we utilized a precision McKenna burner to inflict burns on the human tissue model under well-defined thermal conditions in vitro. Our data showed a localized, temporally restricted polarization of the resident macrophages in the subcutaneous human tissue in response to specific thermal forces. Therefore, our model provides a useful alternative to animal studies for further detailed investigations of human responses to injuries and treatments. PMID:27585227

  12. Commodification of human tissue: implications for feminist and development ethics.

    PubMed

    Dickenson, Donna

    2002-05-01

    One effect of late capitalism--the commodification of practically everything--is to knock down the Chinese walls between the natural and productive realms, to use a Marxist framework. Women's labour in egg extraction and 'surrogate' motherhood might then be seen as what it is, labour which produces something of value. But this does not necessarily mean that women will benefit from the commodification of practically everything, in either North or South. In the newly developing biotechnologies involving stem cells, the reverse is more likely, particular given the the shortage in the North of the egg donors who will be increasingly necessary to therapeutic cloning. Although most of the ethical debate has focused on the status of the embryo, this is to define ethics with no reference to global or gender justice. There has been little or no debate about possible exploitation of women, particularly of ovum donors from the South. Countries of the South without national ethics committees or guidelines may be particularly vulnerable: although there is increasing awareness of the susceptibility of poorer countries to abuses in research ethics, very little has been written about how they might be affected by the enormously profitable new technologies exploiting human tissue. Even in the UK, although the new Medical Research Council guidelines make a good deal of the 'gift relationship', what they are actually about is commodification. If donors believe they are demonstrating altruism, but biotechnology firms and researchers use the discourse of commodity and profit, we have not 'incomplete commodification' but complete commodification with a plausibly human face. PMID:12872770

  13. Measurement of elastic wave dispersion on human femur tissue

    NASA Astrophysics Data System (ADS)

    Strantza, M.; Louis, O.; Polyzos, D.; Boulpaep, F.; Van Hemelrijck, D.; Aggelis, D. G.

    2014-03-01

    Cortical bone is one of the most complex heterogeneous media exhibiting strong wave dispersion. In such media when a burst of energy goes into the formation of elastic waves the different modes tend to separate according to the velocities of the frequency components as usually occurs in waveguides. In this study human femur specimens were subjected to elastic wave measurements. The main objective of the study is using broadband acoustic emission sensors to measure parameters like wave velocity dispersion and attenuation. Additionally, waveform parameters like the duration, rise time and average frequency, are also examined relatively to the propagation distance as a preparation for acoustic emission monitoring during fracture. To do so, four sensors were placed at adjacent positions on the surface of the cortical bone in order to record the transient response after pencil lead break excitation. The results are compared to similar measurements on a bulk metal piece which does not exhibit heterogeneity at the scale of the propagating wave lengths. It is shown that the microstructure of the tissue imposes a dispersive behavior for frequencies below 1 MHz and care should be taken for interpretation of the signals.

  14. High-Resolution NMR Studies of Human Tissue Factor

    PubMed Central

    Nuzzio, Kristin M.; Watt, Eric D.; Boettcher, John M.; Gajsiewicz, Joshua M.; Morrissey, James H.; Rienstra, Chad M.

    2016-01-01

    In normal hemostasis, the blood clotting cascade is initiated when factor VIIa (fVIIa, other clotting factors are named similarly) binds to the integral membrane protein, human tissue factor (TF). The TF/fVIIa complex in turn activates fX and fIX, eventually concluding with clot formation. Several X-ray crystal structures of the soluble extracellular domain of TF (sTF) exist; however, these structures are missing electron density in functionally relevant regions of the protein. In this context, NMR can provide complementary structural information as well as dynamic insights into enzyme activity. The resolution and sensitivity for NMR studies are greatly enhanced by the ability to prepare multiple milligrams of protein with various isotopic labeling patterns. Here, we demonstrate high-yield production of several isotopically labeled forms of recombinant sTF, allowing for high-resolution NMR studies both in the solid and solution state. We also report solution NMR spectra at sub-mM concentrations of sTF, ensuring the presence of dispersed monomer, as well as the first solid-state NMR spectra of sTF. Our improved sample preparation and precipitation conditions have enabled the acquisition of multidimensional NMR data sets for TF chemical shift assignment and provide a benchmark for TF structure elucidation. PMID:27657719

  15. Nattokinase-promoted tissue plasminogen activator release from human cells.

    PubMed

    Yatagai, Chieko; Maruyama, Masugi; Kawahara, Tomoko; Sumi, Hiroyuki

    2008-01-01

    When heated to a temperature of 70 degrees C or higher, the strong fibrinolytic activity of nattokinase in a solution was deactivated. Similar results were observed in the case of using Suc-Ala-Ala-Pro-Phe-pNA and H-D-Val-Leu-Lys-pNA, which are synthetic substrates of nattokinase. In the current study, tests were conducted on the indirect fibrinolytic effects of the substances containing nattokinase that had been deactivated through heating at 121 degrees C for 15 min. Bacillus subtilis natto culture solutions made from three types of bacteria strain were heat-treated and deactivated, and it was found that these culture solutions had the ability to generate tissue plasminogen activators (tPA) from vascular endothelial cells and HeLa cells at certain concentration levels. For example, it was found that the addition of heat-treated culture solution of the Naruse strain (undiluted solution) raises the tPA activity of HeLa cells to about 20 times that of the control. Under the same conditions, tPA activity was raised to a level about 5 times higher for human vascular endothelial cells (HUVEC), and to a level about 24 times higher for nattokinase sold on the market. No change in cell count was observed for HeLa cells and HUVEC in the culture solution at these concentrations, and the level of activity was found to vary with concentration.

  16. Binding of tissue plasminogen activator to cultured human endothelial cells.

    PubMed Central

    Hajjar, K A; Hamel, N M; Harpel, P C; Nachman, R L

    1987-01-01

    Tissue plasminogen activator (t-PA) and urokinase (u-PA), the major activators of plasminogen, are synthesized and released from endothelial cells. We previously demonstrated specific and functional binding of plasminogen to cultured human umbilical vein endothelial cells (HUVEC). In the present study we found that t-PA could bind to HUVEC. Binding of t-PA to HUVEC was specific, saturable, plasminogen-independent, and did not require lysine binding sites. The t-PA bound in a rapid and reversible manner, involving binding sites of both high (Kd, 28.7 +/- 10.8 pM; Bmax, 3,700 +/- 300) and low (Kd, 18.1 +/- 3.8 nM; Bmax 815,000 +/- 146,000) affinity. t-PA binding was 70% inhibited by a 100-fold molar excess of u-PA. When t-PA was bound to HUVEC, its apparent catalytic efficiency increased by three- or fourfold as measured by plasminogen activation. HUVEC-bound t-PA was active site-protected from its rapidly acting inhibitor: plasminogen activator inhibitor. These results demonstrate that t-PA specifically binds to HUVEC and that such binding preserves catalytic efficiency with respect to plasminogen activation. Therefore, endothelial cells can modulate hemostatic and thrombotic events at the cell surface by providing specific binding sites for activation of plasminogen. PMID:3119664

  17. Wave dispersion and attenuation on human femur tissue.

    PubMed

    Strantza, Maria; Louis, Olivia; Polyzos, Demosthenes; Boulpaep, Frans; van Hemelrijck, Danny; Aggelis, Dimitrios G

    2014-01-01

    Cortical bone is a highly heterogeneous material at the microscale and has one of the most complex structures among materials. Application of elastic wave techniques to this material is thus very challenging. In such media the initial excitation energy goes into the formation of elastic waves of different modes. Due to "dispersion", these modes tend to separate according to the velocities of the frequency components. This work demonstrates elastic wave measurements on human femur specimens. The aim of the study is to measure parameters like wave velocity, dispersion and attenuation by using broadband acoustic emission sensors. First, four sensors were placed at small intervals on the surface of the bone to record the response after pencil lead break excitations. Next, the results were compared to measurements on a bulk steel block which does not exhibit heterogeneity at the same wave lengths. It can be concluded that the microstructure of the tissue imposes a dispersive behavior for frequencies below 1 MHz and care should be taken for interpretation of the signals. Of particular interest are waveform parameters like the duration, rise time and average frequency, since in the next stage of research the bone specimens will be fractured with concurrent monitoring of acoustic emission. PMID:25196011

  18. Comparative Transcriptome Analysis Reveals Substantial Tissue Specificity in Human Aortic Valve

    PubMed Central

    Wang, Jun; Wang, Ying; Gu, Weidong; Ni, Buqing; Sun, Haoliang; Yu, Tong; Gu, Wanjun; Chen, Liang; Shao, Yongfeng

    2016-01-01

    RNA sequencing (RNA-seq) has revolutionary roles in transcriptome identification and quantification of different types of tissues and cells in many organisms. Although numerous RNA-seq data derived from many types of human tissues and cell lines, little is known on the transcriptome repertoire of human aortic valve. In this study, we sequenced the total RNA prepared from two calcified human aortic valves and reported the whole transcriptome of human aortic valve. Integrating RNA-seq data of 13 human tissues from Human Body Map 2 Project, we constructed a transcriptome repertoire of human tissues, including 19,505 protein-coding genes and 4,948 long intergenic noncoding RNAs (lincRNAs). Among them, 263 lincRNAs were identified as novel noncoding transcripts in our data. By comparing transcriptome data among different human tissues, we observed substantial tissue specificity of RNA transcripts, both protein-coding genes and lincRNAs, in human aortic valve. Further analysis revealed that aortic valve-specific lincRNAs were more likely to be recently derived from repetitive elements in the primate lineage, but were less likely to be conserved at the nucleotide level. Expression profiling analysis showed significant lower expression levels of aortic valve-specific protein-coding genes and lincRNA genes, when compared with genes that were universally expressed in various tissues. Isoform-level expression analysis also showed that a majority of mRNA genes had a major isoform expressed in the human aortic valve. To our knowledge, this is the first comparative transcriptome analysis between human aortic valve and other human tissues. Our results are helpful to understand the transcriptome diversity of human tissues and the underlying mechanisms that drive tissue specificity of protein-coding genes and lincRNAs in human aortic valve. PMID:27493474

  19. Comparative Transcriptome Analysis Reveals Substantial Tissue Specificity in Human Aortic Valve.

    PubMed

    Wang, Jun; Wang, Ying; Gu, Weidong; Ni, Buqing; Sun, Haoliang; Yu, Tong; Gu, Wanjun; Chen, Liang; Shao, Yongfeng

    2016-01-01

    RNA sequencing (RNA-seq) has revolutionary roles in transcriptome identification and quantification of different types of tissues and cells in many organisms. Although numerous RNA-seq data derived from many types of human tissues and cell lines, little is known on the transcriptome repertoire of human aortic valve. In this study, we sequenced the total RNA prepared from two calcified human aortic valves and reported the whole transcriptome of human aortic valve. Integrating RNA-seq data of 13 human tissues from Human Body Map 2 Project, we constructed a transcriptome repertoire of human tissues, including 19,505 protein-coding genes and 4,948 long intergenic noncoding RNAs (lincRNAs). Among them, 263 lincRNAs were identified as novel noncoding transcripts in our data. By comparing transcriptome data among different human tissues, we observed substantial tissue specificity of RNA transcripts, both protein-coding genes and lincRNAs, in human aortic valve. Further analysis revealed that aortic valve-specific lincRNAs were more likely to be recently derived from repetitive elements in the primate lineage, but were less likely to be conserved at the nucleotide level. Expression profiling analysis showed significant lower expression levels of aortic valve-specific protein-coding genes and lincRNA genes, when compared with genes that were universally expressed in various tissues. Isoform-level expression analysis also showed that a majority of mRNA genes had a major isoform expressed in the human aortic valve. To our knowledge, this is the first comparative transcriptome analysis between human aortic valve and other human tissues. Our results are helpful to understand the transcriptome diversity of human tissues and the underlying mechanisms that drive tissue specificity of protein-coding genes and lincRNAs in human aortic valve. PMID:27493474

  20. Characterization of RNA isolated from eighteen different human tissues: results from a rapid human autopsy program.

    PubMed

    Walker, Douglas G; Whetzel, Alexis M; Serrano, Geidy; Sue, Lucia I; Lue, Lih-Fen; Beach, Thomas G

    2016-09-01

    Many factors affect the integrity of messenger RNA from human autopsy tissues including postmortem interval (PMI) between death and tissue preservation and the pre-mortem agonal and disease states. In this communication, we describe RNA isolation and characterization of 389 samples from 18 different tissues from elderly donors who were participants in a rapid whole-body autopsy program located in Sun City, Arizona ( www.brainandbodydonationprogram.org ). Most tissues were collected within a PMI of 2-6 h (median 3.15 h; N = 455), but for this study, tissue from cases with longer PMIs (1.25-29.25 h) were included. RNA quality was assessed by RNA integrity number (RIN) and total yield (ng RNA/mg tissue). RIN correlated with PMI for heart (r = -0.531, p = 0.009) and liver (r = -558, p = 0.0017), while RNA yield correlated with PMI for colon (r = -485, p = 0.016) and skin (r = -0.460, p = 0.031). RNAs with the lowest integrity were from skin and cervix where 22.7 and 31.4 % of samples respectively failed to produce intact RNA; by contrast all samples from esophagus, lymph node, jejunum, lung, stomach, submandibular gland and kidney produced RNA with measurable RINs. Expression levels in heart RNA of 4 common housekeeping normalization genes showed significant correlations of Ct values with RIN, but only one gene, glyceraldehyde-3 phosphate dehydrogenase, showed a correlation of Ct with PMI. There were no correlations between RIN values obtained for liver, adrenal, cervix, esophagus and lymph node and those obtained from corresponding brain samples. We show that high quality RNA can be produced from most human autopsy tissues, though with significant differences between tissues and donors. The RNA stability and yield did not depend solely on PMI; other undetermined factors are involved, but these do not include the age of the donor.

  1. [Kallikrein-kinin system as a target for diabetic retinopathy treatment].

    PubMed

    Iarovaia, G A; Neshkova, E A; Blokhina, T B; Kochergin, S A; Vorob'eva, I V; Gigineishvili, D N

    2012-01-01

    Multifactor etiology of diabetic retinopathy (DR) determines difficulty of understanding of pathogenesis and need of search of effective approaches to study key mechanisms of development of this microvascular complication of diabetes mellitus (DM). Significant achievements of the last years show the contribution of two proteolytic systems into pathogenesis of DR, that control vascular tone and permeability - kallikrein-kinin (KKS) and renin-angiotensin systems (RAS). Among new approaches to DR treatment one of the most appropriate is an influence on KKS by means of inhibiting kallikrein, that leads to reduction of retinal vascular permeability and allows to prevent the development of macula oedema and other consequences of vascular wall damage in DR. PMID:22994115

  2. Xenotransplantation Models to Study the Effects of Toxicants on Human Fetal Tissues1

    PubMed Central

    Spade, Daniel J.; McDonnell, Elizabeth V.; Heger, Nicholas E.; Sanders, Jennifer A.; Saffarini, Camelia M.; Gruppuso, Philip A.; De Paepe, Monique E.; Boekelheide, Kim

    2015-01-01

    Many diseases that manifest throughout the lifetime are influenced by factors affecting fetal development. Fetal exposure to xenobiotics, in particular, may influence the development of adult diseases. Established animal models provide systems for characterizing both developmental biology and developmental toxicology. However, animal model systems do not allow researchers to assess the mechanistic effects of toxicants on developing human tissue. Human fetal tissue xenotransplantation models have recently been implemented to provide human-relevant mechanistic data on the many tissue-level functions that may be affected by fetal exposure to toxicants. This review describes the development of human fetal tissue xenotransplant models for testis, prostate, lung, liver, and adipose tissue, aimed at studying the effects of xenobiotics on tissue development, including implications for testicular dysgenesis, prostate disease, lung disease, and metabolic syndrome. The mechanistic data obtained from these models can complement data from epidemiology, traditional animal models, and in vitro studies to quantify the risks of toxicant exposures during human development. PMID:25477288

  3. Revisions to Exceptions Applicable to Certain Human Cells, Tissues, and Cellular and Tissue-Based Products. Final rule.

    PubMed

    2016-06-22

    : The Food and Drug Administration (FDA or Agency or we) is issuing this final rule to amend certain regulations regarding donor eligibility, including the screening and testing of donors of particular human cells, tissues, and cellular and tissue-based products (HCT/Ps), and related labeling. This final rule is in response to our enhanced understanding in this area and in response to comments from stakeholders regarding the importance of embryos to individuals and couples seeking access to donated embryos. PMID:27373010

  4. Revisions to Exceptions Applicable to Certain Human Cells, Tissues, and Cellular and Tissue-Based Products. Final rule.

    PubMed

    2016-06-22

    : The Food and Drug Administration (FDA or Agency or we) is issuing this final rule to amend certain regulations regarding donor eligibility, including the screening and testing of donors of particular human cells, tissues, and cellular and tissue-based products (HCT/Ps), and related labeling. This final rule is in response to our enhanced understanding in this area and in response to comments from stakeholders regarding the importance of embryos to individuals and couples seeking access to donated embryos.

  5. What have we learned about the kallikrein-kinin and renin-angiotensin systems in neurological disorders?

    PubMed

    Naffah-Mazzacoratti, Maria da Graça; Gouveia, Telma Luciana Furtado; Simões, Priscila Santos Rodrigues; Perosa, Sandra Regina

    2014-05-26

    The kallikrein-kinin system (KKS) is an intricate endogenous pathway involved in several physiological and pathological cascades in the brain. Due to the pathological effects of kinins in blood vessels and tissues, their formation and degradation are tightly controlled. Their components have been related to several central nervous system diseases such as stroke, Alzheimer's disease, Parkinson's disease, multiple sclerosis, epilepsy and others. Bradykinin and its receptors (B1R and B2R) may have a role in the pathophysiology of certain central nervous system diseases. It has been suggested that kinin B1R is up-regulated in pathological conditions and has a neurodegenerative pattern, while kinin B2R is constitutive and can act as a neuroprotective factor in many neurological conditions. The renin angiotensin system (RAS) is an important blood pressure regulator and controls both sodium and water intake. AngII is a potent vasoconstrictor molecule and angiotensin converting enzyme is the major enzyme responsible for its release. AngII acts mainly on the AT1 receptor, with involvement in several systemic and neurological disorders. Brain RAS has been associated with physiological pathways, but is also associated with brain disorders. This review describes topics relating to the involvement of both systems in several forms of brain dysfunction and indicates components of the KKS and RAS that have been used as targets in several pharmacological approaches. PMID:24921004

  6. Patents on Technologies of Human Tissue and Organ Regeneration from Pluripotent Human Embryonic Stem Cells

    PubMed Central

    Parsons, Xuejun H; Teng, Yang D; Moore, Dennis A; Snyder, Evan Y

    2011-01-01

    Human embryonic stem cells (hESCs) are genetically stable with unlimited expansion ability and unrestricted plasticity, proffering a pluripotent reservoir for in vitro derivation of a large supply of disease-targeted human somatic cells that are restricted to the lineage in need of repair. There is a large healthcare need to develop hESC-based therapeutic solutions to provide optimal regeneration and reconstruction treatment options for the damaged or lost tissue or organ that have been lacking. In spite of controversy surrounding the ownership of hESCs, the number of patent applications related to hESCs is growing rapidly. This review gives an overview of different patent applications on technologies of derivation, maintenance, differentiation, and manipulation of hESCs for therapies. Many of the published patent applications have been based on previously established methods in the animal systems and multi-lineage inclination of pluripotent cells through spontaneous germ-layer differentiation. Innovative human stem cell technologies that are safe and effective for human tissue and organ regeneration in the clinical setting remain to be developed. Our overall view on the current patent situation of hESC technologies suggests a trend towards hESC patent filings on novel therapeutic strategies of direct control and modulation of hESC pluripotent fate, particularly in a 3-dimensional context, when deriving clinically-relevant lineages for regenerative therapies. PMID:23355961

  7. [Tissue engineering and construction of human skin in vitro].

    PubMed

    Arvelo, Francisco

    2007-09-01

    Tissue engineering is the new science that has come to make possible the growth of new organ tissue from small fragments of healthy tissue, thus partially or totally restoring the lost functions of ill tissues or organs, as shown by the achievements made with the culture of skin, cornea or cartilage. Thus far, this new science is able to ensure the recovery of lost functions and, doubtlessly, in a near future will be capable of developing tissues and organs not unlike natural ones. In our laboratory we have began the development of tissue engineering techniques for the successful construction of in vitro skin with the aim at mid term of producing cornea and cartilage. In a first clinical trial, these techniques were applied in the treatment of chronic skin lesions and the advantages and reach of these new tools were demonstrated for the effective solution of problems with would otherwise not be easily solved through the use of conventional treatments.

  8. [Tissue engineering and construction of human skin in vitro].

    PubMed

    Arvelo, Francisco

    2007-09-01

    Tissue engineering is the new science that has come to make possible the growth of new organ tissue from small fragments of healthy tissue, thus partially or totally restoring the lost functions of ill tissues or organs, as shown by the achievements made with the culture of skin, cornea or cartilage. Thus far, this new science is able to ensure the recovery of lost functions and, doubtlessly, in a near future will be capable of developing tissues and organs not unlike natural ones. In our laboratory we have began the development of tissue engineering techniques for the successful construction of in vitro skin with the aim at mid term of producing cornea and cartilage. In a first clinical trial, these techniques were applied in the treatment of chronic skin lesions and the advantages and reach of these new tools were demonstrated for the effective solution of problems with would otherwise not be easily solved through the use of conventional treatments. PMID:17853796

  9. NMDA receptors are expressed in human ovarian cancer tissues and human ovarian cancer cell lines

    PubMed Central

    North, William G; Liu, Fuli; Tian, Ruiyang; Abbasi, Hamza; Akerman, Bonnie

    2015-01-01

    We have earlier demonstrated that breast cancer and small-cell lung cancer express functional NMDA receptors that can be targeted to promote cancer cell death. Human ovarian cancer tissues and human ovarian cancer cell lines (SKOV3, A2008, and A2780) have now been shown to also express NMDA-receptor subunit 1 (GluN1) and subunit 2B (GluN2B). Seventeen ovarian cancers in two arrays were screened by immunohistochemistry using polyclonal antibodies that recognize an extracellular moiety on GluN1 and on GluN2B. These specimens comprised malignant tissue with pathology diagnoses of serous papillary cystadenocarcinoma, endometrioid adenocarcinoma, and clear-cell carcinoma. Additionally, archival tissues defined as ovarian adenocarcinoma from ten patients treated at this institute were also evaluated. All of the cancerous tissues demonstrated positive staining patterns with the NMDA-receptor antibodies, while no staining was found for tumor-adjacent normal tissues or sections of normal ovarian tissue. Human ovarian adenocarcinoma cell lines (A2008, A2780, SKOV3) were demonstrated to express GluN1 by Western blotting, but displayed different levels of expression. Through immunocytochemistry utilizing GluN1 antibodies and imaging using a confocal microscope, we were able to demonstrate that GluN1 protein is expressed on the surface of these cells. In addition to these findings, GluN2B protein was demonstrated to be expressed using polyclonal antibodies against this protein. Treatment of all ovarian cell lines with antibodies against GluN1 was found to result in decreased cell viability (P<0.001), with decreases to 10%–25% that of untreated cells. Treatment of control HEK293 cells with various dilutions of GluN1 antibodies had no effect on cell viability. The GluN1 antagonist MK-801 (dizocilpine maleate) and the GluN2B antagonist ifenprodil, like antibodies, dramatically decreased the viability of A2780 ovarian tumor cells (P<0.01). Treatment of A2780 tumor xenografts with

  10. NMDA receptors are expressed in human ovarian cancer tissues and human ovarian cancer cell lines.

    PubMed

    North, William G; Liu, Fuli; Tian, Ruiyang; Abbasi, Hamza; Akerman, Bonnie

    2015-01-01

    We have earlier demonstrated that breast cancer and small-cell lung cancer express functional NMDA receptors that can be targeted to promote cancer cell death. Human ovarian cancer tissues and human ovarian cancer cell lines (SKOV3, A2008, and A2780) have now been shown to also express NMDA-receptor subunit 1 (GluN1) and subunit 2B (GluN2B). Seventeen ovarian cancers in two arrays were screened by immunohistochemistry using polyclonal antibodies that recognize an extracellular moiety on GluN1 and on GluN2B. These specimens comprised malignant tissue with pathology diagnoses of serous papillary cystadenocarcinoma, endometrioid adenocarcinoma, and clear-cell carcinoma. Additionally, archival tissues defined as ovarian adenocarcinoma from ten patients treated at this institute were also evaluated. All of the cancerous tissues demonstrated positive staining patterns with the NMDA-receptor antibodies, while no staining was found for tumor-adjacent normal tissues or sections of normal ovarian tissue. Human ovarian adenocarcinoma cell lines (A2008, A2780, SKOV3) were demonstrated to express GluN1 by Western blotting, but displayed different levels of expression. Through immunocytochemistry utilizing GluN1 antibodies and imaging using a confocal microscope, we were able to demonstrate that GluN1 protein is expressed on the surface of these cells. In addition to these findings, GluN2B protein was demonstrated to be expressed using polyclonal antibodies against this protein. Treatment of all ovarian cell lines with antibodies against GluN1 was found to result in decreased cell viability (P<0.001), with decreases to 10%-25% that of untreated cells. Treatment of control HEK293 cells with various dilutions of GluN1 antibodies had no effect on cell viability. The GluN1 antagonist MK-801 (dizocilpine maleate) and the GluN2B antagonist ifenprodil, like antibodies, dramatically decreased the viability of A2780 ovarian tumor cells (P<0.01). Treatment of A2780 tumor xenografts with

  11. EDTA separation and recombination of epithelium and connective tissue of human oral mucosa. Studies of tissue transplants in nude mice.

    PubMed

    Holmstrup, P; Dabelsteen, E; Harder, F

    1985-01-01

    A possible epithelial-mesenchymal interaction in determining epithelial histologic features of human oral mucosa was examined. The study comprised 74 biopsies of normal buccal mucosa and 54 biopsies of normal palatal mucosa. Epithelium was separated from connective tissue by the use of 1 mM ethylenediamine tetraacetate dihydrate. Self-recombined and cross-recombined epithelial and connective tissues and connective tissue sheets alone were transplanted to subcutaneous sites of nude mice. Histologic examination of cross-recombined palatal epithelium/buccal connective tissue transplants showed a change in keratinization pattern but no major change in number of epithelial cell layers as the result of connective tissue influence. Transplanted sheets of connective tissue after growth for 14 days showed that complete separation of biopsies from buccal mucosa had been obtained. However, palatal mucosa had been incompletely separated as evidenced by re-epithelialization of most of the connective tissue transplants. The consequences of the incomplete palatal epithelium-connective tissue separation are discussed.

  12. Tissue Specificity of Human Angiotensin I-Converting Enzyme

    PubMed Central

    Kryukova, Olga V.; Tikhomirova, Victoria E.; Golukhova, Elena Z.; Evdokimov, Valery V.; Kalantarov, Gavreel F.; Trakht, Ilya N.; Schwartz, David E.; Dull, Randal O.; Gusakov, Alexander V.; Uporov, Igor V.; Kost, Olga A.; Danilov, Sergei M.

    2015-01-01

    Background Angiotensin-converting enzyme (ACE), which metabolizes many peptides and plays a key role in blood pressure regulation and vascular remodeling, as well as in reproductive functions, is expressed as a type-1 membrane glycoprotein on the surface of endothelial and epithelial cells. ACE also presents as a soluble form in biological fluids, among which seminal fluid being the richest in ACE content - 50-fold more than that in blood. Methods/Principal Findings We performed conformational fingerprinting of lung and seminal fluid ACEs using a set of monoclonal antibodies (mAbs) to 17 epitopes of human ACE and determined the effects of potential ACE-binding partners on mAbs binding to these two different ACEs. Patterns of mAbs binding to ACEs from lung and from seminal fluid dramatically differed, which reflects difference in the local conformations of these ACEs, likely due to different patterns of ACE glycosylation in the lung endothelial cells and epithelial cells of epididymis/prostate (source of seminal fluid ACE), confirmed by mass-spectrometry of ACEs tryptic digests. Conclusions Dramatic differences in the local conformations of seminal fluid and lung ACEs, as well as the effects of ACE-binding partners on mAbs binding to these ACEs, suggest different regulation of ACE functions and shedding from epithelial cells in epididymis and prostate and endothelial cells of lung capillaries. The differences in local conformation of ACE could be the base for the generation of mAbs distingushing tissue-specific ACEs. PMID:26600189

  13. [Purification of kininogen II (LMW) from human plasma].

    PubMed

    Díaz, A; Henriques, O B

    1976-01-01

    Plasma supernatant in which kallikrein has been activated and removed by glass powder whilst kininogen I (HMW) has been consumed by the activated kallikrein, was used for the preparation of kininogen II. It was purified by chromatography on DEAE-cellulose followed by gel filtration on G-200 Sephadex. The purification of kininogen II was assessed from determinations of the amount of kinin released (expressed as bradykinin) as measured on the isolated guinea pig ileum, using samples incubated with human salivary kallikrein or trypsin. A preparation of kininogen II containing an activity equivalent to 8 microgram Br/mg protein, was obtained. Salivary kallikrein released approximately three times more kinin from the substrate as compared to trypsin. PMID:1032267

  14. Needle optical coherence elastography for the measurement of microscale mechanical contrast deep within human breast tissues

    NASA Astrophysics Data System (ADS)

    Kennedy, Kelsey M.; McLaughlin, Robert A.; Kennedy, Brendan F.; Tien, Alan; Latham, Bruce; Saunders, Christobel M.; Sampson, David D.

    2013-12-01

    Optical coherence elastography (OCE) is an emerging imaging technique that probes microscale mechanical contrast in tissues with the potential to differentiate healthy and malignant tissues. However, conventional OCE techniques are limited to imaging the first 1 to 2 mm of tissue in depth. We demonstrate, for the first time, OCE measurements deep within human tissues using needle OCE, extending the potential of OCE as a surgical guidance tool. We use needle OCE to detect tissue interfaces based on mechanical contrast in both normal and malignant breast tissues in freshly excised human mastectomy samples, as validated against histopathology. Further, we demonstrate the feasibility of in situ measurements >4 cm from the tissue surface using ultrasound guidance of the OCE needle probe. With further refinement, our method may potentially aid in accurate detection of the boundary of the tumor to help ensure full removal of all malignant tissues, which is critical to the success of breast-conserving surgery.

  15. 2D Representation of Transcriptomes by t-SNE Exposes Relatedness between Human Tissues

    PubMed Central

    Taskesen, Erdogan; Reinders, Marcel J. T.

    2016-01-01

    The GTEx Consortium reported that hierarchical clustering of RNA profiles from 25 unique tissue types among 1641 individuals accurately distinguished the tissue types, but a multidimensional scaling failed to generate a 2D projection of the data that separates tissue-subtypes. In this study we show that a projection by t-Distributed Stochastic Neighbor Embedding is in line with the cluster analysis which allows a more detailed examination and visualization of human tissue relationships. PMID:26906061

  16. Sharing and Specificity of Co-expression Networks across 35 Human Tissues.

    PubMed

    Pierson, Emma; Koller, Daphne; Battle, Alexis; Mostafavi, Sara; Ardlie, Kristin G; Getz, Gad; Wright, Fred A; Kellis, Manolis; Volpi, Simona; Dermitzakis, Emmanouil T

    2015-05-01

    To understand the regulation of tissue-specific gene expression, the GTEx Consortium generated RNA-seq expression data for more than thirty distinct human tissues. This data provides an opportunity for deriving shared and tissue specific gene regulatory networks on the basis of co-expression between genes. However, a small number of samples are available for a majority of the tissues, and therefore statistical inference of networks in this setting is highly underpowered. To address this problem, we infer tissue-specific gene co-expression networks for 35 tissues in the GTEx dataset using a novel algorithm, GNAT, that uses a hierarchy of tissues to share data between related tissues. We show that this transfer learning approach increases the accuracy with which networks are learned. Analysis of these networks reveals that tissue-specific transcription factors are hubs that preferentially connect to genes with tissue specific functions. Additionally, we observe that genes with tissue-specific functions lie at the peripheries of our networks. We identify numerous modules enriched for Gene Ontology functions, and show that modules conserved across tissues are especially likely to have functions common to all tissues, while modules that are upregulated in a particular tissue are often instrumental to tissue-specific function. Finally, we provide a web tool, available at mostafavilab.stat.ubc.ca/GNAT, which allows exploration of gene function and regulation in a tissue-specific manner.

  17. ESTIV questionnaire on the acquisition and use of primary human cells and tissue in toxicology.

    PubMed

    Sladowski, Dariusz; Combes, Robert; van der Valk, Jan; Nawrot, Ireneusz; Gut, Grzegorz

    2005-10-01

    The ability to use human cells and tissues in toxicology research and testing has the benefit that it obviates the need to undertake species extrapolation when assessing human hazard. However, obtaining and using human cells and tissues is logistically difficult, ethically complex and is a potential source of infections to those coming into contact with human cell material. The issue is also controversial, with the recent EU legislation draft on tissue engineering, and also due to some instances of human material being obtained and used without informed consent. There are also varying regulations and attitudes relating to the use of human cells and tissues throughout Member States of the EU, and there is a need for harmonisation. The European Society of Toxicology in Vitro (ESTIV) Executive Board and the European Network of Human Research Tissue Banks (ENRTB) have conducted a survey to ascertain the extent to which human cells and tissues are used by its members, how these are obtained, what local regulations are in force, how the material is used, and the advantages and disadvantages experienced by members in using such material, as opposed to cell lines. The results obtained have been compared with the results from a previous survey conducted in 2000. It is hoped that this information will help to facilitate the process of acquiring and using human cells and tissues in a safe and effective way to promote the use of non-animal approaches for investigating the mechanisms of toxicity, and for predicting the toxic hazard of substances. PMID:16150566

  18. Mechanical properties of human autologous tubular connective tissues (human biotubes) obtained from patients undergoing peritoneal dialysis.

    PubMed

    Nakayama, Yasuhide; Kaneko, Yoshiyuki; Takewa, Yoshiaki; Okumura, Noriko

    2016-10-01

    Completely autologous in vivo tissue-engineered connective tissue tubes (Biotubes) have promise as arterial vascular grafts in animal implantation studies. In this clinical study of patients undergoing peritoneal dialysis (PD) (n = 11; age: 39-83 years), we evaluated human Biotubes' (h-Biotubes) mechanical properties to determine whether Biotubes with feasibility as vascular grafts could be formed in human bodies. We extracted PD catheters, embedded for 4-47 months, and obtained tubular connective tissues as h-Biotubes (internal diameter: 5 mm) from around the catheter' silicone tubular parts. h-Biotubes were composed mainly of collagen with smooth luminal surfaces. The average wall thickness was 278 ± 178 μm. No relationship was founded between the tubes' mechanical properties and patients' ages or PD catheter embedding periods statistically. However, the elastic modulus (2459 ± 970 kPa) and tensile strength (623 ± 314 g) of h-Biotubes were more than twice as great as those from animal Biotubes, formed from the same PD catheters by embedding in the beagle subcutaneous pouches for 1 month, or beagle arteries. The burst strength (6338 ± 1106 mmHg) of h-Biotubes was almost the same as that of the beagle thoracic or abdominal aorta. h-Biotubes could be formed in humans over a 4-month embedding period, and they satisfied the mechanical requirements for application as vascular grafts. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 1431-1437, 2016.

  19. X-ray microscopy of soft and hard human tissues

    NASA Astrophysics Data System (ADS)

    Müller, Bert; Schulz, Georg; Deyhle, Hans; Stalder, Anja K.; Ilgenstein, Bernd; Holme, Margaret N.; Weitkamp, Timm; Beckmann, Felix; Hieber, Simone E.

    2016-01-01

    The simultaneous post mortem visualization of soft and hard tissues using absorption-based CT remains a challenge. If the photon energy is optimized for the visualization of hard tissue, the surrounding soft tissue components are almost X-ray transparent. Therefore, the combination with other modalities such as phase-contrast CT, magnetic resonance microscopy, and histology is essential to detect the anatomical features. The combination of the 2D and 3D data sets using sophisticated segmentation and registration tools allows for conclusions about otherwise inaccessible anatomical features essential for improved patient treatments.

  20. Reconstruction of genome-scale metabolic models for 126 human tissues using mCADRE

    PubMed Central

    2012-01-01

    Background Human tissues perform diverse metabolic functions. Mapping out these tissue-specific functions in genome-scale models will advance our understanding of the metabolic basis of various physiological and pathological processes. The global knowledgebase of metabolic functions categorized for the human genome (Human Recon 1) coupled with abundant high-throughput data now makes possible the reconstruction of tissue-specific metabolic models. However, the number of available tissue-specific models remains incomplete compared with the large diversity of human tissues. Results We developed a method called metabolic Context-specificity Assessed by Deterministic Reaction Evaluation (mCADRE). mCADRE is able to infer a tissue-specific network based on gene expression data and metabolic network topology, along with evaluation of functional capabilities during model building. mCADRE produces models with similar or better functionality and achieves dramatic computational speed up over existing methods. Using our method, we reconstructed draft genome-scale metabolic models for 126 human tissue and cell types. Among these, there are models for 26 tumor tissues along with their normal counterparts, and 30 different brain tissues. We performed pathway-level analyses of this large collection of tissue-specific models and identified the eicosanoid metabolic pathway, especially reactions catalyzing the production of leukotrienes from arachidnoic acid, as potential drug targets that selectively affect tumor tissues. Conclusions This large collection of 126 genome-scale draft metabolic models provides a useful resource for studying the metabolic basis for a variety of human diseases across many tissues. The functionality of the resulting models and the fast computational speed of the mCADRE algorithm make it a useful tool to build and update tissue-specific metabolic models. PMID:23234303

  1. Effect of metoprolol on 24-hour urinary excretion of adrenal steroids and kallikrein in patients with essential hypertension.

    PubMed Central

    Fritschka, E.; Gotzen, R.; Kittler, R.; Schöneshöfer, M.

    1984-01-01

    Treatment of fifteen patients with essential hypertension over four weeks using the beta 1-adrenoceptor blocking agent, metoprolol, resulted in a decrease in 24 h urinary excretion of kallikrein and aldosterone along with a decrease in plasma renin activity. There was no significant change in 24 h excretion rates of the free adrenal steroids deoxycorticosterone, 18-OH-deoxycorticosterone, corticosterone, cortisol or 18-OH-corticosterone during treatment, which were not significantly different from excretion rates of normal males, thus excluding inhibitory effects of adrenal steroids on urinary kallikrein activity. A positive correlation was found between plasma renin activity and urinary excretion of kallikrein during the control period and after 2 weeks on metoprolol, supporting the assumption of a preserved link between the renin-angiotensin-aldosterone system and the renal excretion of kallikrein in these patients. The decrease in kallikrein excretion during beta 1-adrenoceptor blockade in patients with essential hypertension may be explained by a reduction in sympathetic tone and by reduced activity of the renin-aldosterone system. PMID:6367871

  2. Human keratin diseases: hereditary fragility of specific epithelial tissues.

    PubMed

    Corden, L D; McLean, W H

    1996-12-01

    Keratins are heteropolymeric proteins which form the intermediate filament cytoskeleton in epithelial cells. Since 1991, mutations in several keratin genes have been found to cause a variety of human diseases affecting the epidermis and other epithelial structures. Epidermolysis bullosa simplex (EBS) was the first mechanobullous disease for which the underlying genetic lesion was found, with mutations in both the K5 and K14 genes rendering basal epidermal keratinocytes less resilient to trauma, resulting in skin fragility. The site of mutation in the keratin protein correlates with phenotypic severity in this disorder. Since mutations were identified in the basal cell keratins, the total number of keratin genes associated with diseases has risen to eleven. The rod domains of suprabasal keratins K1 and K10 are mutated in bullous congenital ichthyosiform erythroderma (BCIE; also called epidermolytic hyperkeratosis, EH) and mosaicism for K1/K10 mutations results in a nevoid distribution of EH. An unusual mutation in the VI domain of K1 has also been found to cause diffuse non-epidermolytic palmoplantar keratoderma (DNEPPK). Mutations in palmoplantar specific keratin K9 cause epidermolytic palmoplantar keratoderma (EPPK) and mutations in the late differentiation suprabasal keratin K2e cause ichthyosis bullosa of Siemens (IBS). In the last year or so, mutations were discovered in differentiation specific keratins K6a and K16 causing pachyonychia congenita type 1 and K17 mutations occur in pachyonychia congenita type 2. K16 and K17 mutations have also been reported to produce phenotypes with little or no nail changes: K16 mutations can present as focal non-epidermolytic palmoplantar keratoderma (NEPPK) and K17 mutations can result in a phenotype resembling steatocystoma multiplex. Recently, mutation of mucosal keratin pair K4 and K13 has been shown to underlie white sponge nevus (WSN). This year, the first mutations in a keratin-associated protein, plectin, were shown to

  3. Human intestinal tissue antibiotic concentrations. Clindamycin, gentamicin, and mezlocillin.

    PubMed

    Thadepalli, H; Lou, M A; Prabhala, R H; Mandal, A K

    1990-11-01

    An antibiotic, to be effective for prophylaxis in abdominal trauma, should quickly achieve high concentrations in the intestinal wall and at enough inhibitory levels to kill most aerobic and anaerobic bacteria that are potential contaminants at the site of surgical incision. Therefore, we studied the intestinal tissue levels of clindamycin, gentamicin, and mezlocillin to see whether the tissue levels achieved by these antibiotics in the intestinal tissue were adequate. A single dose of mezlocillin, 4 grams; clindamycin, 600 mg and gentamicin, 80 mg; quickly reached the desired concentrations, i.e., 52.3, 9.69 and 6.1 micrograms/gram of intestinal tissue respectively. These levels were high enough to inhibit the growth of most isolates of E. coli and B. fragilis, common pathogens involved in intra-abdominal abscess.

  4. Electrical impedance characterization of normal and cancerous human hepatic tissue.

    PubMed

    Laufer, Shlomi; Ivorra, Antoni; Reuter, Victor E; Rubinsky, Boris; Solomon, Stephen B

    2010-07-01

    The four-electrode method was used to measure the ex vivo complex electrical impedance of tissues from 14 hepatic tumors and the surrounding normal liver from six patients. Measurements were done in the frequency range 1-400 kHz. It was found that the conductivity of the tumor tissue was much higher than that of the normal liver tissue in this frequency range (from 0.14 +/- 0.06 S m(-1) versus 0.03 +/- 0.01 S m(-1) at 1 kHz to 0.25 +/- 0.06 S m(-1) versus 0.15 +/- 0.03 S m(-1) at 400 kHz). The Cole-Cole models were estimated from the experimental data and the four parameters (rho(0), rho(infinity), alpha, f(c)) were obtained using a least-squares fit algorithm. The Cole-Cole parameters for the cancerous and normal liver are 9 +/- 4 Omega m(-1), 2.2 +/- 0.7 Omega m(-1), 0.5 +/- 0.2, 140 +/- 103 kHz and 50 +/- 28 Omega m(-1), 3.2 +/- 0.6 Omega m(-1), 0.64 +/- 0.04, 10 +/- 7 kHz, respectively. These data can contribute to developing bioelectric applications for tissue diagnostics and in tissue treatment planning with electrical fields such as radiofrequency tissue ablation, electrochemotherapy and gene therapy with reversible electroporation, nanoscale pulsing and irreversible electroporation.

  5. Effects of simulated microgravity on human brain nervous tissue.

    PubMed

    Wang, Xianghan; Du, Jianxin; Wang, Demei; Zeng, Fan; Wei, Yukui; Wang, Fuli; Feng, Chengcheng; Li, Nuomin; Dai, Rongji; Deng, Yulin; Quan, Zhenzhen; Qing, Hong

    2016-08-01

    During spaceflight, the negative effects of space microgravity on astronauts are becoming more and more prominent, and especially, of which on the nervous system is urgently to be solved. For this purpose tissue blocks and primary cells of nervous tissues obtained from glioma of patients were cultivated after culturing for about 7days, explanted tissues and cells were then randomly divided into two groups, one for static culture (control group, C), and the other for rotary processing for 1day, 3days, 5days, 7days and 14days (experiment group, E). Figures captured by inverted microscope revealed that, with short time rotating for 1day or 3days, morphology changes of tissue blocks were not obvious. When the rotary time was extended to 7days or 14days, it was found that cell somas is significantly larger and the ability of adhesion is declined in comparison with that in control group. Additionally, the arrangement of cells migrated from explanted tissues was disorganized, and the migration distance became shorter. In immunofluorescence analysis, β-tubulin filaments in control group appeared to organize into bundles. While in experiment group, β-tubulin was highly disorganized. In conclusion, simulated microgravity treatment for a week affected the morphology of nervous tissue, and caused highly disorganized distribution of cytoskeleton and the increase of cell apoptosis. These morphological changes might be one of the causes of apoptosis induced by simulated microgravity. PMID:27268042

  6. Online quantitative analysis of multispectral images of human body tissues

    SciTech Connect

    Lisenko, S A

    2013-08-31

    A method is developed for online monitoring of structural and morphological parameters of biological tissues (haemoglobin concentration, degree of blood oxygenation, average diameter of capillaries and the parameter characterising the average size of tissue scatterers), which involves multispectral tissue imaging, image normalisation to one of its spectral layers and determination of unknown parameters based on their stable regression relation with the spectral characteristics of the normalised image. Regression is obtained by simulating numerically the diffuse reflectance spectrum of the tissue by the Monte Carlo method at a wide variation of model parameters. The correctness of the model calculations is confirmed by the good agreement with the experimental data. The error of the method is estimated under conditions of general variability of structural and morphological parameters of the tissue. The method developed is compared with the traditional methods of interpretation of multispectral images of biological tissues, based on the solution of the inverse problem for each pixel of the image in the approximation of different analytical models. (biomedical optics)

  7. The effects of corrosive substances on human bone, teeth, hair, nails, and soft tissue.

    PubMed

    Hartnett, Kristen M; Fulginiti, Laura C; Di Modica, Frank

    2011-07-01

    This research investigates the effects of household chemicals on human tissues. Five different human tissues (bone, tooth, hair, fingernails, and skin/muscle/fat) were immersed into six different corrosive agents. These agents consisted of hydrochloric acid, sulfuric acid, lye, bleach, organic septic cleaner, and Coca-Cola(®) soda. Tap water was used as a control. Tissue samples were cut to consistent sizes and submerged in the corrosive liquids. Over time, the appearance, consistency, and weight were documented. Hydrochloric acid was the most destructive agent in this study, consuming most tissues within 24 h. Sulfuric acid was the second most destructive agent in this study. Bleach, lye, and cola had no structural effects on the hard tissues of the body, but did alter the appearance or integrity of the hair, nails, or flesh in some way. The organic septic cleaner and tap water had no effect on any of the human tissue tested during the timeframe of the study. PMID:21447075

  8. Determination of optical parameters of human breast tissue from spatially resolved fluorescence: a diffusion theory model

    NASA Astrophysics Data System (ADS)

    Nair, Maya S.; Ghosh, Nirmalya; Raju, Narisetti Sundar; Pradhan, Asima

    2002-07-01

    We report the measurement of optical transport parameters of pathologically characterized malignant tissues, normal tissues, and different types of benign tumors of the human breast in the visible wavelength region. A spatially resolved steady-state diffuse fluorescence reflectance technique was used to estimate the values for the reduced-scattering coefficient (mu's) and the absorption coefficient (mua) of human breast tissues at three wavelengths (530, 550, and 590 nm). Different breast tissues could be well differentiated from one another, and different benign tumors could also be distinguished by their measured transport parameters. A diffusion theory model was developed to describe fluorescence light energy distribution, especially its spatial variation in a turbid and multiply scattering medium such as human tissue. The validity of the model was checked with a Monte Carlo simulation and also with different tissue phantoms prepared with polystyrene microspheres as scatterers, riboflavin as fluorophores, and methylene blue as absorbers.

  9. The effects of corrosive substances on human bone, teeth, hair, nails, and soft tissue.

    PubMed

    Hartnett, Kristen M; Fulginiti, Laura C; Di Modica, Frank

    2011-07-01

    This research investigates the effects of household chemicals on human tissues. Five different human tissues (bone, tooth, hair, fingernails, and skin/muscle/fat) were immersed into six different corrosive agents. These agents consisted of hydrochloric acid, sulfuric acid, lye, bleach, organic septic cleaner, and Coca-Cola(®) soda. Tap water was used as a control. Tissue samples were cut to consistent sizes and submerged in the corrosive liquids. Over time, the appearance, consistency, and weight were documented. Hydrochloric acid was the most destructive agent in this study, consuming most tissues within 24 h. Sulfuric acid was the second most destructive agent in this study. Bleach, lye, and cola had no structural effects on the hard tissues of the body, but did alter the appearance or integrity of the hair, nails, or flesh in some way. The organic septic cleaner and tap water had no effect on any of the human tissue tested during the timeframe of the study.

  10. Comparison of different fabrication techniques for human adipose tissue engineering in severe combined immunodeficient mice.

    PubMed

    Frerich, Bernhard; Winter, Karsten; Scheller, Konstanze; Braumann, Ulf-Dietrich

    2012-03-01

    Adipose tissue engineering has been advocated for soft-tissue augmentation and for the treatment of soft tissue defects. The efficacy in terms of persistence of the engineered fat is, however, not yet understood and could depend on the nature of fabrication and application. The high metabolic demand of adipose tissue also points to the problem of vascularization. Endothelial cell (EC) cotransplantation could be a solution. Human adipose tissue-derived stromal cells were seeded on collagen microcarriers and submitted to adipogenic differentiation ("microparticles"). In a first run of experiments, these microparticles were implanted under the skin of severe combined immunodeficient (SCID) mice (n = 45) with and without the addition of human umbilical vein ECs (HUVECs). A group of carriers without any cells served as control. In a second run, adipose tissue constructs were fabricated by embedding microparticles in fibrin matrix with and without the addition of HUVEC, and were also implanted in SCID mice (n = 30). The mice were sacrificed after 12 days, 4 weeks, and 4 months. Mature adipose tissue, fibrous tissue, and acellular regions were quantified on whole-specimen histological sections. The implantation of microparticles showed a better sustainment of tissue volume and a higher degree of mature adipose tissue compared with adipose tissue constructs. Immunohistology proved obviously perfused human tissue-engineered vessels. There was a limited but not significant advantage in EC cotransplantation after 4 weeks in terms of tissue volume. In groups with EC cotransplantation, there were significantly fewer acellular/necrotic areas after 4 weeks and 4 months. In conclusion, the size of the implanted tissue equivalents is a crucial parameter, affecting volume maintenance and the gain of mature adipose tissue. EC cotransplantation leads to functional stable vascular networks connecting in part to the host vasculature and contributing to tissue perfusion; however

  11. Anti-Human Tissue Factor Antibody Ameliorated Intestinal Ischemia Reperfusion-Induced Acute Lung Injury in Human Tissue Factor Knock-In Mice

    PubMed Central

    Mura, Marco; Li, Li; Cypel, Marcelo; Soderman, Avery; Picha, Kristen; Yang, Jing; Liu, Mingyao

    2008-01-01

    Background Interaction between the coagulation and inflammation systems plays an important role in the development of acute respiratory distress syndrome (ARDS). Anti-coagulation is an attractive option for ARDS treatment, and this has promoted development of new antibodies. However, preclinical trials for these antibodies are often limited by the high cost and availability of non-human primates. In the present study, we developed a novel alternative method to test the role of a humanized anti-tissue factor mAb in acute lung injury with transgenic mice. Methodology/Principal Findings Human tissue factor knock-in (hTF-KI) transgenic mice and a novel humanized anti-human tissue factor mAb (anti-hTF mAb, CNTO859) were developed. The hTF-KI mice showed a normal and functional expression of hTF. The anti-hTF mAb specifically blocked the pro-coagulation activity of brain extracts from the hTF-KI mice and human, but not from wild type mice. An extrapulmonary ARDS model was used by intestinal ischemia-reperfusion. Significant lung tissue damage in hTF-KI mice was observed after 2 h reperfusion. Administration of CNTO859 (5 mg/kg, i.v.) attenuated the severity of lung tissue injury, decreased the total cell counts and protein concentration in bronchoalveolar lavage fluid, and reduced Evans blue leakage. In addition, the treatment significantly reduced alveolar fibrin deposition, and decreased tissue factor and plasminogen activator inhibitor-1 activity in the serum. This treatment also down-regulated cytokine expression and reduced cell death in the lung. Conclusions This novel anti-hTF antibody showed beneficial effects on intestinal ischemia-reperfusion induced acute lung injury, which merits further investigation for clinical usage. In addition, the use of knock-in transgenic mice to test the efficacy of antibodies against human-specific proteins is a novel strategy for preclinical studies. PMID:18231608

  12. Ethical issues surrounding the transplantation of human fetal tissues.

    PubMed

    Hurd, R E

    1992-12-01

    Organ transplants have been one of the greatest advances in medicine. However, organs from living relatives or cadavers are in short supply, and many people die awaiting a donor organ. Increasing the donor pool by using organs from aborted fetuses has been proposed to increase the supply. In addition, there are benefits of using fetal tissue including its particular usefulness in children, the fact that it is not readily rejected, and its potential for growth. Guidelines for fetal research were issued in 1975, but a research moratorium was imposed in 1988 to allow study of ethical and legal issues. While the federal government delays in lifting the ban, several states have written laws governing experimentation with fetuses. Ethical arguments against using fetal tissue for organ transplant include a concern that this would create a branch of biomedicine which depends on the continuation of induced abortions. This could lead to neglect of research for other therapies. The timing and type of abortion should continue to benefit the mother, rather than the organ recipient. Ethicists debate whether or not use of aborted tissue implies complicity in the abortion process beyond that which exists for all members of a society which permits abortion. They also wonder whether knowing that some good could come of an abortion would influence a woman's decision to have one. Proposals to keep the use of fetal tissue ethical include banning the commercial use of sale of tissues, forbidding designation of the tissue recipient (to prevent harvesting fetal tissue for a relative), separating abortion counseling and management from harvesting of the tissue, and obtaining informed consent (perhaps from a proxy surrogate rather than from the mother) for the use of fetal tissue. When the medical and ethical communities have reached some consensus on these issues, crafted safeguards, and precluded conflicts of interest, then restrictions on government funding should be lifted. Whereas it

  13. Distribution of the human intracellular serpin protease inhibitor 8 in human tissues.

    PubMed

    Strik, Merel C; Bladergroen, Bellinda A; Wouters, Dorine; Kisiel, Walter; Hooijberg, Jan Hendrik; Verlaan, Angelique R; Hordijk, Peter L; Schneider, Pascal; Hack, C Erik; Kummer, J Alain

    2002-11-01

    Ovalbumin-like serine protease inhibitors are mainly localized intracellularly and their in vivo functions are largely unknown. To elucidate their physiological role(s), we studied the expression of one of these inhibitors, protease inhibitor 8 (PI-8), in normal human tissues by immunohistochemistry using a PI-8-specific monoclonal antibody. PI-8 was strongly expressed in the nuclei of squamous epithelium of mouth, pharynx, esophagus, and epidermis, and by the epithelial layer of skin appendages, particularly by more differentiated epithelial cells. PI-8 was also expressed by monocytes and by neuroendocrine cells in the pituitary gland, pancreas, and digestive tract. Monocytes showed nuclear and cytoplasmic localization of PI-8, whereas neuroendocrine cells showed only cytoplasmic staining. In vitro nuclear localization of PI-8 was confirmed by confocal analysis using serpin-transfected HeLa cells. Furthermore, mutation of the P(1) residue did not affect the subcellular distribution pattern of PI-8, indicating that its nuclear localization is independent of the interaction with its target protease. We conclude that PI-8 has a unique distribution pattern in human tissues compared to the distribution patterns of other intracellular serpins. Additional studies must be performed to elucidate its physiological role.

  14. ["Prostatic" kallikreins, sex hormones and insulin-like growth factors: complex of male and female regulatory elements in health and carcinogenesis].

    PubMed

    Zezerov, E G; Severin, E S

    1999-01-01

    Publications (up to the year 1997) on the interactions of "prostatic" kallikreins (prostatic specific antigen-PSA, etc.), sex hormones, insulin-like growth factors (IGF) and proteins binding them (IGFBP) in physiological processes (ageing, menstrual cycle, pregnancy) and oncogenesis (prostatic and mammary cancer) are reviewed. The concentrations of PSA, IGF, and IGFBP in organs and liquid media of men and women are presented. A concept of similarity in the mechanisms of interactions of sex hormones (dihydrotestosterone in men and progesterone in women), PSA, IGFBP, and IGF during activation of anabolic and proliferative processes in health and carcinogenesis is presented as a scheme. The diagnostic and prognostic value of PSA as a cancer marker should not be confined to male tumors (prostatic cancer and benign prostatic hyperplasia). Our data permits us to regard PSA as an oncofetal marker for men and women, indicating normal and neoplastic proliferative processes in the prostatic, mammary, salivary, and other glands and in the lungs and endometrium. Diagnostic and prognostic significance of PSA in breast cancer is shown. The traditional name "PSA" does not reflect its physiological and pathogenetic role as a member of the kallikrein family with chymotrypsin-like activity. PSA is not absolutely specific towards the producer organ and sex. Its relative specificity for the prostate is undoubted, because the content of PSA in prostatic tissue and seminal plasma is 10(6)-10(8) times higher than in the serum and other organs of men and women. Therefore, although the terms "prostatic, "specific", and "antigen" now became trivial and it is difficult to refuse from them, they can be used only in quotation marks.

  15. Identification of cyclopropaneoctanoic acid 2-hexyl in human adipose tissue and serum.

    PubMed

    Sledzinski, Tomasz; Mika, Adriana; Stepnowski, Piotr; Proczko-Markuszewska, Monika; Kaska, Lukasz; Stefaniak, Tomasz; Swierczynski, Julian

    2013-08-01

    Fatty acids containing a cyclopropane ring in their structure (cyclopropane FA) have been found in a wide variety of bacteria, a number of protozoa, and Myriapoda. Little is known about cyclopropane FA in mammal, especially in human tissues. The present study deals with the identification of cyclopropane FA in adipose tissue and serum of humans and rats. Fatty acids extracted from the adipose tissue and serum obtained from obese women during bariatric surgery were methylated and analyzed on GC-MS. We have identified: cyclopropaneoctanoic acid 2-hexyl, cyclopropaneoctanoic acid 2-octyl, cyclopropanenonanoic acid, and 2-[[2-[(2-ethylcyclopropyl)methyl]cyclopropyl]methyl] acid in human adipose tissue. We confirmed the presence of cyclopropaneoctanoic acid 2-hexyl by derivatization of FA extracted from human adipose tissue to picolinyl esters. Cyclopropaneoctanoic acid 2-hexyl was the main cyclopropane FA (approximately 0.4 % of total fatty acids in human adipose tissue, and about 0.2 % of total fatty acids in the serum). In adipose tissue cyclopropaneoctanoic acid 2-hexyl was found mainly in triacylglycerols, whereas in serum in phospholipids and triacylglycerols. The cyclopropaneoctanoic acid 2-hexyl has also been found in serum, and adipose tissue of rats in amounts comparable to humans. The content of cyclopropaneoctanoic acid 2-hexyl decreased in adipose tissue of rats maintained on a restricted diet for 1 month. In conclusion, we demonstrated that cyclopropaneoctanoic acid 2-hexyl is present in human adipose tissue and serum. Adipose tissue cyclopropaneoctanoic acid 2-hexyl is stored mainly in triacylglycerols and the storage of this cyclopropane FA is affected by food restriction.

  16. Human Cardiac Tissue Engineering: From Pluripotent Stem Cells to Heart Repair

    PubMed Central

    Jackman, Christopher P.; Shadrin, Ilya Y.; Carlson, Aaron L.; Bursac, Nenad

    2014-01-01

    Engineered cardiac tissues hold great promise for use in drug and toxicology screening, in vitro studies of human physiology and disease, and as transplantable tissue grafts for myocardial repair. In this review, we discuss recent progress in cell-based therapy and functional tissue engineering using pluripotent stem cell-derived cardiomyocytes and we describe methods for delivery of cells into the injured heart. While significant hurdles remain, notable advances have been made in the methods to derive large numbers of pure human cardiomyocytes, mature their phenotype, and produce and implant functional cardiac tissues, bringing the field a step closer to widespread in vitro and in vivo applications. PMID:25599018

  17. Genome-wide prediction and analysis of human tissue-selective genes using microarray expression data

    PubMed Central

    2013-01-01

    Background Understanding how genes are expressed specifically in particular tissues is a fundamental question in developmental biology. Many tissue-specific genes are involved in the pathogenesis of complex human diseases. However, experimental identification of tissue-specific genes is time consuming and difficult. The accurate predictions of tissue-specific gene targets could provide useful information for biomarker development and drug target identification. Results In this study, we have developed a machine learning approach for predicting the human tissue-specific genes using microarray expression data. The lists of known tissue-specific genes for different tissues were collected from UniProt database, and the expression data retrieved from the previously compiled dataset according to the lists were used for input vector encoding. Random Forests (RFs) and Support Vector Machines (SVMs) were used to construct accurate classifiers. The RF classifiers were found to outperform SVM models for tissue-specific gene prediction. The results suggest that the candidate genes for brain or liver specific expression can provide valuable information for further experimental studies. Our approach was also applied for identifying tissue-selective gene targets for different types of tissues. Conclusions A machine learning approach has been developed for accurately identifying the candidate genes for tissue specific/selective expression. The approach provides an efficient way to select some interesting genes for developing new biomedical markers and improve our knowledge of tissue-specific expression. PMID:23369200

  18. KeyGenes, a Tool to Probe Tissue Differentiation Using a Human Fetal Transcriptional Atlas

    PubMed Central

    Roost, Matthias S.; van Iperen, Liesbeth; Ariyurek, Yavuz; Buermans, Henk P.; Arindrarto, Wibowo; Devalla, Harsha D.; Passier, Robert; Mummery, Christine L.; Carlotti, Françoise; de Koning, Eelco J.P.; van Zwet, Erik W.; Goeman, Jelle J.; Chuva de Sousa Lopes, Susana M.

    2015-01-01

    Summary Differentiated derivatives of human pluripotent stem cells in culture are generally phenotypically immature compared to their adult counterparts. Their identity is often difficult to determine with certainty because little is known about their human fetal equivalents in vivo. Cellular identity and signaling pathways directing differentiation are usually determined by extrapolating information from either human adult tissue or model organisms, assuming conservation with humans. To resolve this, we generated a collection of human fetal transcriptional profiles at different developmental stages. Moreover, we developed an algorithm, KeyGenes, which uses this dataset to quantify the extent to which next-generation sequencing or microarray data resemble specific cell or tissue types in the human fetus. Using KeyGenes combined with the human fetal atlas, we identified multiple cell and tissue samples unambiguously on a limited set of features. We thus provide a flexible and expandable platform to monitor and evaluate the efficiency of differentiation in vitro. PMID:26028532

  19. Activation of the NLRP3/caspase-1 inflammasome in human dental pulp tissue and human dental pulp fibroblasts.

    PubMed

    Jiang, Wenkai; Lv, Haipeng; Wang, Haijing; Wang, Diya; Sun, Shukai; Jia, Qian; Wang, Peina; Song, Bing; Ni, Longxing

    2015-08-01

    The NLRP3/caspase-1 inflammasome pathway plays an important role in cellular immune defence against bacterial infection; however, its function in human dental pulp tissue and human dental pulp fibroblasts remains poorly understood. We demonstrate that NLRP3 protein expression occurs to a greater extent in pulp tissue with irreversible pulpitis than in normal pulp tissue and in tissue with reversible pulpitis. Caspase-1 is present in its active (cleaved) form only in pulp tissue with irreversible pulpitis. NLRP3 and caspase-1 are expressed in the odontoblast layers in normal human dental pulp tissue, whereas in inflamed pulp tissue, the odontoblast layers are disrupted and dental pulp cells are positive for NLRP3 and caspase-1. Additionally, we investigate the role of the NLRP3/caspase-1 inflammasome pathway in human dental pulp fibroblasts and show that ATP activates the P2X7 receptor on the cell membrane triggering K(+) efflux and inducing the gradual recruitment of the membrane pore pannexin-1. Extracellular lipopolysaccharide is able to penetrate the cytosol and activate NLRP3. Furthermore, the low intracellular K(+) concentration in the cytosol triggers reactive oxygen species generation, which also induces the NLRP3 inflammasome. Thus, the NLRP3/caspase-1 pathway has a biological role in the innate immune response mounted by human dental pulp fibroblasts.

  20. Understanding multicellular function and disease with human tissue-specific networks

    PubMed Central

    Greene, Casey S.; Krishnan, Arjun; Wong, Aaron K.; Ricciotti, Emanuela; Zelaya, Rene A.; Himmelstein, Daniel S.; Zhang, Ran; Hartmann, Boris M.; Zaslavsky, Elena; Sealfon, Stuart C.; Chasman, Daniel I.; FitzGerald, Garret A.; Dolinski, Kara; Grosser, Tilo; Troyanskaya, Olga G.

    2016-01-01

    Tissue and cell-type identity lie at the core of human physiology and disease. Understanding the genetic underpinnings of complex tissues and individual cell lineages is crucial for developing improved diagnostics and therapeutics. We present genome-wide functional interaction networks for 144 human tissues and cell types developed using a data-driven Bayesian methodology that integrates thousands of diverse experiments spanning tissue and disease states. Tissue-specific networks predict lineage-specific responses to perturbation, reveal genes’ changing functional roles across tissues, and illuminate disease-disease relationships. We introduce NetWAS, which combines genes with nominally significant GWAS p-values and tissue-specific networks to identify disease-gene associations more accurately than GWAS alone. Our webserver, GIANT, provides an interface to human tissue networks through multi-gene queries, network visualization, analysis tools including NetWAS, and downloadable networks. GIANT enables systematic exploration of the landscape of interacting genes that shape specialized cellular functions across more than one hundred human tissues and cell types. PMID:25915600

  1. Analysis of mechanical interaction between human gluteal soft tissue and body supports.

    PubMed

    Then, C; Menger, J; Benderoth, G; Alizadeh, M; Vogl, T J; Hübner, F; Silber, G

    2008-01-01

    Pressure sores are the most common complication associated with patient immobilization. They develop through sustained localized tissue strain and stress, primarily caused by body supports. Modifying support design can reduce the risk and extent of pressure sore development with computational simulations helping to provide insight into tissue stress-strain distribution. Appropriate material parameters for human soft tissue and support material, as well as precise anatomical modelling, are indispensable in this process. A finite element (FE) model of the human gluteal region based on magnetic resonance imaging (MRI) data has been developed. In vivo human gluteal skin/fat and muscle long-term material parameters as well as open-cell polyurethane foam support long-term material parameters have been characterised. The Ogden form for slightly compressible materials was employed to describe human gluteal soft tissue behaviour. Altering support geometries and support materials, effects on human gluteal soft tissue could be quantified. FE-analysis indicated maximal tissue stress at the muscle-bone interface, not at the skin. Shear strain maxima were found in the muscle layer near the fat-muscle interface. Maximum compressive stress magnitude at the sacral bone depended strongly on the behaviour of the pelvic diaphragm musculature. We hypothesize that the compliance of the muscles forming the pelvic diaphragm govern the relative motion of the buttock tissue to the adjacent bone structure under compression, thus influencing tissue stress magnitudes.

  2. Comparative pathology of canine soft tissue sarcomas: possible models of human non-rhabdomyosarcoma soft tissue sarcomas.

    PubMed

    Milovancev, M; Hauck, M; Keller, C; Stranahan, L W; Mansoor, A; Malarkey, D E

    2015-01-01

    Comparative analyses of canine and human soft tissue sarcomas (STSs) are lacking. This study compared the histological and immunohistochemical (labelling for desmin, smooth muscle actin [SMA], CD31, pancytokeratin, S100 and CD34) appearance of 32 archived, formalin-fixed, paraffin wax-embedded canine STS tumour specimens by board-certified veterinary and medical pathologists, both blinded to the other's interpretations. Comparison between the veterinary and human diagnoses revealed a generally consistent pattern of interpretation with few notable variations. Most tumours (13/32) were judged to display similar histomorphological appearance to human low-grade spindle cell sarcomas, appearing non-distinctive and morphologically of a fibroblastic/myofibroblastic type. Five canine cases resembled human liposarcoma, but with atypical desmin-positive epithelioid cells present. Five canine cases resembled human spindle cell sarcoma with myxoid features and two additional cases resembled human myxofibrosarcoma. Seven canine cases were noted to resemble human undifferentiated sarcoma. Findings in the present study demonstrate that canine STSs display histological and immunohistochemical features similar to their human equivalents. Because of these cross-species similarities, a particular opportunity exists to understand the biology and treatment of human STS by potentially including dogs as clinical models. PMID:25435513

  3. Multiple-Image Radiography for Human Soft Tissue

    SciTech Connect

    Muehleman,C.; Li, J.; Zhong, Z.; Brankov, J.; Wernick, M.

    2006-01-01

    Conventional radiography only provides a measure of the X-ray attenuation caused by an object; thus, it is insensitive to other inherent informative effects, such as refraction. Furthermore, conventional radiographs are degraded by X-ray scatter that can obscure important details of the object being imaged. The novel X-ray technology diffraction-enhanced imaging (DEI) has recently allowed the visualization of nearly scatter-free images displaying both attenuation and refraction properties. A new method termed multiple-image radiography (MIR) is a significant improvement over DEI, corrects errors in DEI, is more robust to noise and produces an additional image that is entirely new to medical imaging. This new image, which portrays ultra-small-angle X-ray scattering (USAXS) conveys the presence of microstructure in the object, thus differentiating homogeneous tissues from tissues that are irregular on a scale of micrometers. The aim of this study was to examine the use of MIR for evaluation of soft tissue, and in particular to conduct a preliminary investigation of the USAXS image, which has not previously been used in tissue imaging.

  4. Assessment of bioburden on human and animal tissues: part 2--results of testing of human tissue and qualification of a composite sample for routine bioburden determination.

    PubMed

    Kowalski, John B; Merritt, Karen; Gocke, David; Osborne, Joel

    2012-08-01

    A quantitative method was developed and validated to assess bioburden on tissue from human donors and to compare bioburden determination results to swab culture results from the same donor. An initial study with allograft tissue from 101 donors showed a wide range of bioburden levels; values from no colony-forming units (CFU) detected to >28,000 CFU were observed. Tissues from donors that had swab cultures negative for objectionable microorganisms generally had lower bioburden than tissues from donors where objectionable microorganisms were recovered by swab culturing. In a follow-up study with 1,445 donors, a wide range of bioburden levels was again observed on tissues from donors that were swab culture negative for objectionable microorganisms. Tissues from 885 (61%) of these donors had no recoverable bioburden (<2 CFU). Importantly, tissues from 560 (39%) of the donors had recoverable bioburden which ranged from 1 to >24,000 CFU. Identification of bioburden isolates showed a diversity of genera and species. In compliance with the recent revision of the American Association of Tissue Banks K2.210 Standard, the quantitative bioburden determination method was validated with a composite tissue sample that contains bone and soft tissue sections tested together in one extraction vessel. A recovery efficiency of 68% was validated and the composite sample was shown to be representative of all of the tissues recovered from a donor. The use of the composite sample in conjunction with the quantitative bioburden determination method will facilitate an accurate assessment of the numbers and types of contaminating microorganisms on allografts prior to disinfection/sterilization. This information will ensure that disinfection/sterilization processes are properly validated and the capability of the overall allograft process is understood on a donor by donor basis.

  5. Disposition of hop prenylflavonoids in human breast tissue

    PubMed Central

    Bolca, Selin; Li, Jinghu; Nikolic, Dejan; Roche, Nathalie; Blondeel, Phillip; Possemiers, Sam; De Keukeleire, Denis; Bracke, Marc; Heyerick, Arne; van Breemen, Richard B.; Depypere, Herman

    2013-01-01

    Hop-derived products may contain xanthohumol (XN), isoxanthohumol (IX), and the potent phytoestrogen 8-prenylnaringenin (8-PN). To evaluate the potential health effects of these prenylflavonoids on breast tissue, their concentration, nature of metabolites, and biodistribution were assessed and compared to 17β-estradiol (E2) exposure. In this dietary intervention study, women were randomly allocated to hop (n=11; 2.04 mg XN, 1.20 mg IX, and 0.1 mg 8-PN per supplement) or control (n=10). After a run-in of ≥4d, 3 supplements were taken daily during 5d preceding an aesthetic breast reduction. Blood and breast biopsies were analyzed using HPLC-ESI-MS/MS. Upon hop administration, XN and IX concentrations ranged between 0.72–17.65 nmol/L and 3.30–31.50 nmol/L, and between 0.26– 5.14 pmol/g and 1.16–83.67 pmol/g in hydrolyzed serum and breast tissue, respectively. 8-PN however, was only detected in samples of moderate and strong 8-PN producers (0.43–7.06 nmol/L and 0.78–4.83 pmol/g). Phase I metabolism appeared to be minor (~10%), whereas extensive glucuronidation was observed (>90%). Total prenylflavonoids showed a breast adipose/glandular tissue distribution of 38/62 and their derived E2-equivalents were negligible compared to E2 in adipose (384.6±118.8 fmol/g, P=0.009) and glandular (241.6±93.1 fmol/g, P<0.001) tissue, respectively. Consequently, low doses of prenylflavonoids are unlikely to elicit estrogenic responses in breast tissue. PMID:20486208

  6. Microhardness of human cancellous bone tissue in progressive hip osteoarthritis.

    PubMed

    Tomanik, Magdalena; Nikodem, Anna; Filipiak, Jarosław

    2016-12-01

    Bone tissue is a biological system in which the dynamic processes of, among others, bone formation or internal reconstruction will determine the spatial structure of the tissue and its mechanical properties. The appearance of a factor disturbing the balance between biological processes, e.g. a disease, will cause changes in the spatial structure of bones, thus affecting its mechanical properties. One of the bone diseases most common in an increasingly ageing population is osteoarthritis, also referred to as degenerative joint disease. It is estimated that in 2050 about 1300 million people will show symptoms of OA. The appearance of a pathological stimulus disturbs the balance of the processes of degradation and synthesis of articular cartilage, chondrocytes and the extracellular matrix, and the subchondral bone layer. As osteoarthritis progresses, study of the epiphysis reveals increasingly widespread changes of the articular surface and the internal structure of bone tissue. In this paper, the authors point out the differences in the mechanical properties of cancellous bone tissue forming the proximal epiphysis of the femoral bone during the progressive stages of OA. In order to determine microproperties of bone trabeculae, specimens from different stages of the disease (N=9) were subjected to microindentation testing, which made it possible to determine the material properties of bone tissue, such as microhardness HV and Young׳s modulus E. In addition, mechanical tests were supplemented with Raman spectroscopy, which determine the degree of bone mineralization, and measurements of structural properties based on analysis using microCT. The conducted tests were used to establish both quantitative and quantitative description of changes in the structural and mechanical properties connected with reorganization of trabeculae making up the bone in the various stages of osteoarthritis. The proposed description will supplement existing knowledge in the literature about

  7. Caspase Induction and BCL2 Inhibition in Human Adipose Tissue

    PubMed Central

    Tinahones, Francisco José; Coín Aragüez, Leticia; Murri, Mora; Oliva Olivera, Wilfredo; Mayas Torres, María Dolores; Barbarroja, Nuria; Gomez Huelgas, Ricardo; Malagón, Maria M.; El Bekay, Rajaa

    2013-01-01

    OBJECTIVE Cell death determines the onset of obesity and associated insulin resistance. Here, we analyze the relationship among obesity, adipose tissue apoptosis, and insulin signaling. RESEARCH DESIGN AND METHODS The expression levels of initiator (CASP8/9) and effector (CASP3/7) caspases as well as antiapoptotic B-cell lymphoma (BCL)2 and inflammatory markers were assessed in visceral (VAT) and subcutaneous (SAT) adipose tissue from patients with different degrees of obesity and without insulin resistance or diabetes. Adipose tissue explants from lean subjects were cultured with TNF-α or IL-6, and the expression of apoptotic and insulin signaling components was analyzed and compared with basal expression levels in morbidly obese subjects. RESULTS SAT and VAT exhibited increased CASP3/7 and CASP8/9 expression levels and decreased BCL2 expression with BMI increase. These changes were accompanied by increased inflammatory cytokine mRNA levels and macrophage infiltration markers. In obese subjects, CASP3/7 activation and BCL2 downregulation correlated with the IRS-1/2–expression levels. Expression levels of caspases, BCL2, p21, p53, IRS-1/2, GLUT4, protein tyrosine phosphatase 1B, and leukocyte antigen-related phosphatase in TNF-α– or IL-6–treated explants from lean subjects were comparable with those found in adipose tissue samples from morbidly obese subjects. These insulin component expression levels were reverted with CASP3/7 inhibition in these TNF-α– or IL-6–treated explants. CONCLUSIONS Body fat mass increase is associated with CASP3/7 and BCL2 expression in adipose tissue. Moreover, this proapoptotic state correlated with insulin signaling, suggesting its potential contribution to the development of insulin resistance. PMID:23193206

  8. Microhardness of human cancellous bone tissue in progressive hip osteoarthritis.

    PubMed

    Tomanik, Magdalena; Nikodem, Anna; Filipiak, Jarosław

    2016-12-01

    Bone tissue is a biological system in which the dynamic processes of, among others, bone formation or internal reconstruction will determine the spatial structure of the tissue and its mechanical properties. The appearance of a factor disturbing the balance between biological processes, e.g. a disease, will cause changes in the spatial structure of bones, thus affecting its mechanical properties. One of the bone diseases most common in an increasingly ageing population is osteoarthritis, also referred to as degenerative joint disease. It is estimated that in 2050 about 1300 million people will show symptoms of OA. The appearance of a pathological stimulus disturbs the balance of the processes of degradation and synthesis of articular cartilage, chondrocytes and the extracellular matrix, and the subchondral bone layer. As osteoarthritis progresses, study of the epiphysis reveals increasingly widespread changes of the articular surface and the internal structure of bone tissue. In this paper, the authors point out the differences in the mechanical properties of cancellous bone tissue forming the proximal epiphysis of the femoral bone during the progressive stages of OA. In order to determine microproperties of bone trabeculae, specimens from different stages of the disease (N=9) were subjected to microindentation testing, which made it possible to determine the material properties of bone tissue, such as microhardness HV and Young׳s modulus E. In addition, mechanical tests were supplemented with Raman spectroscopy, which determine the degree of bone mineralization, and measurements of structural properties based on analysis using microCT. The conducted tests were used to establish both quantitative and quantitative description of changes in the structural and mechanical properties connected with reorganization of trabeculae making up the bone in the various stages of osteoarthritis. The proposed description will supplement existing knowledge in the literature about

  9. Detection of the human endogenous retrovirus ERV3-encoded Env-protein in human tissues using antibody-based proteomics

    PubMed Central

    Fei, Chen; Atterby, Christina; Edqvist, Per-Henrik; Pontén, Fredrik; Zhang, Wei Wei; Larsson, Erik; Ryan, Frank P

    2014-01-01

    Objectives There is growing evidence to suggest that human endogenous retroviruses (HERVs) have contributed to human evolution, being expressed in development, normal physiology and disease. A key difficulty in the scientific evaluation of this potential viral contribution is the accurate demonstration of virally expressed protein in specific human cells and tissues. In this study, we have adopted the endogenous retrovirus, ERV3, as our test model in developing a reliable high-capacity methodology for the expression of such endogenous retrovirus-coded protein. Design Two affinity-purified polyclonal antibodies to ERV3 Env-encoded protein were generated to detect the corresponding protein expression pattern in specific human cells, tissues and organs. Participants Sampling included normal tissues from 144 individuals ranging from childhood to old age. This included more than forty different tissues and organs and some 216 different cancer tissues representing the twenty commonest forms of human cancer. Setting The Rudbeck Laboratory, Uppsala University and Uppsala University Hospital, Uppsala, Sweden. Main Outcome Measures The potential expression at likely physiological level of the ERV3Env encoded protein in a wide range of human cells, tissues and organs. Results We found that ERV3 encoded Env protein is expressed at substantive levels in placenta, testis, adrenal gland, corpus luteum, Fallopian tubes, sebaceous glands, astrocytes, bronchial epithelium and the ducts of the salivary glands. Substantive expression was also seen in a variety of epithelial cells as well as cells known to undergo fusion in inflammation and in normal physiology, including fused macrophages, myocardium and striated muscle. This contrasted strongly with the low levels expressed in other tissues types. These findings suggest that this virus plays a significant role in human physiology and may also play a possible role in disease. Conclusion This technique can now be extended to the study

  10. [Kallikrein-kinin system of blood in patients in the post-infarction period, and its reaction to physical exertion].

    PubMed

    Azimova, F I; Nekrasova, A A; Chernova, N A; Nikolaeva, L F

    1983-09-01

    Blood kallikrein-kinin function was assessed in myocardial infarction survivors. Increased blood kinin-forming activity was demonstrated, accompanied with a simultaneous rise in the system's inhibitor level and excessive response to exercise, with slower regaining of the baseline after exercise was discontinued. It is concluded that functional testing of blood kallikrein-kinin status by means of rationed exercise yields more informative data on this system in various groups of postinfarction patients and that the extent of the system's activation depends on systemic circulatory function.

  11. The Genotype-Tissue Expression (GTEx) pilot analysis: Multitissue gene regulation in humans

    PubMed Central

    2015-01-01

    Understanding the functional consequences of genetic variation, and how it affects complex human disease and quantitative traits, remains a critical challenge for biomedicine. We present an analysis of RNA sequencing data from 1641 samples across 43 tissues from 175 individuals, generated as part of the pilot phase of the Genotype-Tissue Expression (GTEx) project. We describe the landscape of gene expression across tissues, catalog thousands of tissue-specific and shared regulatory expression quantitative trait loci (eQTL) variants, describe complex network relationships, and identify signals from genome-wide association studies explained by eQTLs. These findings provide a systematic understanding of the cellular and biological consequences of human genetic variation and of the heterogeneity of such effects among a diverse set of human tissues. PMID:25954001

  12. Human polyethylene granuloma tissues inhibit bone healing in a novel xenograft animal model.

    PubMed

    Esposito, Christina I; Oliver, Rema A; Campbell, Patricia A; Yu, Yan; Walter, William L; Walter, William K; Walsh, William R

    2014-06-01

    During revision of a conventional polyethylene joint replacement, surgeons usually remove the source of osteolysis (polyethylene) but cannot always remove all of the polyethylene granuloma tissues. We developed a human/rat xenograft model to investigate the effects of polyethylene granuloma tissues on bone healing. Human osteoarthritic and periprosthetic tissues collected during primary and revision hip arthroplasty surgeries were transplanted into the distal femora of athymic nude rats. After 3 weeks in vivo, there was a significant difference in the bone volume fraction (Vf ) between empty, primary, and revision defects (p = 0.02), with a lower Vf in defects with revision granuloma tissues compared to defects with primary osteoarthritic tissues. Polyethylene granuloma tissues in trabecular bone defects inhibited bone healing. Therefore, debridement around a metal-on-polyethylene hip replacement may shorten the time it takes to achieve secondary stability around a revision hip replacement.

  13. Epithelial-connective tissue boundary in the oral part of the human soft palate

    PubMed Central

    PAULSEN, FRIEDRICH; THALE, ANDREAS

    1998-01-01

    The papillary layer of the oral part of the human soft palate was studied in 31 subjects of different age by means of histological, immunohistochemical and scanning electron microscopical methods. For scanning electron microscopy a new maceration method was introduced. Results determine epithelial thickness, height and density of connective tissue papillae and their 3-dimensional architecture inside the lining epithelium as well as the collagenous arrangement of the openings of the glandular ducts. The individual connective tissue papillae of the soft palate are compared with the connective tissue boundary on the other side of the oral cavity. The connective tissue plateaux carrying a variable number of connective tissue papillae were found to be the basic structural units of the papillary body. The function of the epithelial-connective tissue interface and the extracellular matrix arrangement in the lamina propria are discussed in order to promote the comparability of normal with pathologically altered human soft palates. PMID:9877301

  14. Quantification of plasma factor XIIa-Cl(-)-inhibitor and kallikrein-Cl(-)-inhibitor complexes in sepsis.

    PubMed

    Nuijens, J H; Huijbregts, C C; Eerenberg-Belmer, A J; Abbink, J J; Strack van Schijndel, R J; Felt-Bersma, R J; Thijs, L G; Hack, C E

    1988-12-01

    Considerable evidence indicates that activation of the contact system of intrinsic coagulation plays a role in the pathogenesis of septic shock. To monitor contact activation in patients with sepsis, we developed highly sensitive radioimmunoassays (RIAs) for factor XIIa-Cl(-)-inhibitor (Cl(-)-Inh) and kallikrein-Cl(-)-Inh complexes using a monoclonal antibody (MoAb Kok 12) that binds to a neodeterminant exposed on both complexed and cleaved Cl(-)-Inh. Plasma samples were serially collected from 48 patients admitted to the intensive care unit because of severe sepsis. Forty percent of patients on at least one occasion had increased levels of plasma factor XIIa-Cl(-)-Inh (greater than 5 x 10(-4) U/mL) and kallikrein-Cl(-)-Inh (greater than 25 x 10(-4) U/mL), that correlated at a molar ratio of approximately 1:3. Levels of factor XII antigen in plasma and both the highest as well as the levels on admission of plasma factor XIIa-Cl(-)-Inh in 23 patients with septic shock were lower than in 25 normotensive patients (P = .015: factor XII on admission; P = .04: highest factor XIIa-Cl(-)-Inh; P = .01: factor XIIa-Cl(-)-Inh on admission). No significant differences in plasma kallikrein-Cl(-)-Inh or prekallikrein antigen were found between these patients' groups. Elevated Cl(-)-Inh complex levels were measured less frequently in serial samples from patients with septic shock than in those from patients without shock (P less than .0001). Based on these results, we conclude that plasma Cl(-)-Inh complex levels during sepsis may not properly reflect the extent of contact activation.

  15. Cryopreservation, Culture, and Transplantation of Human Fetal Mesencephalic Tissue into Monkeys

    NASA Astrophysics Data System (ADS)

    Redmond, D. E.; Naftolin, F.; Collier, T. J.; Leranth, C.; Robbins, R. J.; Sladek, C. D.; Roth, R. H.; Sladek, J. R.

    1988-11-01

    Studies in animals suggest that fetal neural grafts might restore lost neurological function in Parkinson's disease. In monkeys, such grafts survive for many months and reverse signs of parkinsonism, without attendant graft rejection. The successful and reliable application of a similar transplantation procedure to human patients, however, will require neural tissue obtained from human fetal cadavers, with demonstrated cellular identity, viability, and biological safety. In this report, human fetal neural tissue was successfully grafted into the brains of monkeys. Neural tissue was collected from human fetal cadavers after 9 to 12 weeks of gestation and cryopreserved in liquid nitrogen. Viability after up to 2 months of storage was demonstrated by cell culture and by transplantation into monkeys. Cryopreservation and storage of human fetal neural tissue would allow formation of a tissue bank. The stored cells could then be specifically tested to assure their cellular identity, viability, and bacteriological and virological safety before clinical use. The capacity to collect and maintain viable human fetal neural tissue would also facilitate research efforts to understand the development and function of the human brain and provide opportunities to study neurological diseases.

  16. Intra-Operatively Obtained Human Tissue: Protocols and Techniques for the Study of Neural Stem Cells

    PubMed Central

    Chaichana, Kaisorn; Guerrero-Cazares, Hugo; Capilla-Gonzalez, Vivian; Zamora-Berridi, Grettel; Achanta, Praganthi; Gonzalez-Perez, Oscar; Jallo, George I.; Garcia-Verdugo, Jose Manuel; Quiñones-Hinojosa, Alfredo

    2009-01-01

    The discoveries of neural (NSCs) and brain tumor stem cells (BTSCs) in the adult human brain and in brain tumors, respectively, have led to a new era in neuroscience research. These cells represent novel approaches to studying normal phenomena such as memory and learning, as well as pathological conditions such as Parkinson’s disease, stroke, and brain tumors. This new paradigm stresses the importance of understanding how these cells behave in vitro and in vivo. It also stresses the need to use human-derived tissue to study human disease because animal models may not necessarily accurately replicate the processes that occur in humans. An important, but often underused, source of human tissue and, consequently, both NSCs and BTSCs, is the operating room. This study describes in detail both current and newly developed laboratory techniques, which in our experience are used to process and study human NSCs and BTSCs from tissue obtained directly from the operating room. PMID:19427538

  17. Gene Transfection of Human Turbinate Mesenchymal Stromal Cells Derived from Human Inferior Turbinate Tissues

    PubMed Central

    Kwon, Jin Seon; Park, Seung Hun; Baek, Ji Hye; Dung, Truong Minh; Kim, Sung Won; Min, Byoung Hyun; Kim, Jae Ho; Kim, Moon Suk

    2016-01-01

    Human turbinate mesenchymal stromal cells (hTMSCs) are novel stem cells derived from nasal inferior turbinate tissues. They are easy to isolate from the donated tissue after turbinectomy or conchotomy. In this study, we applied hTMSCs to a nonviral gene delivery system using polyethyleneimine (PEI) as a gene carrier; furthermore, the cytotoxicity and transfection efficiency of hTMSCs were evaluated to confirm their potential as resources in gene therapy. DNA-PEI nanoparticles (NPs) were generated by adding the PEI solution to DNA and were characterized by a gel electrophoresis and by measuring particle size and surface charge of NPs. The hTMSCs were treated with DNA-PEI NPs for 4 h, and toxicity of NPs to hTMSCs and gene transfection efficiency were monitored using MTT assay, fluorescence images, and flow cytometry after 24 h and 48 h. At a high negative-to-positive charge ratio, DNA-PEI NPs treatment led to cytotoxicity of hTMSCs, but the transfection efficiency of DNA was increased due to the electrostatic effect between the NPs and the membranes of hTMSCs. Importantly, the results of this research verified that PEI could deliver DNA into hTMSCs with high efficiency, suggesting that hTMSCs could be considered as untapped resources for applications in gene therapy. PMID:26783402

  18. The kinin-kallikrein system: physiological roles, pathophysiology and its relationship to cancer biomarkers.

    PubMed

    Kashuba, Elena; Bailey, James; Allsup, David; Cawkwell, Lynn

    2013-06-01

    The kinin-kallikrein system (KKS) is an endogenous multiprotein cascade, the activation of which leads to triggering of the intrinsic coagulation pathway and enzymatic hydrolysis of kininogens with the consequent release of bradykinin-related peptides. This system plays a crucial role in inflammation, vasodilation, smooth muscle contraction, cardioprotection, vascular permeability, blood pressure control, coagulation and pain. In this review, we will outline the physiology and pathophysiology of the KKS and also highlight the association of this system with carcinogenesis and cancer progression. PMID:23672534

  19. Analysis of variance in spectroscopic imaging data from human tissues.

    PubMed

    Kwak, Jin Tae; Reddy, Rohith; Sinha, Saurabh; Bhargava, Rohit

    2012-01-17

    The analysis of cell types and disease using Fourier transform infrared (FT-IR) spectroscopic imaging is promising. The approach lacks an appreciation of the limits of performance for the technology, however, which limits both researcher efforts in improving the approach and acceptance by practitioners. One factor limiting performance is the variance in data arising from biological diversity, measurement noise or from other sources. Here we identify the sources of variation by first employing a high throughout sampling platform of tissue microarrays (TMAs) to record a sufficiently large and diverse set data. Next, a comprehensive set of analysis of variance (ANOVA) models is employed to analyze the data. Estimating the portions of explained variation, we quantify the primary sources of variation, find the most discriminating spectral metrics, and recognize the aspects of the technology to improve. The study provides a framework for the development of protocols for clinical translation and provides guidelines to design statistically valid studies in the spectroscopic analysis of tissue.

  20. Linking microstructure and nanochemistry in human dental tissues.

    PubMed

    Srot, Vesna; Bussmann, Birgit; Salzberger, Ute; Koch, Christoph T; van Aken, Peter A

    2012-06-01

    Mineralized dental tissues and dental pulp were characterized using advanced analytical transmission electron microscopy (TEM) methods. Quantitative X-ray energy dispersive spectroscopy was employed to determine the Ca/P and Mg/P concentration ratios. Significantly lower Ca/P concentration ratios were measured in peritubular dentine compared to intertubular dentine, which is accompanied by higher and variable Mg/P concentration ratios. There is strong evidence that magnesium is partially substituting calcium in the hydroxyapatite structure. Electron energy-loss near-edge structures (ELNES) of C-K and O-K from enamel and dentine are noticeably different. We observe a strong influence of beam damage on mineralized dental tissues and dental pulp, causing changes of the composition and consequently also differences in the ELNES. In this article, the importance of TEM sample preparation and specimen damage through electron irradiation is demonstrated.

  1. Production of Tissue Microarrays, Immunohistochemistry Staining and Digitalization Within the Human Protein Atlas

    PubMed Central

    Kampf, Caroline; Olsson, IngMarie; Ryberg, Urban; Sjöstedt, Evelina; Pontén, Fredrik

    2012-01-01

    The tissue microarray (TMA) technology provides the means for high-throughput analysis of multiple tissues and cells. The technique is used within the Human Protein Atlas project for global analysis of protein expression patterns in normal human tissues, cancer and cell lines. Here we present the assembly of 1 mm cores, retrieved from microscopically selected representative tissues, into a single recipient TMA block. The number and size of cores in a TMA block can be varied from approximately forty 2 mm cores to hundreds of 0.6 mm cores. The advantage of using TMA technology is that large amount of data can rapidly be obtained using a single immunostaining protocol to avoid experimental variability. Importantly, only limited amount of scarce tissue is needed, which allows for the analysis of large patient cohorts 1 2. Approximately 250 consecutive sections (4 μm thick) can be cut from a TMA block and used for immunohistochemical staining to determine specific protein expression patterns for 250 different antibodies. In the Human Protein Atlas project, antibodies are generated towards all human proteins and used to acquire corresponding protein profiles in both normal human tissues from 144 individuals and cancer tissues from 216 different patients, representing the 20 most common forms of human cancer. Immunohistochemically stained TMA sections on glass slides are scanned to create high-resolution images from which pathologists can interpret and annotate the outcome of immunohistochemistry. Images together with corresponding pathology-based annotation data are made publically available for the research community through the Human Protein Atlas portal (www.proteinatlas.org) (Figure 1) 3 4. The Human Protein Atlas provides a map showing the distribution and relative abundance of proteins in the human body. The current version contains over 11 million images with protein expression data for 12.238 unique proteins, corresponding to more than 61% of all proteins

  2. Uncovering of melanin fluorescence in human skin tissue

    NASA Astrophysics Data System (ADS)

    Scholz, Matthias; Stankovic, Goran; Seewald, Gunter; Leupold, Dieter

    2007-07-01

    Due to its extremely low fluorescence quantum yield, in the conventionally (one-photon) excited autofluorescence of skin tissue, melanin fluorescence is masked by several other endogenous and possibly also exogenous fluorophores (e.g. NADH, FAD, Porphyrins). A first step to enhance the melanin contribution had been realized by two-photon fs-pulse excitation in the red/near IR, based on the fact that melanin can be excited by stepwise two-photon absorption, whereas all other fluorophores in this spectral region allow only simultaneous two-photon excitation. Now, the next and decisive step has been realized: Using an extremely sensitive detection system, for the first time twophoton fluorescence of skin tissue excited with pulses in the ns-range could be measured. The motivation for this step was based on the fact that the population density of the fluorescent level resulting from a stepwise excitation has a different dependence of the pulse duration than that from a simultaneous excitation (Δt2 vs. Δt). Due to this strong discrimination between the fluorophores, practically pure melanin fluorescence can be obtained. Examples for in-vivo, ex-vivo as well as paraffin embedded skin tissue will be shown. The content of information with respect to early diagnosis of skin deseases will be discussed.

  3. Noninvasive metabolic imaging of engineered 3D human adipose tissue in a perfusion bioreactor.

    PubMed

    Ward, Andrew; Quinn, Kyle P; Bellas, Evangelia; Georgakoudi, Irene; Kaplan, David L

    2013-01-01

    The efficacy and economy of most in vitro human models used in research is limited by the lack of a physiologically-relevant three-dimensional perfused environment and the inability to noninvasively quantify the structural and biochemical characteristics of the tissue. The goal of this project was to develop a perfusion bioreactor system compatible with two-photon imaging to noninvasively assess tissue engineered human adipose tissue structure and function in vitro. Three-dimensional (3D) vascularized human adipose tissues were engineered in vitro, before being introduced to a perfusion environment and tracked over time by automated quantification of endogenous markers of metabolism using two-photon excited fluorescence (TPEF). Depth-resolved image stacks were analyzed for redox ratio metabolic profiling and compared to prior analyses performed on 3D engineered adipose tissue in static culture. Traditional assessments with H&E staining were used to qualitatively measure extracellular matrix generation and cell density with respect to location within the tissue. The distribution of cells within the tissue and average cellular redox ratios were different between static and perfusion cultures, while the trends of decreased redox ratio and increased cellular proliferation with time in both static and perfusion cultures were similar. These results establish a basis for noninvasive optical tracking of tissue structure and function in vitro, which can be applied to future studies to assess tissue development or drug toxicity screening and disease progression.

  4. Phantom and animal tissues for modelling the electrical properties of human liver.

    PubMed

    Stauffer, P R; Rossetto, F; Prakash, M; Neuman, D G; Lee, T

    2003-01-01

    The dielectric properties of human liver were characterized over the frequency range of 0.3-3 GHz for freshly excised tissue samples of primary hepatocellular carcinoma, metastatic colorectal carcinoma, and normal liver tissues resected from the tumour margin. On average, the dielectric constant (epsilon(r)) of freshly excised human liver tumour was 12% higher than that of surrounding normal liver, and the electrical conductivity (sigma) of tumour was 24% higher. In order to establish suitable tissue models for human liver, the electrical properties were compared to measurements of homogenous phantom mixtures, in vitro bovine liver, and in vivo canine and porcine liver tissues. The data demonstrate that there are several animal tissues that can be used to model the average dielectric properties of human liver reasonably accurately, and use of the most readily available bovine liver appears well-justified, even when stored for up to 10 days in a refrigerator. Additionally, the dielectric properties of in vitro liver remained stable over a large temperature range, with sigma rising only 1.1%/ degrees C in porcine liver (15-37 degrees C) and 2.0%/ degrees C in bovine liver (10-90 degrees C), and epsilon(r) decreasing < or =0.2%/ degrees C in both tissues. This effort identifies homogeneous solid and liquid phantom models and several heterogeneous in vitro tissues that adequately model the dielectric properties of human liver tumours for use in quantitative studies of microwave power deposition in liver.

  5. FTIR microscopic comparative study on normal, premalignant, and malignant tissues of human intenstine

    NASA Astrophysics Data System (ADS)

    Mordechai, Shaul; Argov, Shmuel; Salman, Ahmad O.; Cohen, Beny; Ramesh, Jagannathan; Erukhimovitch, Vitaly; Goldstein, Jed; Sinelnikov, Igor

    2000-07-01

    Fourier-Transform Infrared Spectroscopy (FTIR) employs a unique approach to optical diagnosis of tissue pathology based on the characteristic molecular vibrational spectra of the tissue. The architectural changes in the cellular and sub-cellular levels developing in abnormal tissue, including a majority of cancer forms, manifest themselves in different optical signatures, which can be detected in infrared spectroscopy. The biological systems we have studied include normal, premalignant (polyp) and malignant human colonic tissues from three patients. Our method is based on microscopic infrared study (FTIR-microscopy) of thin tissue specimens and a direct comparison with normal histopathological analysis, which serves as a `gold' reference. The normal intestine tissue has a stronger absorption than polyp and cancerous types over a wide region in all three cases. The detailed analysis showed that there is a significant decrease in total phosphate and creatine contents for polyp and cancerous tissue types in comparison to the controls.

  6. Correlating optical coherence elastography based strain measurements with collagen content of the human ovarian tissue.

    PubMed

    Nandy, Sreyankar; Salehi, Hassan S; Wang, Tianheng; Wang, Xiaohong; Sanders, Melinda; Kueck, Angela; Brewer, Molly; Zhu, Quing

    2015-10-01

    In this manuscript, the initial feasibility of a catheter based phase stabilized swept source optical coherence tomography (OCT) system was studied for characterization of the strain inside different human ovarian tissue groups. The ovarian tissue samples were periodically compressed with 500 Hz square wave signal along the axial direction between the surface of an unfocused transducer and a glass cover slide. The displacement and corresponding strain were calculated during loading from different locations for each tissue sample. A total of 27 ex vivo ovaries from 16 patients were investigated. Statistically significant difference (p < 0.001) was observed between the average displacement and strain of the normal and malignant tissue groups. A sensitivity of 93.2% and a specificity of 83% were achieved using 25 microstrain (με) as the threshold. The collagen content of the tissues was quantified from the Sirius Red stained histological sections. The average collagen area fraction (CAF) obtained from the tissue groups were found to have a strong negative correlation (R = -0.75, p < 0.0001) with the amount of strain inside the tissue. This indicates much softer and degenerated tissue structure for the malignant ovaries as compared to the dense, collagen rich structure of the normal ovarian tissue. The initial results indicate that the swept source OCT system can be useful for estimating the elasticity of the human ovarian tissue. PMID:26504631

  7. Raman spectroscopic analysis of human skin tissue sections ex-vivo: evaluation of the effects of tissue processing and dewaxing

    NASA Astrophysics Data System (ADS)

    Ali, Syed M.; Bonnier, Franck; Tfayli, Ali; Lambkin, Helen; Flynn, Kathleen; McDonagh, Vincent; Healy, Claragh; Clive Lee, T.; Lyng, Fiona M.; Byrne, Hugh J.

    2013-06-01

    Raman spectroscopy coupled with K-means clustering analysis (KMCA) is employed to elucidate the biochemical structure of human skin tissue sections and the effects of tissue processing. Both hand and thigh sections of human cadavers were analyzed in their unprocessed and formalin-fixed, paraffin-processed (FFPP), and subsequently dewaxed forms. In unprocessed sections, KMCA reveals clear differentiation of the stratum corneum (SC), intermediate underlying epithelium, and dermal layers for sections from both anatomical sites. The SC is seen to be relatively rich in lipidic content; the spectrum of the subjacent layers is strongly influenced by the presence of melanin, while that of the dermis is dominated by the characteristics of collagen. For a given anatomical site, little difference in layer structure and biochemistry is observed between samples from different cadavers. However, the hand and thigh sections are consistently differentiated for all cadavers, largely based on lipidic profiles. In dewaxed FFPP samples, while the SC, intermediate, and dermal layers are clearly differentiated by KMCA of Raman maps of tissue sections, the lipidic contributions to the spectra are significantly reduced, with the result that respective skin layers from different anatomical sites become indistinguishable. While efficient at removing the fixing wax, the tissue processing also efficiently removes the structurally similar lipidic components of the skin layers. In studies of dermatological processes in which lipids play an important role, such as wound healing, dewaxed samples are therefore not appropriate. Removal of the lipids does however accentuate the spectral features of the cellular and protein components, which may be more appropriate for retrospective analysis of disease progression and biochemical analysis using tissue banks.

  8. Combinations of parabens at concentrations measured in human breast tissue can increase proliferation of MCF-7 human breast cancer cells.

    PubMed

    Charles, Amelia K; Darbre, Philippa D

    2013-05-01

    The alkyl esters of p-hydroxybenzoic acid (parabens), which are used as preservatives in consumer products, possess oestrogenic activity and have been measured in human breast tissue. This has raised concerns for a potential involvement in the development of human breast cancer. In this paper, we have investigated the extent to which proliferation of MCF-7 human breast cancer cells can be increased by exposure to the five parabens either alone or in combination at concentrations as recently measured in 160 human breast tissue samples. Determination of no-observed-effect concentrations (NOEC), lowest-observed-effect concentrations (LOEC), EC50 and EC100 values for stimulation of proliferation of MCF-7 cells by five parabens revealed that 43/160 (27%) of the human breast tissue samples contained at least one paraben at a concentration ≥ LOEC and 64/160 (40%) > NOEC. Proliferation of MCF-7 cells could be increased by combining all five parabens at concentrations down to the 50(th) percentile (median) values measured in the tissues. For the 22 tissue samples taken at the site of ER + PR + primary cancers, 12 contained a sufficient concentration of one or more paraben to stimulate proliferation of MCF-7 cells. This demonstrates that parabens, either alone or in combination, are present in human breast tissue at concentrations sufficient to stimulate the proliferation of MCF-7 cells in vitro, and that functional consequences of the presence of paraben in human breast tissue should be assessed on the basis of all five parabens and not single parabens individually.

  9. Nonlinear optics for the study of human scar tissue

    NASA Astrophysics Data System (ADS)

    Ferro, D. P.; Vieira-Damiani, G.; Adam, R. L.; Cesar, C. L.; Metze, Konradin

    2012-03-01

    Collagen fibers are an essential component of the dynamic process of scarring, which accompanies various diseases. Scar tissue may reveal different morphologic expressions, such as hypertrophic scars or keloids. Collagen fibers can be visualized by fluorescent light when stained with eosin. Second Harmonic Generation (SHG) creates a non linear signal that occurs only in molecules without inversion symmetry and is particularly strong in the collagen fibers arranged in triple helices. The aim of this study was to describe the methodology for the analysis of the density and texture of collagen in keloids, hypertrophic scars and conventional scars. Samples were examined in the National Institute of Science and Technology on Photonics Applied to Cell Biology (INFABIC) at the State University of Campinas. The images were acquired in a multiphoton microscopy LSM 780-NLO Zeiss 40X. Both signals, two-photon fluorescence (TPEF) and SHG, were excited by a Mai-Tai Ti:Sapphire laser at 940 nm. We used a LP490/SP485 NDD filter for SHG, and a BP565-610 NDD filter for fluorescence In each case, ten images were acquired serially (512×512 μm) in Z-stack and joined together to one patchwork-image . Image analysis was performed by a gliding-box-system with in-house made software. Keloids, hypertrophic scars and normal scar tissue show different collagen architecture. Inside an individual case differences of the scar process may be found between central and peripheral parts. In summary, the use of nonlinear optics is a helpful tool for the study of scars tissue.

  10. Mineral density volume gradients in normal and diseased human tissues.

    PubMed

    Djomehri, Sabra I; Candell, Susan; Case, Thomas; Browning, Alyssa; Marshall, Grayson W; Yun, Wenbing; Lau, S H; Webb, Samuel; Ho, Sunita P

    2015-01-01

    Clinical computed tomography provides a single mineral density (MD) value for heterogeneous calcified tissues containing early and late stage pathologic formations. The novel aspect of this study is that, it extends current quantitative methods of mapping mineral density gradients to three dimensions, discretizes early and late mineralized stages, identifies elemental distribution in discretized volumes, and correlates measured MD with respective calcium (Ca) to phosphorus (P) and Ca to zinc (Zn) elemental ratios. To accomplish this, MD variations identified using polychromatic radiation from a high resolution micro-computed tomography (micro-CT) benchtop unit were correlated with elemental mapping obtained from a microprobe X-ray fluorescence (XRF) using synchrotron monochromatic radiation. Digital segmentation of tomograms from normal and diseased tissues (N=5 per group; 40-60 year old males) contained significant mineral density variations (enamel: 2820-3095 mg/cc, bone: 570-1415 mg/cc, cementum: 1240-1340 mg/cc, dentin: 1480-1590 mg/cc, cementum affected by periodontitis: 1100-1220 mg/cc, hypomineralized carious dentin: 345-1450 mg/cc, hypermineralized carious dentin: 1815-2740 mg/cc, and dental calculus: 1290-1770 mg/cc). A plausible linear correlation between segmented MD volumes and elemental ratios within these volumes was established, and Ca/P ratios for dentin (1.49), hypomineralized dentin (0.32-0.46), cementum (1.51), and bone (1.68) were observed. Furthermore, varying Ca/Zn ratios were distinguished in adapted compared to normal tissues, such as in bone (855-2765) and in cementum (595-990), highlighting Zn as an influential element in prompting observed adaptive properties. Hence, results provide insights on mineral density gradients with elemental concentrations and elemental footprints that in turn could aid in elucidating mechanistic processes for pathologic formations. PMID:25856386

  11. Mineral density volume gradients in normal and diseased human tissues

    DOE PAGES

    Djomehri, Sabra I.; Candell, Susan; Case, Thomas; Browning, Alyssa; Marshall, Grayson W.; Yun, Wenbing; Lau, S. H.; Webb, Samuel; Ho, Sunita P.; Aikawa, Elena

    2015-04-09

    Clinical computed tomography provides a single mineral density (MD) value for heterogeneous calcified tissues containing early and late stage pathologic formations. The novel aspect of this study is that, it extends current quantitative methods of mapping mineral density gradients to three dimensions, discretizes early and late mineralized stages, identifies elemental distribution in discretized volumes, and correlates measured MD with respective calcium (Ca) to phosphorus (P) and Ca to zinc (Zn) elemental ratios. To accomplish this, MD variations identified using polychromatic radiation from a high resolution micro-computed tomography (micro-CT) benchtop unit were correlated with elemental mapping obtained from a microprobe X-raymore » fluorescence (XRF) using synchrotron monochromatic radiation. Digital segmentation of tomograms from normal and diseased tissues (N=5 per group; 40-60 year old males) contained significant mineral density variations (enamel: 2820-3095mg/cc, bone: 570-1415mg/cc, cementum: 1240-1340mg/cc, dentin: 1480-1590mg/cc, cementum affected by periodontitis: 1100-1220mg/cc, hypomineralized carious dentin: 345-1450mg/cc, hypermineralized carious dentin: 1815-2740mg/cc, and dental calculus: 1290-1770mg/cc). A plausible linear correlation between segmented MD volumes and elemental ratios within these volumes was established, and Ca/P ratios for dentin (1.49), hypomineralized dentin (0.32-0.46), cementum (1.51), and bone (1.68) were observed. Furthermore, varying Ca/Zn ratios were distinguished in adapted compared to normal tissues, such as in bone (855-2765) and in cementum (595-990), highlighting Zn as an influential element in prompting observed adaptive properties. Hence, results provide insights on mineral density gradients with elemental concentrations and elemental footprints that in turn could aid in elucidating mechanistic processes for pathologic formations.« less

  12. Mineral Density Volume Gradients in Normal and Diseased Human Tissues

    PubMed Central

    Djomehri, Sabra I.; Candell, Susan; Case, Thomas; Browning, Alyssa; Marshall, Grayson W.; Yun, Wenbing; Lau, S. H.; Webb, Samuel; Ho, Sunita P.

    2015-01-01

    Clinical computed tomography provides a single mineral density (MD) value for heterogeneous calcified tissues containing early and late stage pathologic formations. The novel aspect of this study is that, it extends current quantitative methods of mapping mineral density gradients to three dimensions, discretizes early and late mineralized stages, identifies elemental distribution in discretized volumes, and correlates measured MD with respective calcium (Ca) to phosphorus (P) and Ca to zinc (Zn) elemental ratios. To accomplish this, MD variations identified using polychromatic radiation from a high resolution micro-computed tomography (micro-CT) benchtop unit were correlated with elemental mapping obtained from a microprobe X-ray fluorescence (XRF) using synchrotron monochromatic radiation. Digital segmentation of tomograms from normal and diseased tissues (N=5 per group; 40-60 year old males) contained significant mineral density variations (enamel: 2820-3095mg/cc, bone: 570-1415mg/cc, cementum: 1240-1340mg/cc, dentin: 1480-1590mg/cc, cementum affected by periodontitis: 1100-1220mg/cc, hypomineralized carious dentin: 345-1450mg/cc, hypermineralized carious dentin: 1815-2740mg/cc, and dental calculus: 1290-1770mg/cc). A plausible linear correlation between segmented MD volumes and elemental ratios within these volumes was established, and Ca/P ratios for dentin (1.49), hypomineralized dentin (0.32-0.46), cementum (1.51), and bone (1.68) were observed. Furthermore, varying Ca/Zn ratios were distinguished in adapted compared to normal tissues, such as in bone (855-2765) and in cementum (595-990), highlighting Zn as an influential element in prompting observed adaptive properties. Hence, results provide insights on mineral density gradients with elemental concentrations and elemental footprints that in turn could aid in elucidating mechanistic processes for pathologic formations. PMID:25856386

  13. Mineral density volume gradients in normal and diseased human tissues

    SciTech Connect

    Djomehri, Sabra I.; Candell, Susan; Case, Thomas; Browning, Alyssa; Marshall, Grayson W.; Yun, Wenbing; Lau, S. H.; Webb, Samuel; Ho, Sunita P.; Aikawa, Elena

    2015-04-09

    Clinical computed tomography provides a single mineral density (MD) value for heterogeneous calcified tissues containing early and late stage pathologic formations. The novel aspect of this study is that, it extends current quantitative methods of mapping mineral density gradients to three dimensions, discretizes early and late mineralized stages, identifies elemental distribution in discretized volumes, and correlates measured MD with respective calcium (Ca) to phosphorus (P) and Ca to zinc (Zn) elemental ratios. To accomplish this, MD variations identified using polychromatic radiation from a high resolution micro-computed tomography (micro-CT) benchtop unit were correlated with elemental mapping obtained from a microprobe X-ray fluorescence (XRF) using synchrotron monochromatic radiation. Digital segmentation of tomograms from normal and diseased tissues (N=5 per group; 40-60 year old males) contained significant mineral density variations (enamel: 2820-3095mg/cc, bone: 570-1415mg/cc, cementum: 1240-1340mg/cc, dentin: 1480-1590mg/cc, cementum affected by periodontitis: 1100-1220mg/cc, hypomineralized carious dentin: 345-1450mg/cc, hypermineralized carious dentin: 1815-2740mg/cc, and dental calculus: 1290-1770mg/cc). A plausible linear correlation between segmented MD volumes and elemental ratios within these volumes was established, and Ca/P ratios for dentin (1.49), hypomineralized dentin (0.32-0.46), cementum (1.51), and bone (1.68) were observed. Furthermore, varying Ca/Zn ratios were distinguished in adapted compared to normal tissues, such as in bone (855-2765) and in cementum (595-990), highlighting Zn as an influential element in prompting observed adaptive properties. Hence, results provide insights on mineral density gradients with elemental concentrations and elemental footprints that in turn could aid in elucidating mechanistic processes for pathologic formations.

  14. The influence of tissue procurement procedures on RNA integrity, gene expression, and morphology in porcine and human liver tissue.

    PubMed

    Kap, Marcel; Sieuwerts, Anieta M; Kubista, Mikael; Oomen, Monique; Arshad, Shazia; Riegman, Peter

    2015-06-01

    The advent of molecular characterization of tissues has brought an increasing emphasis on the quality of biospecimens, starting with the tissue procurement process. RNA levels are particularly affected by factors in the collection process, but the influence of different pre-analytical factors is not well understood. Here we present the influence of tissue specimen size, as well as the transport and freezing protocols, on RNA quality. Large, medium, and smaller porcine liver samples were stored either dry, on moist gauze, or in salt solution for various times, and then frozen in either liquid nitrogen or in pre-cooled isopentane. Large and small human liver samples were frozen in pre-cooled isopentane either immediately or after one hour at room temperature. The small samples were stored dry, on moist gauze, or in salt solution. RNA was isolated and RIN values were measured. The RNA for six standard reference genes from human liver was analyzed by RT-qPCR, and tissue morphology was assessed for artifacts of freezing. Experiments using porcine liver samples showed that RNA derived from smaller samples was more degraded after one hour of cold ischemia, and that cooled transport is preferable. Human liver samples showed significant RNA degradation after 1 h of cold ischemia, which was more pronounced in smaller samples. RNA integrity was not significantly influenced by the transport or freezing method, but changes in gene expression were observed in samples either transported on gauze or in salt solution. Based on observations in liver samples, smaller samples are more subject to gene expression variability introduced by post-excision sample handling than are larger samples. Small biopsies should be transported on ice and snap frozen as soon as possible after acquisition from the patient. PMID:26035010

  15. The human tissue transglutaminase gene maps on chromosome 20q12 by in situ fluorescence hybridization

    SciTech Connect

    Gentile, V.; Davies, P.J.A. ); Baldini, A. )

    1994-03-15

    A cDNA encoding for the human tissue transglutaminase gene has been used to identify the chromosomal localization of the corresponding structural gene. The precise chromosomal and subregional localizations have been established by using in situ fluorescence mapping with a recombinant [lambda]-Zap phage containing the full cDNA coding sequence. The study showed that the human tissue transglutaminase gene is localized on chromosome 20 and, more precisely, within the band 20q12. To date, this is the third member of the transglutaminase gene family to be mapped. Human factor XIIIa (plasma transglutaminase), human keratinocyte transglutaminase (type I), and human tissue transglutaminase (type II) genes, although codifying for homologous enzymes, are localized on three different chromosomes. 16 refs., 1 fig.

  16. Communication channel modeling of human forearm with muscle fiber tissue characteristics.

    PubMed

    Zhang, Shuang; Pun, Sio Hang; Mak, Peng Un; Qin, Yu-Ping; Liu, Yi-He; Vai, Mang I

    2016-09-14

    Human-Body Communication (HBC) is a wireless communication method using the human body tissue as a transmission medium for signals. This paper on the basis of human muscle fiber tissues' characteristics, it is first proposed to establish the analytical model of galvanic coupling human-body communication channel. In this model, the parallel and the transverse electrical characteristics of muscular tissue are fully considered, and the model accurately presents the transmission mechanism of galvanic coupling human-body communication signals in the channel. At last, through compare with the experimental results and calculation results, the maximum error of the model is 22.4% and the average error is 14.2% within the frequency range.

  17. From cell lines to tissues: extrapolation of transcriptional effects to human tissues (SOT)

    EPA Science Inventory

    A new suite of assays in the metabolically-competent, human hepatocyte-derived HepaRG cell line has been added to the ToxCast screening suite. For 1066 chemicals we have evaluated the chemical treatment-induced changes in expression for a diverse set of 93 genes representative of...

  18. Engineering human tissue and regulation: confronting biology and law to bridge the gaps.

    PubMed

    Longley, D; Lawford, P

    2001-01-01

    There are a number of difficulties confronting the regulation of human tissue engineered products, from the scientific, ethical and legal perspectives. Many of these issues are international in scope and any responses must consider the global implications of marketing and monitoring these products. The article argues that as tissue engineered products become more available regulatory authorities should not be pressured into adopting possibly inappropriate measures, but must consider all the factors relevant to human health, including the need for innovative regulatory mechanisms as well as innovative tissue products themselves.

  19. Organotypic culture of human bone marrow adipose tissue.

    PubMed

    Uchihashi, Kazuyoshi; Aoki, Shigehisa; Shigematsu, Masamori; Kamochi, Noriyuki; Sonoda, Emiko; Soejima, Hidenobu; Fukudome, Kenji; Sugihara, Hajime; Hotokebuchi, Takao; Toda, Shuji

    2010-04-01

    The precise role of bone marrow adipose tissue (BMAT) in the marrow remains unknown. The purpose of the present study was therefore to describe a novel method for studying BMAT using 3-D collagen gel culture of BMAT fragments, immunohistochemistry, ELISA and real-time reverse transcription-polymerase chain reaction. Mature adipocytes and CD45+ leukocytes were retained for >3 weeks. Bone marrow stromal cells (BMSC) including a small number of lipid-laden preadipocytes and CD44+/CD105+ mesenchymal stem cell (MSC)-like cells, developed from BMAT. Dexamethasone (10 micromol/L), but not insulin (20 mU/mL), significantly increased the number of preadipocytes. Dexamethasone and insulin also promoted leptin production and gene expression in BMAT. Adiponectin production by BMAT was <0.8 ng/mL under all culture conditions. Dexamethasone promoted adiponectin gene expression, while insulin inhibited it. This finding suggests that dexamethasone, but not insulin, may serve as a powerful adipogenic factor for BMAT, in which adiponectin protein secretion is normally very low, and that BMAT may exhibit a different phenotype from that of the visceral and subcutaneous adipose tissues. BMAT-osteoblast interactions were also examined, and it was found that osteoblasts inhibited the development of BMSC and reduced leptin production, while BMAT inhibited the growth and differentiation of osteoblasts. The present novel method proved to be useful for the study of BMAT biology.

  20. [Age changes of the connective tissue structures of human penis].

    PubMed

    Klimachev, V V; Neĭmark, A I; Gerval'd, V Ia; Bobrov, I P; Avdalian, A M; Muzalevskaia, N I; Gerval'd, I V; Aliev, R T; Cherdantseva, T M

    2011-01-01

    This investigation was aimed at the study of age changes of penis connective tissue structures. Tissue fragments of penis were obtained from 20 cadavers of men at the age of 20-38 years in group I, and from 20 cadavers of men at the age of 41-59 years in group II. The criteria for the exclusion of material from the research were arterial hypertension, diabetes mellitus, atherosclerosis of internal iliac arteries, Peyronie's disease, and anomalies of genital organ development. It was shown that in the cavernous body of penis, aging was associated with the increased amount and thickening of collagen and argyrophilic fibers, decreased content and thinning of elastic fibers, and the reduced amount of smooth muscle cells (SMC). The average area of fibroblast and SMC nucleolus was not different in both groups studied. The average area of endotheliocyte nucleolus was equal to 1.9+/-0.9 microm2 in group II, being lower than that one in group I, in which this index was equal to 2.1+/-0.9 microm2. No differences in the content of type III and IV collagen were found between the study groups. Age-associated decrease in the average area of endothelial cell nucleolus in the cavernous bodies may reflect the reduction of the activity of these cells and may indicate the development of endothelial dysfunction, which is one of the most important steps in the morphogenesis of age-related male erectile dysfunction.

  1. Atlas of tissue renin-angiotensin-aldosterone system in human: A transcriptomic meta-analysis

    PubMed Central

    Nehme, Ali; Cerutti, Catherine; Dhaouadi, Nedra; Gustin, Marie Paule; Courand, Pierre-Yves; Zibara, Kazem; Bricca, Giampiero

    2015-01-01

    Tissue renin-angiotensin-aldosterone system (RAAS) has attracted much attention because of its physiological and pharmacological implications; however, a clear definition of tissue RAAS is still missing. We aimed to establish a preliminary atlas for the organization of RAAS across 23 different normal human tissues. A set of 37 genes encoding classical and novel RAAS participants including gluco- and mineralo-corticoids were defined as extended RAAS (extRAAS) system. Microarray data sets containing more than 10 normal tissues were downloaded from the GEO database. R software was used to extract expression levels and construct dendrograms of extRAAS genes within each data set. Tissue co-expression modules were then extracted from reproducible gene clusters across data sets. An atlas of the maps of tissue-specific organization of extRAAS was constructed from gene expression and coordination data. Our analysis included 143 data sets containing 4933 samples representing 23 different tissues. Expression data provided an insight on the favored pathways in a given tissue. Gene coordination indicated the existence of tissue-specific modules organized or not around conserved core groups of transcripts. The atlas of tissue-specific organization of extRAAS will help better understand tissue-specific effects of RAAS. This will provide a frame for developing more effective and selective pharmaceuticals targeting extRAAS. PMID:25992767

  2. Tissue Metabonomic Phenotyping for Diagnosis and Prognosis of Human Colorectal Cancer

    PubMed Central

    Tian, Yuan; Xu, Tangpeng; Huang, Jia; Zhang, Limin; Xu, Shan; Xiong, Bin; Wang, Yulan; Tang, Huiru

    2016-01-01

    Colorectal cancer (CRC) is one of the leading causes of cancer-related death worldwide and prognosis based on the conventional histological grading method for CRC remains poor. To better the situation, we analyzed the metabonomic signatures of 50 human CRC tissues and their adjacent non-involved tissues (ANIT) using high-resolution magic-angle spinning (HRMAS) 1H NMR spectroscopy together with the fatty acid compositions of these tissues using GC-FID/MS. We showed that tissue metabolic phenotypes not only discriminated CRC tissues from ANIT, but also distinguished low-grade tumor tissues (stages I-II) from the high-grade ones (stages III-IV) with high sensitivity and specificity in both cases. Metabonomic phenotypes of CRC tissues differed significantly from that of ANIT in energy metabolism, membrane biosynthesis and degradations, osmotic regulations together with the metabolism of proteins and nucleotides. Amongst all CRC tissues, the stage I tumors exhibited largest differentiations from ANIT. The combination of the differentiating metabolites showed outstanding collective power for differentiating cancer from ANIT and for distinguishing CRC tissues at different stages. These findings revealed details in the typical metabonomic phenotypes associated with CRC tissues nondestructively and demonstrated tissue metabonomic phenotyping as an important molecular pathology tool for diagnosis and prognosis of cancerous solid tumors. PMID:26876567

  3. Isolation and Expansion of Mesenchymal Stem/Stromal Cells Derived from Human Placenta Tissue

    PubMed Central

    Pelekanos, Rebecca A.; Sardesai, Varda S.; Futrega, Kathryn; Lott, William B.; Kuhn, Michael; Doran, Michael R.

    2016-01-01

    Mesenchymal stem/stromal cells (MSC) are promising candidates for use in cell-based therapies. In most cases, therapeutic response appears to be cell-dose dependent. Human term placenta is rich in MSC and is a physically large tissue that is generally discarded following birth. Placenta is an ideal starting material for the large-scale manufacture of multiple cell doses of allogeneic MSC. The placenta is a fetomaternal organ from which either fetal or maternal tissue can be isolated. This article describes the placental anatomy and procedure to dissect apart the decidua (maternal), chorionic villi (fetal), and chorionic plate (fetal) tissue. The protocol then outlines how to isolate MSC from each dissected tissue region, and provides representative analysis of expanded MSC derived from the respective tissue types. These methods are intended for pre-clinical MSC isolation, but have also been adapted for clinical manufacture of placental MSC for human therapeutic use. PMID:27340821

  4. Evaluation of tissue-equivalent materials to be used as human brain tissue substitute in dosimetry for diagnostic radiology

    NASA Astrophysics Data System (ADS)

    Ferreira, C. C.; Ximenes Filho, R. E. M.; Vieira, J. W.; Tomal, A.; Poletti, M. E.; Garcia, C. A. B.; Maia, A. F.

    2010-08-01

    Tissue-equivalent materials to be used as substitutes for human brain tissue in dosimetry for diagnostic radiology have been investigated in terms of calculated total mass attenuation coefficient ( μ/ ρ), calculated mass energy-absorption coefficient ( μen/ ρ) and absorbed dose. Measured linear attenuation coefficients ( μ) have been used for benchmarking the calculated total mass attenuation coefficient ( μ/ ρ). The materials examined were bolus, nylon®, orange articulation wax, red articulation wax, PMMA (polymethylmethacrylate), bees wax, paraffin I, paraffin II, pitch and water. The results show that water is the best substitute for brain among the materials investigated. The average percentage differences between the calculated μ/ ρ and μen/ ρ coefficients for water and those for brain were 1.0% and 2.5%, respectively. Absorbed doses determined by Monte Carlo methods confirm water as being the best brain substitute to be used in dosimetry for diagnostic radiology, showing maximum difference of 0.01%. Additionally this study showed that PMMA, a material often used for the manufacturing of head phantoms for computed tomography, cannot be considered to be a suitable substitute for human brain tissue in dosimetry.

  5. Different cross-talk sites between the renin-angiotensin and the kallikrein-kinin systems.

    PubMed

    Su, Jin Bo

    2014-12-01

    Targeting the renin-angiotensin system (RAS) constitutes a major advance in the treatment of cardiovascular diseases. Evidence indicates that angiotensin-converting enzyme inhibitors and angiotensin AT1 receptor blockers act on both the RAS and the kallikrein-kinin system (KKS). In addition to the interaction between the RAS and KKS at the level of angiotensin-converting enzyme catalyzing both angiotensin II generation and bradykinin degradation, the RAS and KKS also interact at other levels: 1) prolylcarboxypeptidase, an angiotensin II inactivating enzyme and a prekallikrein activator; 2) kallikrein, a kinin-generating and prorenin-activating enzyme; 3) angiotensin-(1-7) exerts kininlike effects and potentiates the effects of bradykinin; and 4) the angiotensin AT1 receptor forms heterodimers with the bradykinin B2 receptor. Moreover, angiotensin II enhances B1 and B2 receptor expression via transcriptional mechanisms. These cross-talks explain why both the RAS and KKS are up-regulated in some circumstances, whereas in other circumstances both systems change in the opposite manner, expressed as an activated RAS and a depressed KKS. As the cross-talks between the RAS and the KKS play an important role in response to different stimuli, taking these cross-talks between the two systems into account may help in the development of drugs targeting the two systems. PMID:23386283

  6. The contact activation and kallikrein/kinin systems: pathophysiologic and physiologic activities.

    PubMed

    Schmaier, A H

    2016-01-01

    The contact activation system (CAS) and kallikrein/kinin system (KKS) are older recognized biochemical pathways that include several proteins that skirt the fringes of the blood coagulation, fibrinolytic, complement and renin-angiotensin fields. These proteins initially were proposed as part of the hemostatic pathways because their deficiencies are associated with prolonged clinical assays. However, the absence of bleeding states with deficiencies of factor XII (FXII), prekallikrein (PK) and high-molecular-weight kininogen indicates that the CAS and KKS do not contribute to hemostasis. Since the discovery of the Hageman factor 60 years ago much has been learned about the biochemistry, cell biology and animal physiology of these proteins. The CAS is a pathophysiologic surface defense mechanism against foreign proteins, organisms and artificial materials. The KKS is an inflammatory response mechanism. Targeting their activation through FXIIa or plasma kallikrein inhibition when blood interacts with the artificial surfaces of modern interventional medicine or in acute attacks of hereditary angioedema restores vascular homeostasis. FXII/FXIIa and products that arise with PK deficiency also offer novel ways to reduce arterial and venous thrombosis without an effect on hemostasis. In summary, there is revived interest in the CAS and KKS due to better understanding of their activities. The new appreciation of these systems will lead to several new therapies for a variety of medical disorders. PMID:26565070

  7. Effects of changing salt and water balance on renal kallikrein, kininogen and kinin.

    PubMed

    Weinberg, M; Belknap, S; Trebbin, W; Solomon, R J

    1987-03-01

    The kallikrein-kininogen-kinin system (KKK) has been implicated in the renal sodium excretion response to changes in dietary sodium. However, both increases and decreases in the activity of this system have been observed when urinary sodium excretion is augmented by a variety of maneuvers. To further evaluate the potential physiologic role of this system, we measured three components of the KKK system in urine. Total kallikrein, intact kininogen, and kinin were measured twice in normal individuals during balance on both a high (250 mEq/day) or low (10 mEq/day) sodium intake. A consistent and significant reduction in the activity of all three components of the KKK system was noted during the high salt intake. Furthermore, during the high sodium intake, further acute reductions in components of this system were observed when an acute saline but not water load was administered. The consistent response of the various components of the KKK system to both acute and chronic sodium loading suggests that the system is physiologically linked to the regulation of sodium balance. However, the directional changes argue against a primary natriuretic effect of this system.

  8. The Extreme Anterior Domain Is an Essential Craniofacial Organizer Acting through Kinin-Kallikrein Signaling

    PubMed Central

    Jacox, Laura; Sindelka, Radek; Chen, Justin; Rothman, Alyssa; Dickinson, Amanda; Sive, Hazel

    2014-01-01

    SUMMARY The extreme anterior domain (EAD) is a conserved embryonic region that includes the presumptive mouth. We show that the Kinin-Kallikrein pathway is active in the EAD and necessary for craniofacial development in Xenopus and zebrafish. The mouth failed to form and neural crest (NC) development and migration was abnormal after loss of function (LOF) in the pathway genes kng, encoding Bradykinin (xBdk), carboxypeptidase-N (cpn) that cleaves Bradykinin and neuronal nitric oxide synthase. Consistent with a role for nitric oxide (NO) in face formation, endogenous NO levels declined after LOF in pathway genes but these were restored and a normal face formed after medial implantation of xBdk-beads into LOF embryos. Facial transplants demonstrated that Cpn function from within the EAD is necessary for migration of first arch cranial NC into the face and to promote mouth opening. The study identifies the EAD as an essential craniofacial organizer acting through Kinin-Kallikrein signaling. PMID:25043181

  9. Human tissue lipids: occurrence of fatty acid isomers from dietary hydrogenated oils.

    PubMed

    Ohlrogge, J B; Emken, E A; Gulley, R M

    1981-08-01

    Hydrogenation of vegetable oils produces fatty acids with unusual structures having trans double bonds and double bonds in new positions of the acyl chain. This study was designed to determine which of these fatty acid isomers are incorporated or accumulated in humans during long-term dietary consumption of hydrogenated fats. The double bond position and configuration of the octadecenoate fraction of total lipids extracted from human heart, brain, liver, aorta, and adipose tissue were determined. The level of trans octadecenoate in the tissues as determined by both direct gas-liquid chromatography (GLC) and by GLC after silver nitrate thin-layer chromatography ranged between 0.4 and 5.0%, with an average of 2.7%. Tissues were found to contain trans-octadecenoic isomers having double bonds between the 6 and 15 positions, whereas cis double bonds were found to occur between the 6 and 14 positions. The distribution of double bonds in adipose tissue correlated very closely with the composition of dietary hydrogenated fat. Thus, essentially all of the unusual octadecenoic fatty acid isomers that are produced during vegetable oil hydrogenation are incorporated into human tissue. However, in contrast to results of short-term (1-6 months) feeding studies of animals, our results suggest that long-term (20-60 years) consumption of hydrogenated fats by humans does not lead to substantial preferential accumulation of positional isomers in human tissue total lipids.

  10. Scaffold-Free Human Cardiac Tissue Patch Created from Embryonic Stem Cells

    PubMed Central

    Stevens, Kelly R.; Pabon, Lil; Muskheli, Veronica

    2009-01-01

    Progress in cardiac tissue engineering has been limited by (1) unfavorable cell and host responses to biomaterial scaffolds, (2) lack of suitable human cardiomyocyte sources, and (3) lack of fabrication techniques for scalable production of engineered tissue constructs. Here we report a novel and scalable method to generate scaffold-free human cardiac tissue patches. Human embryonic stem cells were differentiated to cardiomyocytes using activin A and BMP4 and placed into suspension on a rotating orbital shaker. Cells aggregated to form macroscopic disc-shaped patches of beating tissue after 2 days. Patch diameter was directly proportional to input cell number (approximately 11 mm with 12 million cells), and patches were 300–600 μm thick. Cardiomyocytes were concentrated around the patch edges and exhibited increased purity and maturation with time, comprising approximately 80% of total cells after 11 days. Noncardiac cell elements, primarily epithelium, were present at day 2 but were diminished markedly at later time points. Cardiomyocyte proliferation occurred throughout the patches at day 2 but declined by day 8. Patches exhibited automaticity and synchronous calcium transients, indicating electromechanical coupling. These novel scaffold-free human myocardial patches address critical challenges related to human cell sourcing and tissue fabrication that previously inhibited progress in cardiac tissue engineering. PMID:19063661

  11. Modelling soft tissue for kinematic analysis of multi-segment human body models.

    PubMed

    Benham, M P; Wright, D K; Bibb, R

    2001-01-01

    Traditionally biomechanical models represent the musculoskeletal system by a series of rigid links connected by rigidly defined rotational joints. More recently though the mechanics of joints and the action of soft tissues has come under closer scrutiny: biomechanical models might now include a full range of physiological structures. However, soft tissue representation, within multi-segment human body models, presents significant problems; not least in computational speed. We present a method for representing soft tissue physiology which provides for soft tissue wrapping around multiple bony objects; while showing forces at the insertion points, as well as normal reactions due to contact between the soft and bony tissues. These soft tissue representations may therefore be used to constrain the joint, as ligaments would, or to generate motion, like a muscle, so that joints may be modelled which more accurately simulate musculoskeletal motion in all degrees of freedom--rotational and translational. This method produces soft tissues that do not need to be tied to a certain path or route between the bony structures, but may move with the motion of the model; demonstrating a more realistic analysis of soft tissue activity in the musculoskeletal system. The combination of solid geometry models of the skeletal structure, and these novel soft tissue representations, may also provide a useful approach to synthesised human motion.

  12. Distribution of CPP-Protein Complexes in Freshly Resected Human Tissue Material

    PubMed Central

    Saar, Külliki; Saar, Helgi; Hansen, Mats; Langel, Ülo; Pooga, Margus

    2010-01-01

    Interest in cell-penetrating peptides (CPPs) as delivery agents has fuelled a large number of studies conducted on cultured cells and in mice. However, only a few studies have been devoted to the behaviour of CPPs in human tissues. Therefore, we performed ex vivo tissue-dipping experiments where we studied the distribution of CPP-protein complexes in samples of freshly harvested human tissue material. We used the carcinoma or hyperplasia-containing specimens of the uterus and the cervix, obtained as surgical waste from nine hysterectomies. Our aim was to evaluate the tissue of preference (epithelial versus muscular/connective tissue, carcinoma versus adjacent histologically normal tissue) for two well-studied CPPs, the transportan and the TAT-peptide. We complexed biotinylated CPPs with avidin-β-galactosidase (ABG), which enabled us to apply whole-mount X-gal staining as a robust detection method. Our results demonstrate that both peptides enhanced the tissue distribution of ABG. The enhancing effect of the tested CPPs was more obvious in the normal tissue and in some specimens we detected a striking selectivity of CPP-ABG complexes for the normal tissue. This unexpected finding encourages the evaluation of CPPs as local delivery agents in non-malignant situations, for example in the intrauterine gene therapy of benign gynaecological diseases. PMID:27713271

  13. Translation efficiency in humans: tissue specificity, global optimization and differences between developmental stages.

    PubMed

    Waldman, Yedael Y; Tuller, Tamir; Shlomi, Tomer; Sharan, Roded; Ruppin, Eytan

    2010-05-01

    Various studies in unicellular and multicellular organisms have shown that codon bias plays a significant role in translation efficiency (TE) by co-adaptation to the tRNA pool. Yet, in humans and other mammals the role of codon bias is still an open question, with contradictory results from different studies. Here we address this question, performing a large-scale tissue-specific analysis of TE in humans, using the tRNA Adaptation Index (tAI) as a direct measure for TE. We find tAI to significantly correlate with expression levels both in tissue-specific and in global expression measures, testifying to the TE of human tissues. Interestingly, we find significantly higher correlations in adult tissues as opposed to fetal tissues, suggesting that the tRNA pool is more adjusted to the adult period. Optimization based analysis suggests that the tRNA pool-codon bias co-adaptation is globally (and not tissue-specific) driven. Additionally, we find that tAI correlates with several measures related to the protein functionally importance, including gene essentiality. Using inferred tissue-specific tRNA pools lead to similar results and shows that tissue-specific genes are more adapted to their tRNA pool than other genes and that related sets of functional gene groups are translated efficiently in each tissue. Similar results are obtained for other mammals. Taken together, these results demonstrate the role of codon bias in TE in humans, and pave the way for future studies of tissue-specific TE in multicellular organisms. PMID:20097653

  14. Lymphocyte trafficking and HIV infection of human lymphoid tissue in a rotating wall vessel bioreactor

    NASA Technical Reports Server (NTRS)

    Margolis, L. B.; Fitzgerald, W.; Glushakova, S.; Hatfill, S.; Amichay, N.; Baibakov, B.; Zimmerberg, J.

    1997-01-01

    The pathogenesis of HIV infection involves a complex interplay between both the infected and noninfected cells of human lymphoid tissue, the release of free viral particles, the de novo infection of cells, and the recirculatory trafficking of peripheral blood lymphocytes. To develop an in vitro model for studying these various aspects of HIV pathogenesis we have utilized blocks of surgically excised human tonsils and a rotating wall vessel (RWV) cell culture system. Here we show that (1) fragments of the surgically excised human lymphoid tissue remain viable and retain their gross cytoarchitecture for at least 3 weeks when cultured in the RWV system; (2) such lymphoid tissue gradually shows a loss of both T and B cells to the surrounding growth medium; however, this cellular migration is reversible as demonstrated by repopulation of the tissue by labeled cells from the growth medium; (3) this cellular migration may be partially or completely inhibited by embedding the blocks of lymphoid tissue in either a collagen or agarose gel matrix; these embedded tissue blocks retain most of the basic elements of a normal lymphoid cytoarchitecture; and (4) both embedded and nonembedded RWV-cultured blocks of human lymphoid tissue are capable of productive infection by HIV-1 of at least three various strains of different tropism and phenotype, as shown by an increase in both p24 antigen levels and free virus in the culture medium, and by the demonstration of HIV-1 RNA-positive cells inside the tissue identified by in situ hybridization. It is therefore reasonable to suggest that gel-embedded and nonembedded blocks of human lymphoid tissue, cocultured with a suspension of tonsillar lymphocytes in an RWV culture system, constitute a useful model for simulating normal lymphocyte recirculatory traffic and provide a new tool for testing the various aspects of HIV pathogenesis.

  15. History of Diffuse Optical Spectroscopy of Human Tissue

    NASA Astrophysics Data System (ADS)

    Huppert, Theodore J.

    Diffuse optical spectroscopy is a noninvasive method that uses low levels of near-infrared light to measure blood oxygenation in the brain. Over the last 35 years, the number of diffuse optical studies and the range of clinical and research applications have grown steadily. Compared to other neuroimaging methods to measure cerebral blood oxygenation, such as magnetic resonance imaging or positron emission tomography, diffuse optical imaging (DOI) is more cost effective and often uses small portable instrumentation. Wireless and bedside optical systems are currently produced commercially. The portability of these instruments has extended the use of optical methods into several unique applications including brain imaging in infants and children, studies of the brain during ambulatory tasks such as walking or balance, and interoperative brain assessments. This chapter will introduce the history and basic principles of DOI including discussion of the factors contributing to the optical properties of tissue, instrumentation, and an overview of applications of the technology.

  16. Adult human adipose tissue contains several types of multipotent cells.

    PubMed

    Tallone, Tiziano; Realini, Claudio; Böhmler, Andreas; Kornfeld, Christopher; Vassalli, Giuseppe; Moccetti, Tiziano; Bardelli, Silvana; Soldati, Gianni

    2011-04-01

    Multipotent mesenchymal stromal cells (MSCs) are a type of adult stem cells that can be easily isolated from various tissues and expanded in vitro. Many reports on their pluripotency and possible clinical applications have raised hopes and interest in MSCs. In an attempt to unify the terminology and the criteria to label a cell as MSC, in 2006 the International Society for Cellular Therapy (ISCT) proposed a standard set of rules to define the identity of these cells. However, MSCs are still extracted from different tissues, by diverse isolation protocols, are cultured and expanded in different media and conditions. All these variables may have profound effects on the selection of cell types and the composition of heterogeneous subpopulations, on the selective expansion of specific cell populations with totally different potentials and ergo, on the long-term fate of the cells upon in vitro culture. Therefore, specific molecular and cellular markers that identify MSCs subsets as well as standardization of expansion protocols for these cells are urgently needed. Here, we briefly discuss new useful markers and recent data supporting the rapidly emerging concept that many different types of progenitor cells are found in close association with blood vessels. This knowledge may promote the necessary technical improvements required to reduce variability and promote higher efficacy and safety when isolating and expanding these cells for therapeutic use. In the light of the discussed data, particularly the identification of new markers, and advances in the understanding of fundamental MSC biology, we also suggest a revision of the 2006 ISCT criteria.

  17. Molecular Portrait of the Normal Human Breast Tissue and Its Influence on Breast Carcinogenesis

    PubMed Central

    Margan, Madalin Marius; Jitariu, Andreea Adriana; Nica, Cristian; Raica, Marius

    2016-01-01

    Normal human breast tissue consists of epithelial and nonepithelial cells with different molecular profiles and differentiation grades. This molecular heterogeneity is known to yield abnormal clones that may contribute to the development of breast carcinomas. Stem cells that are found in developing and mature breast tissue are either positive or negative for cytokeratin 19 depending on their subtype. These cells are able to generate carcinogenesis along with mature cells. However, scientific data remains controversial regarding the monoclonal or polyclonal origin of breast carcinomas. The majority of breast carcinomas originate from epithelial cells that normally express BRCA1. The consecutive loss of the BRCA1 gene leads to various abnormalities in epithelial cells. Normal breast epithelial cells also express hypoxia inducible factor (HIF) 1α and HIF-2α that are associated with a high metastatic rate and a poor prognosis for malignant lesions. The nuclear expression of estrogen receptor (ER) and progesterone receptor (PR) in normal human breast tissue is maintained in malignant tissue as well. Several controversies regarding the ability of ER and PR status to predict breast cancer outcome remain. Both ER and PR act as modulators of cell activity in normal human breast tissue. Ki-67 positivity is strongly correlated with tumor grade although its specific role in applied therapy requires further studies. Human epidermal growth factor receptor 2 (HER2) oncoprotein is less expressed in normal human breast specimens but is highly expressed in certain malignant lesions of the breast. Unlike HER2, epidermal growth factor receptor expression is similar in both normal and malignant tissues. Molecular heterogeneity is not only found in breast carcinomas but also in normal breast tissue. Therefore, the molecular mapping of normal human breast tissue might represent a key research area to fully elucidate the mechanisms of breast carcinogenesis. PMID:27382385

  18. Molecular Portrait of the Normal Human Breast Tissue and Its Influence on Breast Carcinogenesis.

    PubMed

    Margan, Madalin Marius; Jitariu, Andreea Adriana; Cimpean, Anca Maria; Nica, Cristian; Raica, Marius

    2016-06-01

    Normal human breast tissue consists of epithelial and nonepithelial cells with different molecular profiles and differentiation grades. This molecular heterogeneity is known to yield abnormal clones that may contribute to the development of breast carcinomas. Stem cells that are found in developing and mature breast tissue are either positive or negative for cytokeratin 19 depending on their subtype. These cells are able to generate carcinogenesis along with mature cells. However, scientific data remains controversial regarding the monoclonal or polyclonal origin of breast carcinomas. The majority of breast carcinomas originate from epithelial cells that normally express BRCA1. The consecutive loss of the BRCA1 gene leads to various abnormalities in epithelial cells. Normal breast epithelial cells also express hypoxia inducible factor (HIF) 1α and HIF-2α that are associated with a high metastatic rate and a poor prognosis for malignant lesions. The nuclear expression of estrogen receptor (ER) and progesterone receptor (PR) in normal human breast tissue is maintained in malignant tissue as well. Several controversies regarding the ability of ER and PR status to predict breast cancer outcome remain. Both ER and PR act as modulators of cell activity in normal human breast tissue. Ki-67 positivity is strongly correlated with tumor grade although its specific role in applied therapy requires further studies. Human epidermal growth factor receptor 2 (HER2) oncoprotein is less expressed in normal human breast specimens but is highly expressed in certain malignant lesions of the breast. Unlike HER2, epidermal growth factor receptor expression is similar in both normal and malignant tissues. Molecular heterogeneity is not only found in breast carcinomas but also in normal breast tissue. Therefore, the molecular mapping of normal human breast tissue might represent a key research area to fully elucidate the mechanisms of breast carcinogenesis. PMID:27382385

  19. Immunolocalization of nitric oxide synthase isoforms in human archival and rat tissues, and cultured cells.

    PubMed

    Martins, Antonio R; Zanella, Cesar A B; Zucchi, Fabiola C R; Dombroski, Thaís C D; Costa, Edmar T; Guethe, Liliane M; Oliveira, Alina O; Donatti, Ana L F; Neder, Luciano; Chimelli, Leila; De Nucci, Gilberto; Lee-Ho, Paulo; Murad, Ferid

    2011-05-15

    Nitric oxide (NO) exerts important physiological and pathological roles in humans. The study of NO requires the immunolocalization of its synthesizing enzymes, neuronal, endothelial and inducible NO synthases (NOS). NOS are labile to formalin-fixation and paraffin-embedding, which are used to prepare human archival tissues. This lability has made NOS immunohistochemical studies difficult, and a detailed protocol is not yet available. We describe here a protocol for the immunolocalization of NOS isoforms in human archival cerebellum and non-nervous tissues, and in rat tissues and cultured cells. Neuronal NOS antigenicity in human archival and rat nervous tissue sections was microwave-retrieved in 50 mM Tris-HCl buffer, pH 9.5, for 20 min at 900 W. Neuronal NOS was expressed in stellate, basket, Purkinje and granule cells in human and rat cerebellum. Archival and frozen human cerebellar sections showed the same neuronal NOS staining pattern. Archival cerebellar sections not subjected to antigen retrieval stained weakly. Antigenicity of inducible NOS in human lung was best retrieved in 10 mM sodium citrate buffer, pH 6.0, for 15 min at 900 W. Inflammatory cells in a human lung tuberculoma were strongly stained by anti-inducible NOS antibody. Anti-endothelial NOS strongly stained kidney glomeruli. Cultured PC12 cells were strongly stained by anti-neuronal NOS without antigen retrieving. The present immunohistochemistry protocol is easy to perform, timeless, and suitable for the localization of NOS isoforms in nervous and non-nervous tissues, in human archival and rat tissues. It has been extensively used in our laboratory, and is also appropriate for other antigens.

  20. 78 FR 41403 - Agency Information Collection Activities; Proposed Collection; Comment Request; Human Tissue...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-10

    ... transmission of human immunodeficiency virus, hepatitis B, and hepatitis C, through the use of human tissue for... 1270.31(a) and (b) also requires recording and justification of any deviation from the written... (5 CFR 1320.3(b)(2)). The recordkeeping burden, thus, is estimated for the remaining...

  1. [Organochlorine pesticide residues in human adipose tissue in Costa Rica].

    PubMed

    Barquero, M; Constenla, M A

    1986-06-01

    Organochlorine pesticide residues were found in 82 samples of human adipose material from 82 surgical cases in 16 Costa Rica hospitals. Identification was made by gas-liquid chromatography. The highest pesticide concentration was that of DDT and its metabolites (33.16 micrograms/g). Residues of almost all commercial pesticides were also found. Concentrations of alpha-chlordane. Aldrin and Polychlorinated biphenyls were not significant.

  2. Genome-wide quantification of rare somatic mutations in normal human tissues using massively parallel sequencing.

    PubMed

    Hoang, Margaret L; Kinde, Isaac; Tomasetti, Cristian; McMahon, K Wyatt; Rosenquist, Thomas A; Grollman, Arthur P; Kinzler, Kenneth W; Vogelstein, Bert; Papadopoulos, Nickolas

    2016-08-30

    We present the bottleneck sequencing system (BotSeqS), a next-generation sequencing method that simultaneously quantifies rare somatic point mutations across the mitochondrial and nuclear genomes. BotSeqS combines molecular barcoding with a simple dilution step immediately before library amplification. We use BotSeqS to show age- and tissue-dependent accumulations of rare mutations and demonstrate that somatic mutational burden in normal human tissues can vary by several orders of magnitude, depending on biologic and environmental factors. We further show major differences between the mutational patterns of the mitochondrial and nuclear genomes in normal tissues. Lastly, the mutation spectra of normal tissues were different from each other, but similar to those of the cancers that arose in them. This technology can provide insights into the number and nature of genetic alterations in normal tissues and can be used to address a variety of fundamental questions about the genomes of diseased tissues.

  3. Genome-wide quantification of rare somatic mutations in normal human tissues using massively parallel sequencing

    PubMed Central

    Hoang, Margaret L.; Kinde, Isaac; Tomasetti, Cristian; McMahon, K. Wyatt; Rosenquist, Thomas A.; Grollman, Arthur P.; Kinzler, Kenneth W.; Vogelstein, Bert; Papadopoulos, Nickolas

    2016-01-01

    We present the bottleneck sequencing system (BotSeqS), a next-generation sequencing method that simultaneously quantifies rare somatic point mutations across the mitochondrial and nuclear genomes. BotSeqS combines molecular barcoding with a simple dilution step immediately before library amplification. We use BotSeqS to show age- and tissue-dependent accumulations of rare mutations and demonstrate that somatic mutational burden in normal human tissues can vary by several orders of magnitude, depending on biologic and environmental factors. We further show major differences between the mutational patterns of the mitochondrial and nuclear genomes in normal tissues. Lastly, the mutation spectra of normal tissues were different from each other, but similar to those of the cancers that arose in them. This technology can provide insights into the number and nature of genetic alterations in normal tissues and can be used to address a variety of fundamental questions about the genomes of diseased tissues. PMID:27528664

  4. Acknowledging tissue donation: Human cadaveric specimens in musculoskeletal research.

    PubMed

    Winkelmann, Andreas; Heinze, Anne-Kathrin; Hendrix, Sven

    2016-01-01

    Human cadaveric specimens are an important resource for research, particularly in biomechanical studies, but their use also raises ethical questions and cannot simply be taken for granted. It was asked how much information authors publishing musculoskeletal research actually give about such specimens and about how they were acquired. The aim was to formulate recommendations on how this reporting might be improved. Relevant articles published between 2009 and 2012 in four North American or European journals were scanned for information regarding the characteristics of the human specimens used, their institutional source and the ethical or legal context of their acquisition. While the majority of articles report biological characteristics of specimens (sex, age at death, preservation method), only 40% of articles refer to body donation, only 23% report the institution that provided specimens, and only 17% refer to some kind of formalized approval of their research. There were regional and journal-to-journal differences. No standard for reporting studies involving human specimens could be detected. It is suggested that such a standard be developed by researchers and editors. Information on the source of specimens and on the ethical or legal basis should be regularly reported to acknowledge this unique research resource and to preserve the good relationship between researchers and the communities, that provide the required specimens by body donation and upon which researchers depend.

  5. Design and characterization of protein films for modeling near-infrared spectra of human tissue.

    PubMed

    Karunathilaka, Sanjeewa R; Small, Gary W

    2015-06-21

    Near-infrared (near-IR) spectroscopy has been investigated for use in direct measurements of human tissue for the purpose of quantifying clinically relevant analytes such as glucose. Spectra collected by transmitting near-IR light through human tissue reveal the presence of both aqueous components and solid structures within the optical path of the measurement. Developing technology for use in making these measurements requires either the availability of human subjects or an in vitro experimental platform that can effectively simulate the spectroscopic properties of tissue. This paper describes the preparation and testing of films of collagen and keratin, the two proteins that comprise the bulk of the solid material in the human epidermis and dermis. By placing these films in the optical path of a near-IR spectrometer together with an aqueous sample cell, a phantom can be constructed that allows experiments to be performed that simulate noninvasive measurements of tissue. In this work, both constant and variable thickness films are prepared and evaluated by use of a regression fit to spectra of human tissue. The stability and spectral reproducibility of the prepared films are also assessed. The regression fits to the human subject spectra yield values of R(2) ranging from 0.97 to 0.99 and the films are found to yield spectra that vary by less than a 2% relative standard deviation under three different reproducibility tests.

  6. Methods of Assessing Human Tendon Metabolism and Tissue Properties in Response to Changes in Mechanical Loading.

    PubMed

    Heinemeier, Katja M; Kjaer, Michael; Magnusson, S Peter

    2016-01-01

    In recent years a number of methodological developments have improved the opportunities to study human tendon. Microdialysis enables sampling of interstitial fluid in the peritendon tissue, while sampling of human tendon biopsies allows direct analysis of tendon tissue for gene- and protein expression as well as protein synthesis rate. Further the (14)C bomb-pulse method has provided data on long-term tissue turnover in human tendon. Non-invasive techniques allow measurement of tendon metabolism (positron emission tomography (PET)), tendon morphology (magnetic resonance imaging (MRI)), and tendon mechanical properties (ultrasonography combined with force measurement during movement). Finally, 3D cell cultures of human tendon cells provide the opportunity to investigate cell-matrix interactions in response to various interventions. PMID:27535251

  7. Brown adipose tissue improves whole-body glucose homeostasis and insulin sensitivity in humans.

    PubMed

    Chondronikola, Maria; Volpi, Elena; Børsheim, Elisabet; Porter, Craig; Annamalai, Palam; Enerbäck, Sven; Lidell, Martin E; Saraf, Manish K; Labbe, Sebastien M; Hurren, Nicholas M; Yfanti, Christina; Chao, Tony; Andersen, Clark R; Cesani, Fernando; Hawkins, Hal; Sidossis, Labros S

    2014-12-01

    Brown adipose tissue (BAT) has attracted scientific interest as an antidiabetic tissue owing to its ability to dissipate energy as heat. Despite a plethora of data concerning the role of BAT in glucose metabolism in rodents, the role of BAT (if any) in glucose metabolism in humans remains unclear. To investigate whether BAT activation alters whole-body glucose homeostasis and insulin sensitivity in humans, we studied seven BAT-positive (BAT(+)) men and five BAT-negative (BAT(-)) men under thermoneutral conditions and after prolonged (5-8 h) cold exposure (CE). The two groups were similar in age, BMI, and adiposity. CE significantly increased resting energy expenditure, whole-body glucose disposal, plasma glucose oxidation, and insulin sensitivity in the BAT(+) group only. These results demonstrate a physiologically significant role of BAT in whole-body energy expenditure, glucose homeostasis, and insulin sensitivity in humans, and support the notion that BAT may function as an antidiabetic tissue in humans.

  8. Species-Specific Metastasis of Human Tumor Cells in the Severe Combined Immunodeficiency Mouse Engrafted with Human Tissue

    NASA Astrophysics Data System (ADS)

    Shtivelman, Emma; Namikawa, Reiko

    1995-05-01

    We have attempted to model human metastatic disease by implanting human target organs into the immunodeficient C.B-17 scid/scid (severe combined immunodeficiency; SCID) mouse, creating SCID-hu mice. Preferential metastasis to implants of human fetal lung and human fetal bone marrow occurred after i.v. injection of human small cell lung cancer (SCLC) cells into SCID-hu mice; the homologous mouse organs were spared. Clinically more aggressive variant SCLC cells metastasized more efficiently to human fetal lung implants than did cells from classic SCLC. Metastasis of variant SCLC to human fetal bone marrow was enhanced in SCID-hu mice exposed to γ-irradiation or to interleukin 1α. These data indicate that the SCID-hu mice may provide a model in which to study species- and tissue-specific steps of the human metastatic process.

  9. Expression of ceramide-metabolising enzymes in subcutaneous and intra-abdominal human adipose tissue

    PubMed Central

    2012-01-01

    Background Inflammation and increased ceramide concentrations characterise adipose tissue of obese women with high liver fat content compared to equally obese women with normal liver fat content. The present study characterises enzymes involved in ceramide metabolism in subcutaneous and intra-abdominal adipose tissue. Methods Pathways leading to increased ceramide concentrations in inflamed versus non-inflamed adipose tissue were investigated by quantifying expression levels of key enzymes involved in ceramide metabolism. Sphingomyelinases (sphingomyelin phosphodiesterases SMPD1-3) were investigated further using immunohistochemistry to establish their location within adipose tissue, and their mRNA expression levels were determined in subcutaneous and intra-abdominal adipose tissue from both non-obese and obese subject. Results Gene expression levels of sphingomyelinases, enzymes that hydrolyse sphingomyelin to ceramide, rather than enzymes involved in de novo ceramide synthesis, were higher in inflamed compared to non-inflamed adipose tissue of obese women (with high and normal liver fat contents respectively). Sphingomyelinases were localised to both macrophages and adipocytes, but also to blood vessels and to extracellular regions surrounding vessels within adipose tissue. Expression levels of SMPD3 mRNA correlated significantly with concentrations of different ceramides and sphingomyelins. In both non-obese and obese subjects SMPD3 mRNA levels were higher in the more inflamed intra-abdominal compared to the subcutaneous adipose tissue depot. Conclusions Generation of ceramides within adipose tissue as a result of sphingomyelinase action may contribute to inflammation in human adipose tissue. PMID:22974251

  10. Detection of VEGF-Axxxb Isoforms in Human Tissues

    PubMed Central

    Bates, David O.; Mavrou, Athina; Qiu, Yan; Carter, James G.; Hamdollah-Zadeh, Maryam; Barratt, Shaney; Gammons, Melissa V.; Millar, Ann B.; Salmon, Andrew H. J.; Oltean, Sebastian; Harper, Steven J.

    2013-01-01

    Vascular Endothelial Growth Factor-A (VEGF-A) can be generated as multiple isoforms by alternative splicing. Two families of isoforms have been described in humans, pro-angiogenic isoforms typified by VEGF-A165a, and anti-angiogenic isoforms typified by VEGF-A165b. The practical determination of expression levels of alternative isoforms of the same gene may be complicated by experimental protocols that favour one isoform over another, and the use of specific positive and negative controls is essential for the interpretation of findings on expression of the isoforms. Here we address some of the difficulties in experimental design when investigating alternative splicing of VEGF isoforms, and discuss the use of appropriate control paradigms. We demonstrate why use of specific control experiments can prevent assumptions that VEGF-A165b is not present, when in fact it is. We reiterate, and confirm previously published experimental design protocols that demonstrate the importance of using positive controls. These include using known target sequences to show that the experimental conditions are suitable for PCR amplification of VEGF-A165b mRNA for both q-PCR and RT-PCR and to ensure that mispriming does not occur. We also provide evidence that demonstrates that detection of VEGF-A165b protein in mice needs to be tightly controlled to prevent detection of mouse IgG by a secondary antibody. We also show that human VEGF165b protein can be immunoprecipitated from cultured human cells and that immunoprecipitating VEGF-A results in protein that is detected by VEGF-A165b antibody. These findings support the conclusion that more information on the biology of VEGF-A165b isoforms is required, and confirm the importance of the experimental design in such investigations, including the use of specific positive and negative controls. PMID:23935865

  11. Positive Selection Shaped the Convergent Evolution of Independently Expanded Kallikrein Subfamilies Expressed in Mouse and Rat Saliva Proteomes

    PubMed Central

    Karn, Robert C.; Laukaitis, Christina M.

    2011-01-01

    We performed proteomics studies of salivas from the genome mouse (C57BL/6 strain) and the genome rat (BN/SsNHsd/Mcwi strain). Our goal was to identify salivary proteins with one or more of three characteristics that may indicate that they have been involved in adaptation: 1) rapid expansion of their gene families; 2) footprints of positive selection; and/or 3) sex-limited expression. The results of our proteomics studies allow direct comparison of the proteins expressed and their levels between the sexes of the two rodent species. Twelve members of the Mus musculus species-specific kallikrein subfamily Klk1b showed sex-limited expression in the mouse saliva proteomes. By contrast, we did not find any of the Rattus norvegicus species-specific kallikrein subfamily Klk1c proteins in male or female genome rat, nor transcripts in their submandibular glands. On the other hand, we detected expression of this family as transcripts in the submandibular glands of both sexes of Sprague-Dawley rats. Using the CODEML program in the PAML package, we demonstrate that the two rodent kallikrein subfamilies have apparently evolved rapidly under the influence of positive selection that continually remodeled the amino acid sites on the same face in the members of the subfamilies. Thus, although their kallikrein subfamily expansions were independent, this evolutionary pattern has occurred in parallel in the two rodent species, suggesting a form of convergent evolution at the molecular level. On the basis of this new data, we suggest that the previous speculative function of the species-specific rodent kallikreins as important solely in wound healing in males be investigated further. In addition to or instead of that function, we propose that their sex-limited expression, coupled with their rapid evolution may be clues to an as-yet-undetermined interaction between the sexes. PMID:21695125

  12. Integrated interactions database: tissue-specific view of the human and model organism interactomes.

    PubMed

    Kotlyar, Max; Pastrello, Chiara; Sheahan, Nicholas; Jurisica, Igor

    2016-01-01

    IID (Integrated Interactions Database) is the first database providing tissue-specific protein-protein interactions (PPIs) for model organisms and human. IID covers six species (S. cerevisiae (yeast), C. elegans (worm), D. melonogaster (fly), R. norvegicus (rat), M. musculus (mouse) and H. sapiens (human)) and up to 30 tissues per species. Users query IID by providing a set of proteins or PPIs from any of these organisms, and specifying species and tissues where IID should search for interactions. If query proteins are not from the selected species, IID enables searches across species and tissues automatically by using their orthologs; for example, retrieving interactions in a given tissue, conserved in human and mouse. Interaction data in IID comprises three types of PPI networks: experimentally detected PPIs from major databases, orthologous PPIs and high-confidence computationally predicted PPIs. Interactions are assigned to tissues where their proteins pairs or encoding genes are expressed. IID is a major replacement of the I2D interaction database, with larger PPI networks (a total of 1,566,043 PPIs among 68,831 proteins), tissue annotations for interactions, and new query, analysis and data visualization capabilities. IID is available at http://ophid.utoronto.ca/iid.

  13. Development of human nervous tissue upon differentiation of embryonic stem cells in three-dimensional culture.

    PubMed

    Preynat-Seauve, Olivier; Suter, David M; Tirefort, Diderik; Turchi, Laurent; Virolle, Thierry; Chneiweiss, Herve; Foti, Michelangelo; Lobrinus, Johannes-Alexander; Stoppini, Luc; Feki, Anis; Dubois-Dauphin, Michel; Krause, Karl Heinz

    2009-03-01

    Researches on neural differentiation using embryonic stem cells (ESC) require analysis of neurogenesis in conditions mimicking physiological cellular interactions as closely as possible. In this study, we report an air-liquid interface-based culture of human ESC. This culture system allows three-dimensional cell expansion and neural differentiation in the absence of added growth factors. Over a 3-month period, a macroscopically visible, compact tissue developed. Histological coloration revealed a dense neural-like neural tissue including immature tubular structures. Electron microscopy, immunochemistry, and electrophysiological recordings demonstrated a dense network of neurons, astrocytes, and oligodendrocytes able to propagate signals. Within this tissue, tubular structures were niches of cells resembling germinal layers of human fetal brain. Indeed, the tissue contained abundant proliferating cells expressing markers of neural progenitors. Finally, the capacity to generate neural tissues on air-liquid interface differed for different ESC lines, confirming variations of their neurogenic potential. In conclusion, this study demonstrates in vitro engineering of a human neural-like tissue with an organization that bears resemblance to early developing brain. As opposed to previously described methods, this differentiation (a) allows three-dimensional organization, (b) yields dense interconnected neural tissue with structurally and functionally distinct areas, and (c) is spontaneously guided by endogenous developmental cues.

  14. Insulin Regulates the Unfolded Protein Response in Human Adipose Tissue

    PubMed Central

    Boden, Guenther; Cheung, Peter; Salehi, Sajad; Homko, Carol; Loveland-Jones, Catherine; Jayarajan, Senthil; Stein, T. Peter; Williams, Kevin Jon; Liu, Ming-Lin; Barrero, Carlos A.; Merali, Salim

    2014-01-01

    Endoplasmic reticulum (ER) stress is increased in obesity and is postulated to be a major contributor to many obesity-related pathologies. Little is known about what causes ER stress in obese people. Here, we show that insulin upregulated the unfolded protein response (UPR), an adaptive reaction to ER stress, in vitro in 3T3-L1 adipocytes and in vivo, in subcutaneous (sc) adipose tissue of nondiabetic subjects, where it increased the UPR dose dependently over the entire physiologic insulin range (from ∼35 to ∼1,450 pmol/L). The insulin-induced UPR was not due to increased glucose uptake/metabolism and oxidative stress. It was associated, however, with increased protein synthesis, with accumulation of ubiquitination associated proteins, and with multiple posttranslational protein modifications (acetylations, methylations, nitrosylations, succinylation, and ubiquitinations), some of which are potential causes for ER stress. These results reveal a new physiologic role of insulin and provide a putative mechanism for the development of ER stress in obesity. They may also have clinical and therapeutic implications, e.g., in diabetic patients treated with high doses of insulin. PMID:24130338

  15. Luminescence lifetime determination for oxygen imaging in human tissue

    NASA Astrophysics Data System (ADS)

    Lochmann, C.; Häupl, T.; Beuthan, J.

    2008-02-01

    Imaging and monitoring of biochemical parameters in vitro and in vivo have become the goal of many investigations in medical physics. The main imaging technique used is laser-induced luminescence due to its cost effectiveness and diversity of applications [1]. One key parameter in medical investigations, for instance to control photodynamic therapy, is the molecular oxygen concentration. The use of optical methods provides possible means of measuring molecular oxygen. The basis of such a method is the measurement of the luminescence lifetime of a dye which is quenched by molecular oxygen. The molecular oxygen concentration can be monitored two-dimensionally by pixel-wise determination of the luminescence lifetime with a CCD-camera. An oxygen imaging system based on this principle was built and tested with a commercially available oxygen sensitive sol gel-layer. The embedded ruthenium complex is quenched by molecular oxygen and because of the oxygen permeability of the layer; it is suitable for oxygen measurements. The characteristics and dependence on the pH-value and temperature of the luminescence lifetime of the layer were examined in preparation for measurements on tissue to exclude cross-correlation of other quenching processes.

  16. Functional Local Renin-Angiotensin System in Human and Rat Periodontal Tissue

    PubMed Central

    Santos, Carlos F.; Morandini, Ana C.; Dionísio, Thiago J.; Faria, Flávio A.; Lima, Marta C.; Figueiredo, Caio M.; Colombini-Ishikiriama, Bella L.; Sipert, Carla R.; Maciel, Rubens P.; Akashi, Ana P.; Souza, Gabriela P.; Garlet, Gustavo P.; Rodini, Camila O.; Amaral, Sandra L.; Becari, Christiane; Salgado, Maria C.; Oliveira, Eduardo B.; Matus, Isaac; Didier, Daniela N.; Greene, Andrew S.

    2015-01-01

    The initiation or progression of periodontitis might involve a local renin-angiotensin system (RAS) in periodontal tissue. The aim of this study was to further characterize the local RAS in human and rat periodontal tissues between healthy and periodontally-affected tissue. Components of the RAS were investigated using in vitro, ex vivo and in vivo experiments involving both human and Wistar rat periodontium. Although not upregulated when challenged with P. gingivalis-lipopolysaccharide, human gingival and periodontal ligament fibroblasts expressed RAS components. Likewise, healthy and inflamed human gingiva expressed RAS components, some of which were shown to be functional, yet no differences in expression were found between healthy and diseased gingiva. However, in inflamed tissue the immunoreactivity was greater for the AT1R compared to AT2R in fibroblasts. When compared to healthy tissue, ACE activity was increased in human gingiva from volunteers with gingivitis. Human-gingiva homogenates generated Ang II, Ang 1-9 and Ang 1-7 when incubated with precursors. In gingiva homogenates, Ang II formation from Ang I was nearly abolished only when captopril and chymostatin were combined. Ang 1-7 formation was significantly greater when human gingiva homogenates were incubated with chymostatin alone compared to incubation without any inhibitor, only captopril, or captopril and chymostatin. In rat gingiva, RAS components were also found; their expression was not different between healthy and experimentally induced periodontitis (EP) groups. However, renin inhibition (aliskiren) and an AT1R antagonist (losartan) significantly blocked EP-alveolar-bone loss in rats. Collectively, these data are consistent with the hypothesis that a local RAS system is not only present but is also functional in both human and rat periodontal tissue. Furthermore, blocking AT1R and renin can significantly prevent periodontal bone loss induced by EP in rats. PMID:26244896

  17. Mesenchymal Stromal Cells Derived from Human Umbilical Cord Tissues: Primitive Cells with Potential for Clinical and Tissue Engineering Applications

    NASA Astrophysics Data System (ADS)

    Moretti, Pierre; Hatlapatka, Tim; Marten, Dana; Lavrentieva, Antonina; Majore, Ingrida; Hass, Ralf; Kasper, Cornelia

    Mesenchymal stem or stromal cells (MSCs) have a high potential for cell-based therapies as well as for tissue engineering applications. Since Friedenstein first isolated stem or precursor cells from the human bone marrow (BM) stroma that were capable of osteogenesis, BM is currently the most common source for MSCs. However, BM presents several disadvantages, namely low frequency of MSCs, high donor-dependent variations in quality, and painful invasive intervention. Thus, tremendous research efforts have been observed during recent years to find alternative sources for MSCs.

  18. Immunohistochemical Study of Expression of Sohlh1 and Sohlh2 in Normal Adult Human Tissues

    PubMed Central

    Zhang, Xiaoli; Liu, Ruihua; Su, Zhongxue; Zhang, Yuecun; Zhang, Wenfang; Liu, Xinyu; Wang, Fuwu; Guo, Yuji; Li, Chuangang; Hao, Jing

    2015-01-01

    The expression pattern of Sohlh1 (spermatogenesis and oogenesis specific basic helix-loop-helix 1) and Sohlh2 in mice has been reported in previous studies. Sohlh1 and Sohlh2 are specifically expressed in spermatogonia, prespermatogonia in male mice and oocytes of primordial and primary follicles in female mice. In this report, we studied the expression pattern of Sohlh1 and Sohlh2 in human adult tissues. Immunohistochemical staining of Sohlh1 and Sohlh2 was performed in 5 samples of normal ovaries and testes, respectively. The results revealed that Sohlh genes are not only expressed in oocytes and spermatogonia, but also in granular cells, theca cells, Sertoli cells and Leydig cells, and in smooth muscles of blood vessel walls. To further investigate the expression of Sohlh genes in other adult human tissues, we collected representative normal adult tissues developed from three embryonic germ layers. Compared with the expression in mice, Sohlhs exhibited a much more extensive expression pattern in human tissues. Sohlhs were detected in testis, ovary and epithelia developed from embryonic endoderm, ectoderm and tissues developed from embryonic mesoderm. Sohlh signals were found in spermatogonia, Sertoli cells and also Leydig cells in testis, while in ovary, the expression was mainly in oocytes of primordial and primary follicles, granular cells and theca cells of secondary follicles. Compared with Sohlh2, the expression of Sohlh1 was stronger and more extensive. Our study explored the expression of Sohlh genes in human tissues and might provide insights for functional studies of Sohlh genes. PMID:26375665

  19. Decellularization of human stromal refractive lenticules for corneal tissue engineering

    PubMed Central

    Yam, Gary Hin-Fai; Yusoff, Nur Zahirah Binte M.; Goh, Tze-Wei; Setiawan, Melina; Lee, Xiao-Wen; Liu, Yu-Chi; Mehta, Jodhbir S.

    2016-01-01

    Small incision lenticule extraction (SMILE) becomes a procedure to correct myopia. The extracted lenticule can be used for other clinical scenarios. To prepare for allogeneic implantation, lenticule decellularization with preserved optical property, stromal architecture and chemistry would be necessary. We evaluated different methods to decellularize thin human corneal stromal lenticules created by femtosecond laser. Treatment with 0.1% sodium dodecylsulfate (SDS) followed by extensive washes was the most efficient protocol to remove cellular and nuclear materials. Empty cell space was found inside the stroma, which displayed aligned collagen fibril architecture similar to native stroma. The SDS-based method was superior to other treatments with hyperosmotic 1.5 M sodium chloride, 0.1% Triton X-100 and nucleases (from 2 to 10 U/ml DNase and RNase) in preserving extracellular matrix content (collagens, glycoproteins and glycosaminoglycans). The stromal transparency and light transmittance was indifferent to untreated lenticules. In vitro recellularization showed that the SDS-treated lenticules supported corneal stromal fibroblast growth. In vivo re-implantation into a rabbit stromal pocket further revealed the safety and biocompatibility of SDS-decellularized lenticules without short- and long-term rejection risk. Our results concluded that femtosecond laser-derived human stromal lenticules decellularized by 0.1% SDS could generate a transplantable bioscaffold with native-like stromal architecture and chemistry. PMID:27210519

  20. Breast Cancer Cell Colonization of the Human Bone Marrow Adipose Tissue Niche1

    PubMed Central

    Templeton, Zach S.; Lie, Wen-Rong; Wang, Weiqi; Rosenberg-Hasson, Yael; Alluri, Rajiv V.; Tamaresis, John S.; Bachmann, Michael H.; Lee, Kitty; Maloney, William J.; Contag, Christopher H.; King, Bonnie L.

    2015-01-01

    BACKGROUND/OBJECTIVES: Bone is a preferred site of breast cancer metastasis, suggesting the presence of tissue-specific features that attract and promote the outgrowth of breast cancer cells. We sought to identify parameters of human bone tissue associated with breast cancer cell osteotropism and colonization in the metastatic niche. METHODS: Migration and colonization patterns of MDA-MB-231-fLuc-EGFP (luciferase-enhanced green fluorescence protein) and MCF-7-fLuc-EGFP breast cancer cells were studied in co-culture with cancellous bone tissue fragments isolated from 14 hip arthroplasties. Breast cancer cell migration into tissues and toward tissue-conditioned medium was measured in Transwell migration chambers using bioluminescence imaging and analyzed as a function of secreted factors measured by multiplex immunoassay. Patterns of breast cancer cell colonization were evaluated with fluorescence microscopy and immunohistochemistry. RESULTS: Enhanced MDA-MB-231-fLuc-EGFP breast cancer cell migration to bone-conditioned versus control medium was observed in 12/14 specimens (P = .0014) and correlated significantly with increasing levels of the adipokines/cytokines leptin (P = .006) and IL-1β (P = .001) in univariate and multivariate regression analyses. Fluorescence microscopy and immunohistochemistry of fragments underscored the extreme adiposity of adult human bone tissues and revealed extensive breast cancer cell colonization within the marrow adipose tissue compartment. CONCLUSIONS: Our results show that breast cancer cells migrate to human bone tissue-conditioned medium in association with increasing levels of leptin and IL-1β, and colonize the bone marrow adipose tissue compartment of cultured fragments. Bone marrow adipose tissue and its molecular signals may be important but understudied components of the breast cancer metastatic niche. PMID:26696367

  1. Human Circulating and Tissue-Resident CD56bright Natural Killer Cell Populations

    PubMed Central

    Melsen, Janine E.; Lugthart, Gertjan; Lankester, Arjan C.; Schilham, Marco W.

    2016-01-01

    Two human natural killer (NK) cell subsets are usually distinguished, displaying the CD56dimCD16+ and the CD56brightCD16−/+ phenotype. This distinction is based on NK cells present in blood, where the CD56dim NK cells predominate. However, CD56bright NK cells outnumber CD56dim NK cells in the human body due to the fact that they are predominant in peripheral and lymphoid tissues. Interestingly, within the total CD56bright NK cell compartment, a major phenotypical and functional diversity is observed, as demonstrated by the discovery of tissue-resident CD56bright NK cells in the uterus, liver, and lymphoid tissues. Uterus-resident CD56bright NK cells express CD49a while the liver- and lymphoid tissue-resident CD56bright NK cells are characterized by co-expression of CD69 and CXCR6. Tissue-resident CD56bright NK cells have a low natural cytotoxicity and produce little interferon-γ upon monokine stimulation. Their distribution and specific phenotype suggest that the tissue-resident CD56bright NK cells exert tissue-specific functions. In this review, we examine the CD56bright NK cell diversity by discussing the distribution, phenotype, and function of circulating and tissue-resident CD56bright NK cells. In addition, we address the ongoing debate concerning the developmental relationship between circulating CD56bright and CD56dim NK cells and speculate on the position of tissue-resident CD56bright NK cells. We conclude that distinguishing tissue-resident CD56bright NK cells from circulating CD56bright NK cells is a prerequisite for the better understanding of the specific role of CD56bright NK cells in the complex process of human immune regulation. PMID:27446091

  2. Lack of tissue renewal in human adult Achilles tendon is revealed by nuclear bomb 14C

    PubMed Central

    Heinemeier, Katja Maria; Schjerling, Peter; Heinemeier, Jan; Magnusson, Stig Peter; Kjaer, Michael

    2013-01-01

    Tendons are often injured and heal poorly. Whether this is caused by a slow tissue turnover is unknown, since existing data provide diverging estimates of tendon protein half-life that range from 2 mo to 200 yr. With the purpose of determining life-long turnover of human tendon tissue, we used the 14C bomb-pulse method. This method takes advantage of the dramatic increase in atmospheric levels of 14C, produced by nuclear bomb tests in 1955–1963, which is reflected in all living organisms. Levels of 14C were measured in 28 forensic samples of Achilles tendon core and 4 skeletal muscle samples (donor birth years 1945–1983) with accelerator mass spectrometry (AMS) and compared to known atmospheric levels to estimate tissue turnover. We found that Achilles tendon tissue retained levels of 14C corresponding to atmospheric levels several decades before tissue sampling, demonstrating a very limited tissue turnover. The tendon concentrations of 14C approximately reflected the atmospheric levels present during the first 17 yr of life, indicating that the tendon core is formed during height growth and is essentially not renewed thereafter. In contrast, 14C levels in muscle indicated continuous turnover. Our observation provides a fundamental premise for understanding tendon function and pathology, and likely explains the poor regenerative capacity of tendon tissue.—Heinemeier, K. M., Schjerling, P., Heinemeier, J., Magnusson, S. P., Kjaer, M. Lack of tissue renewal in human adult Achilles tendon is revealed by nuclear bomb 14C. PMID:23401563

  3. A convex optimization approach for identification of human tissue-specific interactomes

    PubMed Central

    Mohammadi, Shahin; Grama, Ananth

    2016-01-01

    Motivation: Analysis of organism-specific interactomes has yielded novel insights into cellular function and coordination, understanding of pathology, and identification of markers and drug targets. Genes, however, can exhibit varying levels of cell type specificity in their expression, and their coordinated expression manifests in tissue-specific function and pathology. Tissue-specific/tissue-selective interaction mechanisms have significant applications in drug discovery, as they are more likely to reveal drug targets. Furthermore, tissue-specific transcription factors (tsTFs) are significantly implicated in human disease, including cancers. Finally, disease genes and protein complexes have the tendency to be differentially expressed in tissues in which defects cause pathology. These observations motivate the construction of refined tissue-specific interactomes from organism-specific interactomes. Results: We present a novel technique for constructing human tissue-specific interactomes. Using a variety of validation tests (Edge Set Enrichment Analysis, Gene Ontology Enrichment, Disease-Gene Subnetwork Compactness), we show that our proposed approach significantly outperforms state-of-the-art techniques. Finally, using case studies of Alzheimer’s and Parkinson’s diseases, we show that tissue-specific interactomes derived from our study can be used to construct pathways implicated in pathology and demonstrate the use of these pathways in identifying novel targets. Availability and implementation: http://www.cs.purdue.edu/homes/mohammas/projects/ActPro.html Contact: mohammadi@purdue.edu PMID:27307623

  4. A 3D bioprinting system to produce human-scale tissue constructs with structural integrity.

    PubMed

    Kang, Hyun-Wook; Lee, Sang Jin; Ko, In Kap; Kengla, Carlos; Yoo, James J; Atala, Anthony

    2016-03-01

    A challenge for tissue engineering is producing three-dimensional (3D), vascularized cellular constructs of clinically relevant size, shape and structural integrity. We present an integrated tissue-organ printer (ITOP) that can fabricate stable, human-scale tissue constructs of any shape. Mechanical stability is achieved by printing cell-laden hydrogels together with biodegradable polymers in integrated patterns and anchored on sacrificial hydrogels. The correct shape of the tissue construct is achieved by representing clinical imaging data as a computer model of the anatomical defect and translating the model into a program that controls the motions of the printer nozzles, which dispense cells to discrete locations. The incorporation of microchannels into the tissue constructs facilitates diffusion of nutrients to printed cells, thereby overcoming the diffusion limit of 100-200 μm for cell survival in engineered tissues. We demonstrate capabilities of the ITOP by fabricating mandible and calvarial bone, cartilage and skeletal muscle. Future development of the ITOP is being directed to the production of tissues for human applications and to the building of more complex tissues and solid organs. PMID:26878319

  5. A 3D bioprinting system to produce human-scale tissue constructs with structural integrity.

    PubMed

    Kang, Hyun-Wook; Lee, Sang Jin; Ko, In Kap; Kengla, Carlos; Yoo, James J; Atala, Anthony

    2016-03-01

    A challenge for tissue engineering is producing three-dimensional (3D), vascularized cellular constructs of clinically relevant size, shape and structural integrity. We present an integrated tissue-organ printer (ITOP) that can fabricate stable, human-scale tissue constructs of any shape. Mechanical stability is achieved by printing cell-laden hydrogels together with biodegradable polymers in integrated patterns and anchored on sacrificial hydrogels. The correct shape of the tissue construct is achieved by representing clinical imaging data as a computer model of the anatomical defect and translating the model into a program that controls the motions of the printer nozzles, which dispense cells to discrete locations. The incorporation of microchannels into the tissue constructs facilitates diffusion of nutrients to printed cells, thereby overcoming the diffusion limit of 100-200 μm for cell survival in engineered tissues. We demonstrate capabilities of the ITOP by fabricating mandible and calvarial bone, cartilage and skeletal muscle. Future development of the ITOP is being directed to the production of tissues for human applications and to the building of more complex tissues and solid organs.

  6. Inosculation and perfusion of pre-vascularized tissue patches containing aligned human microvessels after myocardial infarction.

    PubMed

    Riemenschneider, Sonja B; Mattia, Donald J; Wendel, Jacqueline S; Schaefer, Jeremy A; Ye, Lei; Guzman, Pilar A; Tranquillo, Robert T

    2016-08-01

    A major goal of tissue engineering is the creation of pre-vascularized tissues that have a high density of organized microvessels that can be rapidly perfused following implantation. This is especially critical for highly metabolic tissues like myocardium, where a thick myocardial engineered tissue would require rapid perfusion within the first several days to survive transplantation. In the present work, tissue patches containing human microvessels that were either randomly oriented or aligned were placed acutely on rat hearts post-infarction and for each case it was determined whether rapid inosculation could occur and perfusion of the patch could be maintained for 6 days in an infarct environment. Patches containing self-assembled microvessels were formed by co-entrapment of human blood outgrowth endothelial cells and human pericytes in fibrin gel. Cell-induced gel contraction was mechanically-constrained resulting in samples with high densities of microvessels that were either randomly oriented (with 420 ± 140 lumens/mm(2)) or uniaxially aligned (with 940 ± 240 lumens/mm(2)) at the time of implantation. These patches were sutured onto the epicardial surface of the hearts of athymic rats following permanent ligation of the left anterior descending artery. In both aligned and randomly oriented microvessel patches, inosculation occurred and perfusion of the transplanted human microvessels was maintained, proving the in vivo vascularization potential of these engineered tissues. No difference was found in the number of human microvessels that were perfused in the randomly oriented (111 ± 75 perfused lumens/mm(2)) and aligned (173 ± 97 perfused lumens/mm(2)) patches. Our results demonstrate that tissue patches containing a high density of either aligned or randomly oriented human pre-formed microvessels achieve rapid perfusion in the myocardial infarct environment - a necessary first-step toward the creation of a thick, perfusable heart patch.

  7. Energy absorption buildup factors of human organs and tissues at energies and penetration depths relevant for radiotherapy and diagnostics.

    PubMed

    Manohara, S R; Hanagodimath, S M; Gerward, L

    2011-11-15

    Energy absorption geometric progression (GP) fitting parameters and the corresponding buildup factors have been computed for human organs and tissues, such as adipose tissue, blood (whole), cortical bone, brain (grey/white matter), breast tissue, eye lens, lung tissue, skeletal muscle, ovary, testis, soft tissue, and soft tissue (4-component), for the photon energy range 0.015-15 MeV and for penetration depths up to 40 mfp (mean free path). The chemical composition of human organs and tissues is seen to influence the energy absorption buildup factors. It is also found that the buildup factor of human organs and tissues changes significantly with the change of incident photon energy and effective atomic number, Z(eff). These changes are due to the dominance of different photon interaction processes in different energy regions and different chemical compositions of human organs and tissues. With the proper knowledge of buildup factors of human organs and tissues, energy absorption in the human body can be carefully controlled. The present results will help in estimating safe dose levels for radiotherapy patients and also useful in diagnostics and dosimetry. The tissue-equivalent materials for skeletal muscle, adipose tissue, cortical bone, and lung tissue are also discussed. It is observed that water and MS20 are good tissue equivalent materials for skeletal muscle in the extended energy range.

  8. [Obtention of human skin sheets by means of tissue engineering].

    PubMed

    Arvelo, Francisco; Pérez, Pedro; Cotte, Carlos

    2004-01-01

    The aim of this "in vitro" study was to develop a new system for keratinocyte culture on a dermal equivalent that enables treatment of different skin injuries. The keratinocyte where obtained from primary cell cultures derived from skin biopsies, seeded over a fibrin matrix enhanced with live human fibroblast. Cells growing over the dermal equivalent, rapidly confluences and a stratified epithelium was obtained within 20-25 days culture. Detachment of composite culture from flask is a simple and quick procedure with no need for chemical or enzyme treatments. The method described provides a number of advantages which include the large expansion of keratinocyte from the primary cell cultures without the need of a feeder layer, the availability of plasma from blood banks, and the versatile and safe manipulation of composite obtained "in vitro". All these facts allow to assure that this system could result very efficient for the treatment of all type of skin injuries.

  9. Cadmium concentrations in human renal cortex tissue (necropsies)

    SciTech Connect

    Lopez-Artiguez, M.; Repetto, M.; Camean, A.; Gonzalez, G.

    1995-06-01

    Cadmium is toxic to most living organisms. It occurs as part of different types of rocks, sedimentation sludges, coals and mineral oils; in minerals, cadmium (Cd) is frequently associated with zinc. Its world wide presence and considerable industrial use has given rise to an increase in its content in trophic food chains, which contribute mainly to human exposure. Oral absorption is relatively low and is influenced by the solubility of the compound, type of diet, and individual nutritional state. Interest in Cd contamination began after the outbreak of itai-itai disease in Japan. Evaluation of Cd contamination has been carried out in all the countries of the European Economic Community, and it has been estimated that in Spain emissions to the atmosphere and water are respectively 6.89 and 3.79% of total emissions in the European Communities. After exposure, the kidney is the organ which contains the highest concentrations of the Cd and retains it longest. When critical body concentration is reached, renal malfunction and damage are produced. Moreover, studies on humans not occupationally exposed to Cd show 50% of the body burden is found in the kidneys. Cd in the renal cortex increases with age, reaching a maximum between 40-60 years. Differences found among populations have been associated with daily intake in the diet and smoking habits. Taking into consideration the lack of studies on factors influencing the Cd burden in renal cortex in our country, the aim of the present paper was to find out the levels of Cd in renal cortex samples obtained from necropsies of inhabitants of Andalusia, Spain, and compare them with levels in other populations not occupationally exposed to the element; also to investigate the influence of individual factors, such as sex, age and drug addition on said Cd levels. 21 refs., 1 fig., 2 tabs.

  10. Calprotectin is released from human skeletal muscle tissue during exercise

    PubMed Central

    Mortensen, Ole Hartvig; Andersen, Kasper; Fischer, Christian; Nielsen, Anders Rinnov; Nielsen, Søren; Åkerström, Thorbjörn; Aastrøm, Maj-brit; Borup, Rehannah; Pedersen, Bente Klarlund

    2008-01-01

    Skeletal muscle has been identified as a secretory organ. We hypothesized that IL-6, a cytokine secreted from skeletal muscle during exercise, could induce production of other secreted factors in skeletal muscle. IL-6 was infused for 3 h into healthy young males (n = 7) and muscle biopsies obtained at time points 0, 3 and 6 h in these individuals and in resting controls. Affymetrix microarray analysis of gene expression changes in skeletal muscle biopsies identified a small set of genes changed by IL-6 infusion. RT-PCR validation confirmed that S100A8 and S100A9 mRNA were up-regulated 3-fold in skeletal muscle following IL-6 infusion compared to controls. Furthermore, S100A8 and S100A9 mRNA levels were up-regulated 5-fold in human skeletal muscle following cycle ergometer exercise for 3 h at ∼60% of in young healthy males (n = 8). S100A8 and S100A9 form calprotectin, which is known as an acute phase reactant. Plasma calprotectin increased 5-fold following acute cycle ergometer exercise in humans, but not following IL-6 infusion. To identify the source of calprotectin, healthy males (n = 7) performed two-legged dynamic knee extensor exercise for 3 h with a work load of ∼50% of peak power output and arterial–femoral venous differences were obtained. Arterial plasma concentrations for calprotectin increased 2-fold compared to rest and there was a net release of calprotectin from the working muscle. In conclusion, IL-6 infusion and muscle contractions induce expression of S100A8 and S100A9 in skeletal muscle. However, IL-6 alone is not a sufficient stimulus to facilitate release of calprotectin from skeletal muscle. PMID:18511485

  11. Bioceramic-collagen scaffolds loaded with human adipose-tissue derived stem cells for bone tissue engineering.

    PubMed

    Daei-Farshbaf, Neda; Ardeshirylajimi, Abdolreza; Seyedjafari, Ehsan; Piryaei, Abbas; Fadaei Fathabady, Fatemeh; Hedayati, Mehdi; Salehi, Mohammad; Soleimani, Masoud; Nazarian, Hamid; Moradi, Sadegh-Lotfalah; Norouzian, Mohsen

    2014-02-01

    The combination of bioceramics and stem cells has attracted the interest of research community for bone tissue engineering applications. In the present study, a combination of Bio-Oss(®) and type 1 collagen gel as scaffold were loaded with human adipose-tissue derived mesenchymal stem cells (AT-MSCs) after isolation and characterization, and the capacity of them for bone regeneration was investigated in rat critical size defects using digital mammography, multi-slice spiral computed tomography imaging and histological analysis. 8 weeks after implantation, no mortality or sign of inflammation was observed in the site of defect. According to the results of imaging analysis, a higher level of bone regeneration was observed in the rats receiving Bio-Oss(®)-Gel compared to untreated group. In addition, MSC-seeded Bio-Oss-Gel induced the highest bone reconstruction among all groups. Histological staining confirmed these findings and impressive osseointegration was observed in MSC-seeded Bio-Oss-Gel compared with Bio-Oss-Gel. On the whole, it was demonstrated that combination of AT-MSCs, Bio-Oss and Gel synergistically enhanced bone regeneration and reconstruction and also could serve as an appropriate structure to bone regenerative medicine and tissue engineering application.

  12. Computational Analyses of Simple Sequence Repeats on Human Tissue Specific Genes Promoters

    NASA Astrophysics Data System (ADS)

    FeiFei, Zhao; XiuJun, Gong; XinMi, Liu; LiFeng, Dong

    Promoter region of gene closely related with tissue specific expression and SSRs (simple sequence repeats) have been shown to have a variety of effects on an organism. This paper used a heuristic method to find SSRs and compared the most frequently SSRs on promoter region of both human tissues specific genes and human housekeeping genes. We used kidney and testis tissue as examples to show the final results. Especially, we found that (AGG)n is kidney specific SSR and (GCG)n is testis specific SSR. We also analyzed the SSRs frequency density distribution on different promoter regions of both tissue specific genes and housekeeping genes, and we found the density of housekeeping genes on core-promoter region is much higher than on other promoter regions.

  13. Bystander CD4+ T lymphocytes survive in HIV-infected human lymphoid tissue

    NASA Technical Reports Server (NTRS)

    Grivel, Jean-Charles; Biancotto, Angelique; Ito, Yoshinori; Lima, Rosangela G.; Margolis, Leonid B.

    2003-01-01

    HIV infection is associated with depletion of CD4(+) T cells. The mechanisms of this phenomenon remain to be understood. In particular, it remains controversial whether and to what extent uninfected ("bystander") CD4(+) T cells die in HIV-infected individuals. We address this question using a system of human lymphoid tissue ex vivo. Tissue blocks were inoculated with HIV-1. After productive infection was established, they were treated with the reverse transcriptase inhibitor nevirapine to protect from infection those CD4(+) T cells that had not yet been infected. These CD4(+) T cells residing in HIV-infected tissue are by definition bystanders. Our results demonstrate that after nevirapine application the number of bystander CD4(+) T cells is conserved. Thus, in the context of HIV-infected human lymphoid tissue, productive HIV infection kills infected cells but is not sufficient to cause the death of a significant number of uninfected CD4(+) T cells.

  14. Mobile encapsulated adipose tissue (MEAT) of cows and humans: a distinct nonneoplastic entity.

    PubMed

    Burgdorf, Walter H C; Hurt, Mark A

    2011-10-01

    Mobile encapsulated adipose tissue can be found in both the subcutis and peritoneal cavity. The cutaneous lesions are more common and better described; they are usually designated as "mobile encapsulated lipoma" or "nodular-cystic fat necrosis." The clinical name of abacus tumor describes best the small marble-like nodules that often can be moved freely through the subcutaneous tissue planes. Histopathologically, the nodules are composed of a dense fibrous capsule surrounding fat that may show varying degrees of necrosis, calcification, and lipomembranous changes. The peritoneal nodules are thought to originate from detached epiploic appendices. They are described occasionally in humans but more commonly in cows. Because these bovine mobile peritoneal bodies are clinically and histopathologically identical to the nodules in human subcutaneous tissue, the authors suggest that one can better understand both processes by comparing them. Because the lesions are reactive, not neoplastic, and necrosis is not an invariable feature, the authors suggest the neutral designation of mobile encapsulated adipose tissue.

  15. Thorium-232 in human tissues: Metabolic parameters and radiation doses

    SciTech Connect

    Stehney, A.F.

    1994-09-01

    Higher than environmental levels of {sup 232}Th have been found in autopsy samples of lungs and other organs from four former employees of a Th refinery. Working periods of the subjects ranged from 3 to 24 years, and times from end of work to death ranged from 6 to 31 years. Concentrations of {sup 232}Th in these samples and in tissues from two cases of non-occupational exposure were examined for compatibility with dosimetric models in Publication 30 of the International Commission on Radiological Protection (ICPP 1979a). The concentrations of {sup 232}Th in the lungs of the Th workers relative to the concentrations in bone or liver were much higher than calculated from the model for class Y aerosols of Th and the exposure histories of the subjects, and concentrations in the pulmonary lymph nodes were much lower than calculated for three of the Th workers and both non-occupational cases. Least-squares fits to the measured concentrations showed that the biological half-times of Th in liver, spleen, and kidneys are similar to the half-time in bone instead of the factor of 10 less suggested in Publication 30, and the fractions translocated from body fluids were found to be about 0.03, 0.02, and 0.005, respectively, when the fraction to bone was held at the suggested value of 0.7. Fitted values of the respiratory parameters differed significantly between cases and the differences were ascribable to aerosol differences. Average inhalation rates calculated for individual Th workers ranged from 50 to 110 Bq {sup 232}Th y{sup {minus}1}, and dose equivalents as high as 9.3 Sv to the lungs, 2.0 Sv to bone surfaces, and 1.1 Sv effective dose equivalent were calculated from the inhalation rates and fitted values of the metabolic parameters. The radiation doses were about the same when calculated from parameter values fitted with an assumed translocation fraction of 0.2 from body fluids to bone instead of 0.7.

  16. Molecular cloning and synthesis of biologically active human tissue inhibitor of metalloproteinases in yeast

    SciTech Connect

    Kaczorek, M.; Honore, N.; Ribes, V.; Dehoux, P.; Cornet, P.; Cartwright, T.; Streeck, R.E.

    1987-06-01

    Tissue inhibitor of metalloproteinases (TIMP) is a widely distributed glycoprotein that stochiometrically inactivates metalloproteinases involved in connective tissue catabolism. Here they report the cDNA cloning of TIMP from human fibroblastic MRC5 cells using a single 42-base oligonucleotide probe. Expression in S. cerevisiae of complete TIMP cDNA yielded insoluble protein aggregates. Biologically active TIMP was reconstituted from the yeast product by a denaturation/renaturation procedure.

  17. Tissue culture system for infection with human hepatitis delta virus.

    PubMed Central

    Sureau, C; Jacob, J R; Eichberg, J W; Lanford, R E

    1991-01-01

    An in vitro culture system was developed for assaying the infectivity of the human hepatitis delta virus (HDV). Hepatocytes were isolated from chimpanzee liver and grown in a serum-free medium. Cells were shown to be infectible by HDV and to remain susceptible to infection for at least 3 weeks in culture, as evidenced by the appearance of RNA species characteristic of HDV replication as early as 6 days postinfection. When repeated experiments were carried out on cells derived from an animal free of hepatitis B virus (HBV), HDV infection occurred in a consistent fashion but there was no indication of infection with the HBV that was present in the inoculum. Despite numerous attempts with different sources of HBV inocula free of HDV, there was no evidence that indicated susceptibility of these cells to HBV infection. This observation may indicate that HBV and HDV use different modes of entry into hepatocytes. When cells derived from an HBV-infected animal were exposed to HDV, synthesis and release of progeny HDV particles were obtained in addition to HBV replication and production of Dane particles. Although not infectible with HBV, primary cultures of chimpanzee hepatocytes are capable of supporting part of the life cycle of HBV and the entire life cycle of HDV. Images PMID:2041075

  18. Divergent viral presentation among human tumors and adjacent normal tissues

    PubMed Central

    Cao, Song; Wendl, Michael C.; Wyczalkowski, Matthew A.; Wylie, Kristine; Ye, Kai; Jayasinghe, Reyka; Xie, Mingchao; Wu, Song; Niu, Beifang; Grubb, Robert; Johnson, Kimberly J.; Gay, Hiram; Chen, Ken; Rader, Janet S.; Dipersio, John F.; Chen, Feng; Ding, Li

    2016-01-01

    We applied a newly developed bioinformatics system called VirusScan to investigate the viral basis of 6,813 human tumors and 559 adjacent normal samples across 23 cancer types and identified 505 virus positive samples with distinctive, organ system- and cancer type-specific distributions. We found that herpes viruses (e.g., subtypes HHV4, HHV5, and HHV6) that are highly prevalent across cancers of the digestive tract showed significantly higher abundances in tumor versus adjacent normal samples, supporting their association with these cancers. We also found three HPV16-positive samples in brain lower grade glioma (LGG). Further, recurrent HBV integration at the KMT2B locus is present in three liver tumors, but absent in their matched adjacent normal samples, indicating that viral integration induced host driver genetic alterations are required on top of viral oncogene expression for initiation and progression of liver hepatocellular carcinoma. Notably, viral integrations were found in many genes, including novel recurrent HPV integrations at PTPN13 in cervical cancer. Finally, we observed a set of HHV4 and HBV variants strongly associated with ethnic groups, likely due to viral sequence evolution under environmental influences. These findings provide important new insights into viral roles of tumor initiation and progression and potential new therapeutic targets. PMID:27339696

  19. Autofluorescence multiphoton microscopy for visualization of tissue morphology and cellular dynamics in murine and human airways

    PubMed Central

    Kretschmer, Sarah; Pieper, Mario; Hüttmann, Gereon; Bölke, Torsten; Wollenberg, Barbara; Marsh, Leigh M; Garn, Holger; König, Peter

    2016-01-01

    The basic understanding of inflammatory airway diseases greatly benefits from imaging the cellular dynamics of immune cells. Current imaging approaches focus on labeling specific cells to follow their dynamics but fail to visualize the surrounding tissue. To overcome this problem, we evaluated autofluorescence multiphoton microscopy for following the motion and interaction of cells in the airways in the context of tissue morphology. Freshly isolated murine tracheae from healthy mice and mice with experimental allergic airway inflammation were examined by autofluorescence multiphoton microscopy. In addition, fluorescently labeled ovalbumin and fluorophore-labeled antibodies were applied to visualize antigen uptake and to identify specific cell populations, respectively. The trachea in living mice was imaged to verify that the ex vivo preparation reflects the in vivo situation. Autofluorescence multiphoton microscopy was also tested to examine human tissue from patients in short-term tissue culture. Using autofluorescence, the epithelium, underlying cells, and fibers of the connective tissue, as well as blood vessels, were identified in isolated tracheae. Similar structures were visualized in living mice and in the human airway tissue. In explanted murine airways, mobile cells were localized within the tissue and we could follow their migration, interactions between individual cells, and their phagocytic activity. During allergic airway inflammation, increased number of eosinophil and neutrophil granulocytes were detected that moved within the connective tissue and immediately below the epithelium without damaging the epithelial cells or connective tissues. Contacts between granulocytes were transient lasting 3 min on average. Unexpectedly, prolonged interactions between granulocytes and antigen-uptaking cells were observed lasting for an average of 13 min. Our results indicate that autofluorescence-based imaging can detect previously unknown immune cell

  20. Autofluorescence multiphoton microscopy for visualization of tissue morphology and cellular dynamics in murine and human airways.

    PubMed

    Kretschmer, Sarah; Pieper, Mario; Hüttmann, Gereon; Bölke, Torsten; Wollenberg, Barbara; Marsh, Leigh M; Garn, Holger; König, Peter

    2016-08-01

    The basic understanding of inflammatory airway diseases greatly benefits from imaging the cellular dynamics of immune cells. Current imaging approaches focus on labeling specific cells to follow their dynamics but fail to visualize the surrounding tissue. To overcome this problem, we evaluated autofluorescence multiphoton microscopy for following the motion and interaction of cells in the airways in the context of tissue morphology. Freshly isolated murine tracheae from healthy mice and mice with experimental allergic airway inflammation were examined by autofluorescence multiphoton microscopy. In addition, fluorescently labeled ovalbumin and fluorophore-labeled antibodies were applied to visualize antigen uptake and to identify specific cell populations, respectively. The trachea in living mice was imaged to verify that the ex vivo preparation reflects the in vivo situation. Autofluorescence multiphoton microscopy was also tested to examine human tissue from patients in short-term tissue culture. Using autofluorescence, the epithelium, underlying cells, and fibers of the connective tissue, as well as blood vessels, were identified in isolated tracheae. Similar structures were visualized in living mice and in the human airway tissue. In explanted murine airways, mobile cells were localized within the tissue and we could follow their migration, interactions between individual cells, and their phagocytic activity. During allergic airway inflammation, increased number of eosinophil and neutrophil granulocytes were detected that moved within the connective tissue and immediately below the epithelium without damaging the epithelial cells or connective tissues. Contacts between granulocytes were transient lasting 3 min on average. Unexpectedly, prolonged interactions between granulocytes and antigen-uptaking cells were observed lasting for an average of 13 min. Our results indicate that autofluorescence-based imaging can detect previously unknown immune cell

  1. Characterization and assessment of hyperelastic and elastic properties of decellularized human adipose tissues.

    PubMed

    Omidi, Ehsan; Fuetterer, Lydia; Reza Mousavi, Seyed; Armstrong, Ryan C; Flynn, Lauren E; Samani, Abbas

    2014-11-28

    Decellularized adipose tissue (DAT) has shown potential as a regenerative scaffold for plastic and reconstructive surgery to augment or replace damaged or missing adipose tissue (e.g. following lumpectomy or mastectomy). The mechanical properties of soft tissue substitutes are of paramount importance in restoring the natural shape and appearance of the affected tissues, and mechanical mismatching can lead to unpredictable scar tissue formation and poor implant integration. The goal of this work was to assess the linear elastic and hyperelastic properties of decellularized human adipose tissue and compare them to those of normal breast adipose tissue. To assess the influence of the adipose depot source on the mechanical properties of the resultant decellularized scaffolds, we performed indentation tests on DAT samples sourced from adipose tissue isolated from the breast, subcutaneous abdominal region, omentum, pericardial depot and thymic remnant, and their corresponding force-displacement data were acquired. Elastic and hyperelastic parameters were estimated using inverse finite element algorithms. Subsequently, a simulation was conducted in which the estimated hyperelastic parameters were tested in a real human breast model under gravity loading in order to assess the suitability of the scaffolds for implantation. Results of these tests showed that in the human breast, the DAT would show similar deformability to that of native normal tissue. Using the measured hyperelastic parameters, we were able to assess whether DAT derived from different depots exhibited different intrinsic nonlinearities. Results showed that DAT sourced from varying regions of the body exhibited little intrinsic nonlinearity, with no statistically significant differences between the groups.

  2. Allelic imbalance of tissue-type plasminogen activator (t-PA) gene expression in human brain tissue.

    PubMed

    Tjarnlund-Wolf, A; Hultman, K; Curtis, M A; Faull, R L M; Medcalf, R L; Jern, C

    2011-06-01

    We have identified a single-nucleotide polymorphism (SNP) in the t-PA enhancer (-7351C>T), which is associated with endothelial t-PA release in vivo. In vitro studies demonstrated that this SNP is functional at the level of transcription. In the brain, t-PA has been implicated in both physiologic and pathophysiologic processes. The aim of the present study was to examine the effect of the t-PA -7351C>T SNP on t-PA gene expression in human brain tissue. Allelic mRNA expression was measured in heterozygous post-mortem brain tissues using quantitative TaqMan genotyping assay. Protein-DNA interactions were assessed using electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). Significantly higher levels of t-PA mRNA were generated from chromosomes that harboured the wild-type -7351C allele, as compared to those generated from the mutant T allele (for the hippocampus, C to T allelic ratio of ~1.3, p=0.010, n=12; and for the cortex, C to T allelic ratio of ~1.2, p=0.017, n=12). EMSA showed reduced neuronal and astrocytic nuclear protein binding affinity to the T allele, and identified Sp1 and Sp3 as the major transcription factors that bound to the -7351 site. ChIP analyses confirmed that Sp1 recognises this site in intact cells. In conclusion, the t-PA -7351C>T SNP affects t-PA gene expression in human brain tissue. This finding might have clinical implications for neurological conditions associated with enhanced t-PA levels, such as in the acute phase of cerebral ischaemia, and also for stroke recovery.

  3. Adipose tissue glycogen accumulation is associated with obesity-linked inflammation in humans

    PubMed Central

    Ceperuelo-Mallafré, Victòria; Ejarque, Miriam; Serena, Carolina; Duran, Xavier; Montori-Grau, Marta; Rodríguez, Miguel Angel; Yanes, Oscar; Núñez-Roa, Catalina; Roche, Kelly; Puthanveetil, Prasanth; Garrido-Sánchez, Lourdes; Saez, Enrique; Tinahones, Francisco J.; Garcia-Roves, Pablo M.; Gómez-Foix, Anna Ma; Saltiel, Alan R.; Vendrell, Joan; Fernández-Veledo, Sonia

    2015-01-01

    Objective Glycogen metabolism has emerged as a mediator in the control of energy homeostasis and studies in murine models reveal that adipose tissue might contain glycogen stores. Here we investigated the physio(patho)logical role of glycogen in human adipose tissue in the context of obesity and insulin resistance. Methods We studied glucose metabolic flux of hypoxic human adipoctyes by nuclear magnetic resonance and mass spectrometry-based metabolic approaches. Glycogen synthesis and glycogen content in response to hypoxia was analyzed in human adipocytes and macrophages. To explore the metabolic effects of enforced glycogen deposition in adipocytes and macrophages, we overexpressed PTG, the only glycogen-associated regulatory subunit (PP1-GTS) reported in murine adipocytes. Adipose tissue gene expression analysis was performed on wild type and homozygous PTG KO male mice. Finally, glycogen metabolism gene expression and glycogen accumulation was analyzed in adipose tissue, mature adipocytes and resident macrophages from lean and obese subjects with different degrees of insulin resistance in 2 independent cohorts. Results We show that hypoxia modulates glucose metabolic flux in human adipocytes and macrophages and promotes glycogenesis. Enforced glycogen deposition by overexpression of PTG re-orients adipocyte secretion to a pro-inflammatory response linked to insulin resistance and monocyte/lymphocyte migration. Furthermore, glycogen accumulation is associated with inhibition of mTORC1 signaling and increased basal autophagy flux, correlating with greater leptin release in glycogen-loaded adipocytes. PTG-KO mice have reduced expression of key inflammatory genes in adipose tissue and PTG overexpression in M0 macrophages induces a pro-inflammatory and glycolytic M1 phenotype. Increased glycogen synthase expression correlates with glycogen deposition in subcutaneous adipose tissue of obese patients. Glycogen content in subcutaneous mature adipocytes is associated

  4. Proteomic Analysis of Human Brown Adipose Tissue Reveals Utilization of Coupled and Uncoupled Energy Expenditure Pathways.

    PubMed

    Müller, Sebastian; Balaz, Miroslav; Stefanicka, Patrik; Varga, Lukas; Amri, Ez-Zoubir; Ukropec, Jozef; Wollscheid, Bernd; Wolfrum, Christian

    2016-01-01

    Human brown adipose tissue (BAT) has become an attractive target to combat the current epidemical spread of obesity and its associated co-morbidities. Currently, information on its functional role is primarily derived from rodent studies. Here, we present the first comparative proteotype analysis of primary human brown adipose tissue versus adjacent white adipose tissue, which reveals significant quantitative differences in protein abundances and in turn differential functional capabilities. The majority of the 318 proteins with increased abundance in BAT are associated with mitochondrial metabolism and confirm the increased oxidative capacity. In addition to uncoupling protein 1 (UCP1), the main functional effector for uncoupled respiration, we also detected the mitochondrial creatine kinases (CKMT1A/B, CKMT2), as effective modulators of ATP synthase coupled respiration, to be exclusively expressed in BAT. The abundant expression and utilization of both energy expenditure pathways in parallel highlights the complex functional involvement of BAT in human physiology. PMID:27418403

  5. Proteomic Analysis of Human Brown Adipose Tissue Reveals Utilization of Coupled and Uncoupled Energy Expenditure Pathways

    PubMed Central

    Müller, Sebastian; Balaz, Miroslav; Stefanicka, Patrik; Varga, Lukas; Amri, Ez-Zoubir; Ukropec, Jozef; Wollscheid, Bernd; Wolfrum, Christian

    2016-01-01

    Human brown adipose tissue (BAT) has become an attractive target to combat the current epidemical spread of obesity and its associated co-morbidities. Currently, information on its functional role is primarily derived from rodent studies. Here, we present the first comparative proteotype analysis of primary human brown adipose tissue versus adjacent white adipose tissue, which reveals significant quantitative differences in protein abundances and in turn differential functional capabilities. The majority of the 318 proteins with increased abundance in BAT are associated with mitochondrial metabolism and confirm the increased oxidative capacity. In addition to uncoupling protein 1 (UCP1), the main functional effector for uncoupled respiration, we also detected the mitochondrial creatine kinases (CKMT1A/B, CKMT2), as effective modulators of ATP synthase coupled respiration, to be exclusively expressed in BAT. The abundant expression and utilization of both energy expenditure pathways in parallel highlights the complex functional involvement of BAT in human physiology. PMID:27418403

  6. Advanced Imaging and Tissue Engineering of the Human Limbal Epithelial Stem Cell Niche

    PubMed Central

    Massie, Isobel; Dziasko, Marc; Kureshi, Alvena; Levis, Hannah J.; Morgan, Louise; Neale, Michael; Sheth, Radhika; Tovell, Victoria E.; Vernon, Amanda J.; Funderburgh, James L.; Daniels, Julie T.

    2015-01-01

    The limbal epithelial stem cell niche provides a unique, physically protective environment in which limbal epithelial stem cells reside in close proximity with accessory cell types and their secreted factors. The use of advanced imaging techniques is described to visualize the niche in three dimensions in native human corneal tissue. In addition, a protocol is provided for the isolation and culture of three different cell types, including human limbal epithelial stem cells from the limbal niche of human donor tissue. Finally, the process of incorporating these cells within plastic compressed collagen constructs to form a tissue-engineered corneal limbus is described and how immunohistochemical techniques may be applied to characterize cell phenotype therein. PMID:25388395

  7. Proteomic Analysis of Human Brown Adipose Tissue Reveals Utilization of Coupled and Uncoupled Energy Expenditure Pathways.

    PubMed

    Müller, Sebastian; Balaz, Miroslav; Stefanicka, Patrik; Varga, Lukas; Amri, Ez-Zoubir; Ukropec, Jozef; Wollscheid, Bernd; Wolfrum, Christian

    2016-07-15

    Human brown adipose tissue (BAT) has become an attractive target to combat the current epidemical spread of obesity and its associated co-morbidities. Currently, information on its functional role is primarily derived from rodent studies. Here, we present the first comparative proteotype analysis of primary human brown adipose tissue versus adjacent white adipose tissue, which reveals significant quantitative differences in protein abundances and in turn differential functional capabilities. The majority of the 318 proteins with increased abundance in BAT are associated with mitochondrial metabolism and confirm the increased oxidative capacity. In addition to uncoupling protein 1 (UCP1), the main functional effector for uncoupled respiration, we also detected the mitochondrial creatine kinases (CKMT1A/B, CKMT2), as effective modulators of ATP synthase coupled respiration, to be exclusively expressed in BAT. The abundant expression and utilization of both energy expenditure pathways in parallel highlights the complex functional involvement of BAT in human physiology.

  8. "The state of the heart": Recent advances in engineering human cardiac tissue from pluripotent stem cells.

    PubMed

    Sirabella, Dario; Cimetta, Elisa; Vunjak-Novakovic, Gordana

    2015-08-01

    The pressing need for effective cell therapy for the heart has led to the investigation of suitable cell sources for tissue replacement. In recent years, human pluripotent stem cell research expanded tremendously, in particular since the derivation of human-induced pluripotent stem cells. In parallel, bioengineering technologies have led to novel approaches for in vitro cell culture. The combination of these two fields holds potential for in vitro generation of high-fidelity heart tissue, both for basic research and for therapeutic applications. However, this new multidisciplinary science is still at an early stage. Many questions need to be answered and improvements need to be made before clinical applications become a reality. Here we discuss the current status of human stem cell differentiation into cardiomyocytes and the combined use of bioengineering approaches for cardiac tissue formation and maturation in developmental studies, disease modeling, drug testing, and regenerative medicine.

  9. Brown Adipose Tissue Activation Is Linked to Distinct Systemic Effects on Lipid Metabolism in Humans.

    PubMed

    Chondronikola, Maria; Volpi, Elena; Børsheim, Elisabet; Porter, Craig; Saraf, Manish K; Annamalai, Palam; Yfanti, Christina; Chao, Tony; Wong, Daniel; Shinoda, Kosaku; Labbė, Sebastien M; Hurren, Nicholas M; Cesani, Fernardo; Kajimura, Shingo; Sidossis, Labros S

    2016-06-14

    Recent studies suggest that brown adipose tissue (BAT) plays a role in energy and glucose metabolism in humans. However, the physiological significance of human BAT in lipid metabolism remains unknown. We studied 16 overweight/obese men during prolonged, non-shivering cold and thermoneutral conditions using stable isotopic tracer methodologies in conjunction with hyperinsulinemic-euglycemic clamps and BAT and white adipose tissue (WAT) biopsies. BAT volume was significantly associated with increased whole-body lipolysis, triglyceride-free fatty acid (FFA) cycling, FFA oxidation, and adipose tissue insulin sensitivity. Functional analysis of BAT and WAT demonstrated the greater thermogenic capacity of BAT compared to WAT, while molecular analysis revealed a cold-induced upregulation of genes involved in lipid metabolism only in BAT. The accelerated mobilization and oxidation of lipids upon BAT activation supports a putative role for BAT in the regulation of lipid metabolism in humans. PMID:27238638

  10. An experimental model of tool mark striations by a serrated blade in human soft tissues.

    PubMed

    Jacques, Rebekah; Kogon, Stanley; Shkrum, Michael

    2014-03-01

    Tool mark analysis is a method of matching a weapon with the injury it caused. In a homicidal stabbing using a serrated knife, a stab wound that involves a cartilage may leave striations from the serration points on the blade edge. Assessing tissue striations is a means of identifying the weapon as having a serrated blade. This prospective study examines the possibility that similar striations may be produced in human soft tissues. Using tissues taken at the time of hospital-consented autopsies, stab wound tracks were assessed in a variety of human tissues (aorta, skin, liver, kidney, and cardiac and skeletal muscle). Stab wounds were produced postmortem with similar serrated and smooth-edged blades. The walls of the stab wounds were exposed, documented by photography and cast with dental impression material. Striations were identified by naked-eye examination in the skin and aorta. Photodocumentation of fresh tissue was best achieved in the aorta. Striations were not identified in wound tracks produced by the smooth-edged blade. Three blinded forensic pathologists were assessed for their ability to detect striations in photographs of wound tracks and had substantial interobserver agreement (κ = 0.76) identifying striations. This study demonstrates that tool mark striations can be present in some noncartilaginous human tissues.

  11. An experimental model of tool mark striations by a serrated blade in human soft tissues.

    PubMed

    Jacques, Rebekah; Kogon, Stanley; Shkrum, Michael

    2014-03-01

    Tool mark analysis is a method of matching a weapon with the injury it caused. In a homicidal stabbing using a serrated knife, a stab wound that involves a cartilage may leave striations from the serration points on the blade edge. Assessing tissue striations is a means of identifying the weapon as having a serrated blade. This prospective study examines the possibility that similar striations may be produced in human soft tissues. Using tissues taken at the time of hospital-consented autopsies, stab wound tracks were assessed in a variety of human tissues (aorta, skin, liver, kidney, and cardiac and skeletal muscle). Stab wounds were produced postmortem with similar serrated and smooth-edged blades. The walls of the stab wounds were exposed, documented by photography and cast with dental impression material. Striations were identified by naked-eye examination in the skin and aorta. Photodocumentation of fresh tissue was best achieved in the aorta. Striations were not identified in wound tracks produced by the smooth-edged blade. Three blinded forensic pathologists were assessed for their ability to detect striations in photographs of wound tracks and had substantial interobserver agreement (κ = 0.76) identifying striations. This study demonstrates that tool mark striations can be present in some noncartilaginous human tissues. PMID:24457587

  12. The fractional viscoelastic response of human breast tissue cells

    NASA Astrophysics Data System (ADS)

    Carmichael, B.; Babahosseini, H.; Mahmoodi, S. N.; Agah, M.

    2015-07-01

    The mechanical response of a living cell is notoriously complicated. The complex, heterogeneous characteristics of cellular structure introduce difficulties that simple linear models of viscoelasticity cannot overcome, particularly at deep indentation depths. Herein, a nano-scale stress-relaxation analysis performed with an atomic force microscope reveals that isolated human breast cells do not exhibit simple exponential relaxation capable of being modeled by the standard linear solid (SLS) model. Therefore, this work proposes the application of the fractional Zener (FZ) model of viscoelasticity to extract mechanical parameters from the entire relaxation response, improving upon existing physical techniques to probe isolated cells. The FZ model introduces a new parameter that describes the fractional time-derivative dependence of the response. The results show an exceptional increase in conformance to the experimental data compared to that predicted by the SLS model, and the order of the fractional derivative (α) is remarkably homogeneous across the populations, with a median value of 0.48 ± 0.06 for the malignant population and 0.51 ± 0.07 for the benign. The cells’ responses exhibit power-law behavior and complexity not associated with simple relaxation (SLS, α = 1) that supports the application of a fractional model. The distributions of some of the FZ parameters also preserve the distinction between the malignant and benign sample populations seen from the linear model and previous results while including the contribution of fast-relaxation behavior. The resulting viscosity, measured by a composite relaxation time, exhibits considerably less dispersion due to residual error than the distribution generated by the linear model and therefore serves as a more powerful marker for cell differentiation.

  13. Biorepository standards and protocols for collecting, processing, and storing human tissues.

    PubMed

    Troyer, Dean

    2008-01-01

    Recent advances in high-throughput assays for gene expression (genomics), proteins (proteomics), and metabolites (metabolomics) have engendered a parallel need for well-annotated human biological samples. Samples from both diseased and unaffected normal tissues are often required. Biorepositories consist of a specimen bank linked to a database of information. Assuring chain of custody and annotation of samples with relevant clinical information is required. The value of samples to end users is generally commensurate with the quality and extent of relevant clinical data included with the samples. Procurement of tissues is often done with parallel pre- and/or post-treatment venipuncture to obtain blood and tissue samples from the same subject. Biorepositories must also process, preserve, and distribute samples to end users. Like traditional libraries, biorepositories are meant to be used, and they are most useful when the needs of end users (researchers) are considered in the planning and development process. Ethics review and an awareness of regulatory requirements for storage, transport, and distribution are required. In the USA, Institutional Review Boards are the local regulatory entities that review protocols for banking of human biological tissues. Governmental and professional agencies and organizations provide some guidelines for standard operating procedures. The Food and Drug Administration (FDA), the Centers For Disease Control (CDC), and professional organizations such as the American Association of Tissue Banks (AATB), the American Association of Blood Banks, The International Red Cross, International Society for Biological Repositories (ISBER) and other organizations provide guidelines for biorepositories and banking of human tissues (see Table 1). To date, these guidelines are directed largely toward procurement, banking, and distribution of human tissues for therapeutic uses. In the international setting, the World Health Organization provides ethical

  14. Comparison of organochlorine residues in human adipose tissue autopsy samples from two Ontario municipalities

    SciTech Connect

    Williams, D.T.; LeBel, G.L.; Junkins, E.

    1984-01-01

    Human adipose tissue samples obtained during autopsies in a Canadian Great Lakes community, Kingston, Ontario, and a second community, Ottawa, Ontario, were analyzed for organochlorine pesticides, polychlorobiphenyls, chlorobenzenes, and chlorophenols. Significantly different levels of Dichlorodiphenyl-dichlorethane, mirex, hexachlorobenzene, and 2,3,4,6-tetrachlorophenol were found in Kingston adipose tissues compared to Ottawa tissues. Residue levels of oxychlordane, mirex, and polychlorinated biphenyls were significantly different in Kingston males versus Kingston females. The means and ranges of residue levels were contrasted with those reported in previous Canadian surveys.

  15. Human breast tissue disposition and bioactivity of limonene in women with early-stage breast cancer.

    PubMed

    Miller, Jessica A; Lang, Julie E; Ley, Michele; Nagle, Ray; Hsu, Chiu-Hsieh; Thompson, Patricia A; Cordova, Catherine; Waer, Amy; Chow, H-H Sherry

    2013-06-01

    Limonene is a bioactive food component found in citrus peel oil that has shown chemopreventive and chemotherapeutic activities in preclinical studies. We conducted an open-label pilot clinical study to determine the human breast tissue disposition of limonene and its associated bioactivity. We recruited 43 women with newly diagnosed operable breast cancer electing to undergo surgical excision to take 2 grams of limonene daily for two to six weeks before surgery. Blood and breast tissue were collected to determine drug/metabolite concentrations and limonene-induced changes in systemic and tissue biomarkers of breast cancer risk or carcinogenesis. Limonene was found to preferentially concentrate in the breast tissue, reaching high tissue concentration (mean = 41.3 μg/g tissue), whereas the major active circulating metabolite, perillic acid, did not concentrate in the breast tissue. Limonene intervention resulted in a 22% reduction in cyclin D1 expression (P = 0.002) in tumor tissue but minimal changes in tissue Ki67 and cleaved caspase-3 expression. No significant changes in serum leptin, adiponectin, TGF-β1, insulin-like growth factor binding protein-3 (IGFBP-3), and interleukin-6 (IL-6) levels were observed following limonene intervention. There was a small but statistically significant postintervention increase in insulin-like growth factor I (IGF-I) levels. We conclude that limonene distributed extensively to human breast tissue and reduced breast tumor cyclin D1 expression that may lead to cell-cycle arrest and reduced cell proliferation. Furthermore, placebo-controlled clinical trials and translational research are warranted to establish limonene's role for breast cancer prevention or treatment.

  16. Influence of trace elements in human tissue in low-energy photon brachytherapy dosimetry

    NASA Astrophysics Data System (ADS)

    White, Shane A.; Landry, Guillaume; van Gils, Francis; Verhaegen, Frank; Reniers, Brigitte

    2012-06-01

    The aim of this paper is to determine the dosimetric impact of trace elements in human tissues for low-energy photon sources used in brachytherapy. Monte Carlo dose calculations were used to investigate the dosimetric effect of trace elements present in normal or cancerous human tissues. The effect of individual traces (atomic number Z = 11-30) was studied in soft tissue irradiated by low-energy brachytherapy sources. Three other tissue types (prostate, adipose and mammary gland) were also simulated with varying trace concentrations to quantify the contribution of each trace to the dose distribution. The dose differences between cancerous and healthy prostate tissues were calculated in single- and multi-source geometries. The presence of traces in a tissue produces a difference in the dose distribution that is dependent on Z and the concentration of the trace. Low-Z traces (Na) have a negligible effect (<0.3%) in all tissues, while higher Z (K) had a larger effect (>3%). There is a potentially significant difference in the dose distribution between cancerous and healthy prostate tissues (4%) and even larger if compared to the trace-free composition (15%) in both single- and multi-sourced geometries. Trace elements have a non-negligible (up to 8% in prostate D90) effect on the dose in tissues irradiated with low-energy photon sources. This study underlines the need for further investigation into accurate determination of the trace composition of tissues associated with low-energy brachytherapy. Alternatively, trace elements could be incorporated as a source of uncertainty in dose calculations. This work was part of an invited presentation at the ‘International Workshop on Recent Advances in Monte Carlo Techniques for Radiation Therapy’, held in Montreal, June 8-10, 2011.

  17. The appropriateness of swab cultures for the release of human allograft tissue.

    PubMed

    Ronholdt, Chad J; Bogdansky, Simon

    2005-08-01

    Surgeries utilizing human allograft tissues have increased dramatically in recent years. With this increase has come a greater reliance on the use of swab culturing to assess allograft tissues for microbial contamination prior to distribution. In contrast to the typical industrial microbiological uses for swabs, the tissue banking industry has relied on swab cultures as a sterility release method for allograft tissues. It has been reported in the literature that swabs have limitations, both in sensitivity and reproducibility, so their suitability as a final sterility release method was evaluated in this study. Two different swab-culturing systems were evaluated (COPAN, EZ Culturette) using human allograft tissues spiked with low levels of multiple bacterial and fungal microorganisms. The average microbial recoveries for all challenge microorganisms for each tissue type and each swab system were calculated. Percent recoveries for each challenge microorganism were also calculated and reported. The results indicated that both swab systems exhibited low and highly variable recoveries from the seeded allograft tissues. Further analysis indicated there was no statistical difference ( proportional, variant=0.05) between the two swab systems. It is the recommendation of the authors that swab culturing not be used to assess relatively low levels of microbial contamination on allografts. Instead, alternative validated microbial detection methods with improved sensitivity and reproducibility should be employed and validated for this critical task. PMID:15973533

  18. When fat becomes an ally of the enemy: adipose tissue as collaborator in human breast cancer.

    PubMed

    Lapeire, Lore; Denys, Hannelore; Cocquyt, Véronique; De Wever, Olivier

    2015-07-01

    Since the discovery of leptin in 1994, our vision of adipose tissue as a static organ regulating mainly lipid storage and release has been completely overthrown, and adipose tissue is now seen as an active and integral organ in human physiology. In the past years, extensive research has tremendously given us more insights in the mechanisms and pathways involved not only in normal but also in 'sick' adipose tissue, for example, in obesity and lipodystrophy. With growing evidence of a link between obesity and several types of cancer, research focusing on the interaction between adipose tissue and cancer has begun to unravel the interesting but complex multi-lateral communication between the different players. With breast cancer as one of the first cancer types where a positive correlation between obesity and breast cancer incidence and prognosis in post-menopausal women was found, we have focused this review on the paracrine and endocrine role of adipose tissue in breast cancer initiation and progression. As important inter-species differences in adipose tissue occur, we mainly selected human adipose tissue- and breast cancer-based studies with a short reflection on therapeutic possibilities. This review is part of the special issue on "Adiposopathy in Cancer and (Cardio)Metabolic Diseases".

  19. Technical report: immunofluorescence and TUNEL staining of celloidin embedded human temporal bone tissues.

    PubMed

    Markaryan, Adam; Nelson, Erik G; Tretiakova, Maria; Hinojosa, Raul

    2008-07-01

    The large archival human temporal bone collections of the world have been fixed in formalin and embedded in celloidin. These treatments have created challenges to the use of contemporary probes, which are routinely used in the evaluation of fresh and frozen tissues, for the analysis of archival temporal bone tissues. Formalin alters the configuration of proteins and can obscure antigens by modifying the epitopes recognized by antibodies. Celloidin embedding provides superior support of the delicate membranous structures of the inner ear to maintain tissue integrity during sectioning, however, inadequate removal of celloidin may limit tissue permeability resulting in poor penetration of large molecules. Methods are described in this manuscript that have allowed reproducible immunofluorescence and TUNEL (terminal deoxynucleotidyl transferase mediated dUTP nick end labeling) staining results in these archival tissues. To our knowledge, successful immunofluorescence staining of type I collagen, immunofluorescence staining of cytochrome c oxidase subunit III (COX III), and TUNEL staining in archival human temporal bone tissues with confocal microscopy has not been previously reported. These results demonstrate the utility of developing techniques to evaluate the existing collections of archival temporal bones which remain our greatest source of tissue for investigating the causes of ear diseases.

  20. Drug analysis in blowfly larvae and in human tissues: a comparative study.

    PubMed

    Campobasso, Carlo P; Gherardi, Mirella; Caligara, Marina; Sironi, Luca; Introna, Francesco

    2004-08-01

    The present study investigates the correlation between concentrations of drugs in human tissues and Diptera larvae feeding on these tissues. Samples of liver were taken from 18 cases in which preliminary toxicological screening indicated the presence of drugs. Blowfly larvae (Diptera: Calliphoridae) were reared on these samples and subsequently analyzed for drug content. Toxicological analyses were carried out using ONLINE Abuscreen (Roche) and GC/MS for available body fluids (blood, urine and bile) as well as liver samples and maggots. All drugs detected in human tissues were also detected in insect specimens. Opiates, cocaine and barbiturates as well as some antidepressants (clomipramine, amitryptiline, nortryptiline, levomepromezine and tioridazine) were observed. Comparisons of drug concentrations between those in human tissues and blowfly larvae showed different patterns of distribution that may be attributed to differences in physiology. Results confirm the reliability of entomological specimens for qualitative analyses, although quantitative extrapolations are unreliable. All xenobiotics detected were in higher concentrations in human tissues than in maggots. Concentrations in post-feeding maggots were significantly lower than for feeding maggots, suggesting that the feeding state of maggots may affect toxicological analyses as they metabolize and eliminate drugs during development.

  1. Mitochondrial Respiration Chain Enzymatic Activities in the Human Brain: Methodological Implications for Tissue Sampling and Storage.

    PubMed

    Ronsoni, Marcelo Fernando; Remor, Aline Pertile; Lopes, Mark William; Hohl, Alexandre; Troncoso, Iris H Z; Leal, Rodrigo Bainy; Boos, Gustavo Luchi; Kondageski, Charles; Nunes, Jean Costa; Linhares, Marcelo Neves; Lin, Kátia; Latini, Alexandra Susana; Walz, Roger

    2016-04-01

    Mitochondrial respiratory chain complexes enzymatic (MRCCE) activities were successfully evaluated in frozen brain samples. Epilepsy surgery offers an ethical opportunity to study human brain tissue surgically removed to treat drug resistant epilepsies. Epilepsy surgeries are done with hemodynamic and laboratory parameters to maintain physiology, but there are no studies analyzing the association among these parameters and MRCCE activities in the human brain tissue. We determined the intra-operative parameters independently associated with MRCCE activities in middle temporal neocortex (Cx), amygdala (AMY) and head of hippocampus (HIP) samples of patients (n = 23) who underwent temporal lobectomy using multiple linear regressions. MRCCE activities in Cx, AMY and HIP are differentially associated to trans-operative mean arterial blood pressure, O2 saturation, hemoglobin, and anesthesia duration to time of tissue sampling. The time-course between the last seizure occurrence and tissue sampling as well as the sample storage to biochemical assessments were also associated with enzyme activities. Linear regression models including these variables explain 13-17 % of MRCCE activities and show a moderate to strong effect (r = 0.37-0.82). Intraoperative hemodynamic and laboratory parameters as well as the time from last seizure to tissue sampling and storage time are associated with MRCCE activities in human samples from the Cx, AMYG and HIP. Careful control of these parameters is required to minimize confounding biases in studies using human brain samples collected from elective neurosurgery. PMID:26586405

  2. Optical detection of carotenoid antioxidants in human bone and surrounding tissue.

    PubMed

    Ermakov, Igor V; Ermakova, Maia R; Rosenberg, Thomas D; Gellermann, Werner

    2013-11-01

    Carotenoids are known to play an important role in health and disease state of living human tissue based on their antioxidant and optical filtering functions. In this study, we show that carotenoids exist in human bone and surrounding fatty tissue both in significant and individually variable concentrations. Measurements of biopsied tissue samples with molecule-specific Raman spectroscopy and high-performance liquid chromatography reveal that all carotenoids that are known to exist in human skin are also present in human bone. This includes all carotenes, lycopene, β-cryptoxanthin, lutein, and zeaxanthin. We propose quantitative reflection imaging as a noncontact optical method suitable for the measurement of composite carotenoid levels in bone and surrounding tissue exposed during open surgeries such as total knee arthroplasty, and as a proof of concept, demonstrate carotenoid measurements in biopsied bone samples. This will allow one to establish potential correlations between internal tissue carotenoid levels and levels in skin and to potentially use already existing optical skin carotenoid tests as surrogate marker for bone carotenoid status.

  3. Tissue-specific expression of human CD4 in transgenic mice.

    PubMed

    Gillespie, F P; Doros, L; Vitale, J; Blackwell, C; Gosselin, J; Snyder, B W; Wadsworth, S C

    1993-05-01

    The gene for the human CD4 glycoprotein, which serves as the receptor for human immunodeficiency virus type 1, along with approximately 23 kb of sequence upstream of the translational start site, was cloned. The ability of 5' flanking sequences to direct tissue-specific expression was tested in cell culture and in transgenic mice. A 5' flanking region of 6 kb was able to direct transcription of the CD4 gene in NIH 3T3 cells but did not result in detectable expression in the murine T-cell line EL4 or in four lines of transgenic mice. A larger 5' flanking region of approximately 23 kb directed high-level CD4 transcription in the murine T-cell line EL4 and in three independent lines of transgenic mice. Human CD4 expression in all tissues analyzed was tightly correlated with murine CD4 expression; the highest levels of human CD4 RNA expression were found in the thymus and spleen, with relatively low levels detected in other tissues. Expression of human CD4 protein in peripheral blood mononuclear cells was examined by flow cytometry in these transgenic animals and found to be restricted to the murine CD4+ subset of lymphocytes. Human CD4 protein, detected with an anti-human CD4 monoclonal antibody, was present on the surface of 45 to 50% of the peripheral blood mononuclear cells from all transgenic lines. PMID:8474453

  4. The release of spasmogenic substances from human chopped lung tissue and its inhibition

    PubMed Central

    Piper, Priscilla J.; Walker, Joyce L.

    1973-01-01

    1. Human lung tissue, passively sensitized with reaginic antibodies, released prostaglandins E1, E2 and F2α in addition to histamine and slow reacting substance (SRS-A), when exposed to the appropriate antigen. No rabbit aorta contracting substance (RCS) was detected. 2. Experiments with rats and guinea-pigs showed that the release of RCS is not confined to anaphylactic reactions mediated by non-reaginic antibodies but may be a feature of anaphylaxis in guinea-pigs alone. 3. Human lung tissue gently agitated with a blunt nylon rod liberated an E-type prostaglandin and RCS in addition to histamine and SRS-A. 4. Human isolated bronchial muscle was contracted by RCS. 5. Disodium cromoglycate antagonized the release of prostaglandins during anaphylaxis but not during agitation of human lung tissue, whereas indomethacin blocked the release of prostaglandins during agitation and anaphylaxis. 6. The release of an E-type prostaglandin during anaphylaxis in human lung tissue, which inhibits the further release of histamine could be another example of the regulatory role of prostaglandins in body functions. PMID:4352867

  5. Comparison of DNA adducts from exposure to complex mixtures in various human tissues and experimental systems

    PubMed Central

    Lewtas, Joellen; Mumford, Judy; Everson, Richard B.; Hulka, Barbara; Wilcosky, Tim; Kozumbo, Walter; Thompson, Claudia; George, Michael; Dobiáš, Lubomir; Šrám, Radim; Li, Xueming; Gallagher, Jane

    1993-01-01

    DNA adducts derived from complex mixtures of polycyclic aromatic compounds emitted from tobacco smoke are compared to industrial pollution sources (e.g., coke ovens and aluminum smelters), smoky coal burning, and urban air pollution. Exposures to coke oven emissions and smoky coal, both potent rodent skin tumor initiators and lung carcinogens in humans, result in high levels of DNA adducts compared to tobacco smoke in the in vitro calf thymus DNA model system, in cultured lymphocytes, and in the mouse skin assay. Using tobacco smoke as a model in human studies, we have compared relative DNA adduct levels detected in blood lymphocytes, placental tissue, bronchoalveolar lung lavage cells, sperm, and autopsy tissues of smokers and nonsmokers. Adduct levels in DNA isolated from smokers were highest in human heart and lung tissue with smaller but detectable differences in placental tissue and lung lavage cells. Comparison of the DNA adduct levels resulting from human exposure to different complex mixtures shows that emissions from coke ovens, aluminum smelters, and smoky coal result in higher DNA adduct levels than tobacco smoke exposure. These studies suggest that humans exposed to complex combustion mixtures will have higher DNA adduct levels in target cells (e.g., lung) as compared to nontarget cells (e.g., lymphocytes) and that the adduct levels will be dependent on the genotoxic and DNA adduct-forming potency of the mixture. ImagesFIGURE 1.FIGURE 1.FIGURE 2.FIGURE 3.FIGURE 3.FIGURE 3.FIGURE 3.FIGURE 3.FIGURE 3.FIGURE 4. PMID:8319665

  6. Effective atomic numbers for photon energy absorption and photon attenuation of tissues from human organs.

    PubMed

    Shivaramu

    2002-01-01

    Effective atomic numbers for photon energy- absorption (Z(PEA)eff) and photon interaction (ZPI(eff)) of human organs and tissues such as cortical bone, ovary, eye lens, testis, breast tissue, adipose tissue, lung tissue, soft tissue, soft tissue, (4-component), blood (whole), brain (grey/white matter), and skeletal muscle have been calculated by a direct method in the energy region of 1 keV to 20 MeV. The ZPEAeff and ZPIeff values steadily increase, up to 8-50 keV, and steadily decrease up to 1.25-2.0 MeV for all of the substances studied. From 2.0 MeV, the values rise with the increase in energy, up to 20 MeV. Significant differences exist between the ZPIeff and ZPEAeff in the energy region of 20-400 keV and 3-20 MeV for cortical bone; 15-150 keV for soft tissue, ovary, testis, blood, brain, lung, and skeletal muscle; 15-100 keV for breast tissue, eye lens, and soft tissue (4-component); and 10-100 keV for adipose tissue. A maximum difference of 28.37% is observed at 100 keV for cortical bone, and 30.43% at 40 keV for adipose tissue. For ovary, eye lens, testis, breast tissue, lung tissue, soft tissue, soft tissue (4-component), blood (whole), brain (grey/white matter), and skeletal muscle, a maximum difference of 31.74%, 29.60%, 31.87%, 30.61%, 31.47%, 31.52%, 29.95%, 31.63%, 32.36%, and 31.42%, respectively, is seen at 50 keV. The energy positions at which the maximum of ZPEAeff and ZPIeff occurs differ. The single effective atomic number directly obtained using the program XMuDat (Z(XMUDATTeff)) are found to be higher compared to those of ZPEAeff and ZPIeff values. The effect of absorption edge on effective atomic numbers, and its variation with photon energy and the possibility of defining 2 set values of effective atomic numbers below the absorption edges of elements present in the organs and tissues, are discussed.

  7. Critical insights into the beneficial and protective actions of the kallikrein-kinin system.

    PubMed

    Regoli, Domenico; Gobeil, Fernand

    2015-01-01

    Hypertension is characterized by an imbalance between the renin-angiotensin system (RAS) and the kallikrein-kinin system (KKS). Angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin II AT-1 receptor antagonists (also known as sartans or ARBs) are potent modulators of these systems and are highly effective as first-line treatments for hypertension, diabetic nephropathies, and diseases of the brain and coronary arteries. However, these agents are mechanistically distinct and should not be considered interchangeable. In this mini-review, we provide novel insights into the often neglected roles of the KKS in the beneficial, protective, and reparative actions of ACEIs. Indeed, ACEIs are the only antihypertensive drugs that properly reduce the imbalance between the RAS and the KKS, thereby restoring optimal cardiovascular homeostasis and significantly reducing morbidity and the risk of all-cause mortality among individuals affected by hypertension and other cardiovascular diseases. PMID:25579779

  8. Tissue Engineering for Human Urethral Reconstruction: Systematic Review of Recent Literature

    PubMed Central

    de Kemp, Vincent; de Graaf, Petra; Fledderus, Joost O.; Ruud Bosch, J. L. H.; de Kort, Laetitia M. O.

    2015-01-01

    Background Techniques to treat urethral stricture and hypospadias are restricted, as substitution of the unhealthy urethra with tissue from other origins (skin, bladder or buccal mucosa) has some limitations. Therefore, alternative sources of tissue for use in urethral reconstructions are considered, such as ex vivo engineered constructs. Purpose To review recent literature on tissue engineering for human urethral reconstruction. Methods A search was made in the PubMed and Embase databases restricted to the last 25 years and the English language. Results A total of 45 articles were selected describing the use of tissue engineering in urethral reconstruction. The results are discussed in four groups: autologous cell cultures, matrices/scaffolds, cell-seeded scaffolds, and clinical results of urethral reconstructions using these materials. Different progenitor cells were used, isolated from either urine or adipose tissue, but slightly better results were obtained with in vitro expansion of urothelial cells from bladder washings, tissue biopsies from the bladder (urothelium) or the oral cavity (buccal mucosa). Compared with a synthetic scaffold, a biological scaffold has the advantage of bioactive extracellular matrix proteins on its surface. When applied clinically, a non-seeded matrix only seems suited for use as an onlay graft. When a tubularized substitution is the aim, a cell-seeded construct seems more beneficial. Conclusions Considerable experience is available with tissue engineering of urethral tissue in vitro, produced with cells of different origin. Clinical and in vivo experiments show promising results. PMID:25689740

  9. Human and Mouse Brown Adipose Tissue Mitochondria Have Comparable UCP1 Function.

    PubMed

    Porter, Craig; Herndon, David N; Chondronikola, Maria; Chao, Tony; Annamalai, Palam; Bhattarai, Nisha; Saraf, Manish K; Capek, Karel D; Reidy, Paul T; Daquinag, Alexes C; Kolonin, Mikhail G; Rasmussen, Blake B; Borsheim, Elisabet; Toliver-Kinsky, Tracy; Sidossis, Labros S

    2016-08-01

    Brown adipose tissue (BAT) plays an important role in mammalian thermoregulation. The component of BAT mitochondria that permits this function is the inner membrane carrier protein uncoupling protein 1 (UCP1). To the best of our knowledge, no studies have directly quantified UCP1 function in human BAT. Further, whether human and rodent BAT have comparable thermogenic function remains unknown. We employed high-resolution respirometry to determine the respiratory capacity, coupling control, and, most importantly, UCP1 function of human supraclavicular BAT and rodent interscapular BAT. Human BAT was sensitive to the purine nucleotide GDP, providing the first direct evidence that human BAT mitochondria have thermogenically functional UCP1. Further, our data demonstrate that human and rodent BAT have similar UCP1 function per mitochondrion. These data indicate that human and rodent BAT are qualitatively similar in terms of UCP1 function. PMID:27508873

  10. Plasma Kallikrein-Kinin system mediates immune-mediated renal injury in trichloroethylene-sensitized mice.

    PubMed

    Wang, Hui; Zhang, Jia-Xiang; Ye, Liang-Ping; Li, Shu-Long; Wang, Feng; Zha, Wan-Sheng; Shen, Tong; Wu, Changhao; Zhu, Qi-Xing

    2016-07-01

    Trichloroethylene (TCE) is a major environmental pollutant. An immunological response is a newly-recognized mechanism for TCE-induced kidney damage. However, the role of the plasma kallikrein-kinin system (KKS) in immune-mediated kidney injury has never been examined. This study aimed to explore the role of the key components of the KKS, i.e. plasma kallikrein (PK), bradykinin (BK) and its receptors B1R and B2R, in TCE-induced kidney injury. A mouse model of skin sensitization was used to explore the mechanism of injury with or without a PK inhibitor PKSI. Kidney function was evaluated by measuring blood urea nitrogen (BUN) and creatinine (Cr) in conjunction with histopathologic characterization. Plasma BK was determined by ELISA; Renal C5b-9 membrane attack complex was evaluated by immunohistochemistry. Expression of BK and PK in the kidney was detected by immunofluorescence. mRNA and protein levels of B1R and B2R were assessed by real-time qPCR and Western blot. As expected, numerous inflammatory cell infiltration and tubular epithelial cell vacuolar degeneration were observed in TCE-sensitized mice. Moreover, serum BUN and Cr and plasma BK were increased. In addition, deposition of BK, PK and C5b-9 were observed and B1R and B2R mRNA and proteins levels were up-regulated. Pre-treatment with PKSI, a highly selective inhibitor of PK, alleviated TCE-induced renal damage. In addition, PKSI attenuated TCE-induced up-regulation of BK, PK and its receptors and C5b-9. These results provided the first evidence that activation of the KKS contributed to immune-mediated renal injury induced by TCE and also helped to identify the KKS as a potential therapeutic target for mitigating chemical sensitization-induced renal damage. PMID:27027470

  11. Plasma-kallikrein clearance by the liver of acetaminophen-intoxicated rats.

    PubMed

    de Toledo, C F; Borges, D R

    1993-01-01

    The liver synthesizes prokallikrein and is the main organ to clear the active enzyme (plasma-kallikrein) from circulation. This clearance, a receptor-mediated endocytosis, is calcium-independent and not affected by the blockade of Kupffer cells. The effects of endothelial cells blockade and of acetaminophen intoxication on the clearance of 10 nM rat plasma-kallikrein (RPK) by the isolated, exsanguinated and perfused rat liver are now reported. Endothelial cells blockade obtained by the addition of large excess (30 uM) of formaldehyde-treated serum albumin to the perfusion fluid does not affect the hepatic clearance of RPK (the half-lives of hepatic uptake were 15.5 +/- 1.0 min in the absence versus 16.5 +/- 1.4 min in the presence of the treated protein, p > 0.05). Some livers were perfused 24 hours after acetaminophen intoxication: 6.6 mmol/kg given i.p. after a 42-hour period of fast. Hepatocyte injury suggested by elevated aminotransferase activity (ALT 10 times control value, AST 30 times control value), acute phase inflammatory response (serum alpha 2-macroglobulin increase) and reduced synthetic function (serum albumin decrease), was confirmed histologically and only zone 3 hepatocytes were necrotic. A 66-hour period of fast does not affect by itself the hepatic clearance of RPK (16.9 +/- 1.3 min of half-life of hepatic uptake) when compared with the control group (15.5 +/- 1.0 min, p > 0.05). On the other hand the RPK clearance by the livers of rats previously intoxicated with acetaminophen was markedly deficient (the half-life of hepatic uptake was 39.2 +/- 3.2 min). These findings suggest that RPK is internalized by hepatocytes, preferentially by those of the perivenular zone of the hepatic acinus.

  12. Total mass attenuation coefficient evaluation of ten materials commonly used to simulate human tissue

    NASA Astrophysics Data System (ADS)

    Ferreira, C. C.; Ximenes, R. E.; Garcia, C. A. B.; Vieira, J. W.; Maia, A. F.

    2010-11-01

    To study the doses received by patient submitted to ionizing radiation, several materials are used to simulate the human tissue and organs. The total mass attenuation coefficient is a reasonable way for evaluating the usage in dosimetry of these materials. The total mass attenuation coefficient is determined by photon energy and constituent elements of the material. Currently, the human phantoms are composed by a unique material that presents characteristics similar to the mean proprieties of the different tissues within the region. Therefore, the phantoms are usually homogeneous and filled with a material similar to soft tissue. We studied ten materials used as soft tissue-simulating. These materials were named: bolus, nylon®, orange articulation wax, red articulation wax, PMMA, modelling clay, bee wax, paraffin 1, paraffin 2 and pitch. The objective of this study was to verify the best material to simulate the human cerebral tissue. We determined the elementary composition, mass density and, therefore, calculated the total mass attenuation coefficient of each material. The results were compared to the values established by the International Commission on Radiation Units and Measurements - ICRU, report n° 44, and by the International Commission on Radiation Protection - ICRP, report n° 89, to determine the best material for this energy interval. These results indicate that new head phantoms can be constructed with nylon®.

  13. Infrared Spectra of Human Breast Tumor Tissue and Experimental Animal Tumors

    NASA Astrophysics Data System (ADS)

    Tolstorozhev, G. B.; Belkov, M. V.; Skornyakov, I. V.; Pekhnyo, V. I.; Kozachkova, A. N.; Tsarik, H. V.; Kutsenko, I. P.; Sharykina, N. I.; Butra, V. A.

    2015-01-01

    We have used Fourier transform IR spectroscopy methods to conduct comparative studies of human breast tumors and sarcoma 180 tumor grafted into mice. The IR spectral parameters used to identify tumor tissue in mice with the sarcoma 180 strain proved to be identical to the parameters for human breast tissue in cancer. In the presence of a malignant tumor in humans, the most intense C=O vibrational bands in the protein molecules are observed in the interval 1710-1680 cm-1. For a benign tumor, in the IR spectra of breast tissue the intense bands are located in the interval 1670-1650 cm-1. We spectroscopically monitored the diagnosis and the chemotherapy process using the model of sarcoma 180 in mice. As the therapeutic drugs, we used synthesized coordination compounds based on palladium complexes with diphosphonic acid derivatives. We demonstrate the promising potential of palladium complexes with zoledronic acid as an effective cytostatic. In therapy using a palladium complex with zoledronic acid, the effect of tumor growth inhibition is accompanied by a change in its spectral characteristics. The parameters of the IR spectra for tumor tissue after treatment are close to those of the IR spectra for healthy tissue.

  14. Generating human intestinal tissue from pluripotent stem cells in vitro.

    PubMed

    McCracken, Kyle W; Howell, Jonathan C; Wells, James M; Spence, Jason R

    2011-12-01

    Here we describe a protocol for generating 3D human intestinal tissues (called organoids) in vitro from human pluripotent stem cells (hPSCs). To generate intestinal organoids, pluripotent stem cells are first differentiated into FOXA2(+)SOX17(+) endoderm by treating the cells with activin A for 3 d. After endoderm induction, the pluripotent stem cells are patterned into CDX2(+) mid- and hindgut tissue using FGF4 and WNT3a. During this patterning step, 3D mid- or hindgut spheroids bud from the monolayer epithelium attached to the tissue culture dish. The 3D spheroids are further cultured in Matrigel along with prointestinal growth factors, and they proliferate and expand over 1-3 months to give rise to intestinal tissue, complete with intestinal mesenchyme and epithelium comprising all of the major intestinal cell types. To date, this is the only method for efficiently directing the differentiation of hPSCs into 3D human intestinal tissue in vitro. PMID:22082986

  15. Effect of Human Ovarian Tissue Vitrification/Warming on the Expression of Genes Related to Folliculogenesis

    PubMed Central

    Shams Mofarahe, Zahra; Ghaffari Novin, Marefat; Jafarabadi, Mina; Salehnia, Mojdeh; Noroozian, Mohsen; Ghorbanmehr, Nassim

    2015-01-01

    Background: Ovarian tissue cryopreservation is an alternative strategy to preserve the fertility of women predicted to undergo premature ovarian failure. This study was designed to evaluate the expression of folliculogenesis-related genes, including factor in the germline alpha (FIGLA), growth differentiation factor-9 (GDF-9), follicle-stimulating hormone receptor (FSHR), and KIT LIGAND after vitrification/warming of human ovarian tissue. Methods: Human ovarian tissue samples were collected from five transsexual women. In the laboratory, the ovarian medullary part was removed by a surgical blade, and the cortical tissue was cut into small pieces. Some pieces were vitrified and warmed and the others were considered as non-vitrified group (control). Follicular normality was assessed with morphological observation by a light microscope, and the expression of FIGLA, KIT LIGAND, GDF-9,, and FSHR genes was examined using real-time RT-PCR in both the vitrified and non-vitrified groups. Results: Overall, 85% of the follicles preserved their normal morphologic feature after warming. The percentage of normal follicles and the expression of FIGLA, KIT LIGAND, GDF-9, and FSHR genes were similar in both vitrified and non-vitrified groups (P > 0.05). Conclusion: Vitrification/warming of human ovarian tissue had no remarkable effect on the expression of folliculogenesis-related genes. PMID:26175108

  16. Importance of good manufacturing practices in microbiological monitoring in processing human tissues for transplant.

    PubMed

    Pianigiani, Elisa; Ierardi, Francesca; Fimiani, Michele

    2013-12-01

    Skin allografts represent an important therapeutic resource in the treatment of severe skin loss. The risk associated with application of processed tissues in humans is very low, however, human material always carries the risk of disease transmission. To minimise the risk of contamination of grafts, processing is carried out in clean rooms where air quality is monitored. Procedures and quality control tests are performed to standardise the production process and to guarantee the final product for human use. Since we only validate and distribute aseptic tissues, we conducted a study to determine what type of quality controls for skin processing are the most suitable for detecting processing errors and intercurrent contamination, and for faithfully mapping the process without unduly increasing production costs. Two different methods for quality control were statistically compared using the Fisher exact test. On the basis of the current study we selected our quality control procedure based on pre- and post-processing tissue controls, operator and environmental controls. Evaluation of the predictability of our control methods showed that tissue control was the most reliable method of revealing microbial contamination of grafts. We obtained 100 % sensitivity by doubling tissue controls, while maintaining high specificity (77 %). PMID:23271587

  17. Tissue-engineered human bioartificial muscles expressing a foreign recombinant protein for gene therapy

    NASA Technical Reports Server (NTRS)

    Powell, C.; Shansky, J.; Del Tatto, M.; Forman, D. E.; Hennessey, J.; Sullivan, K.; Zielinski, B. A.; Vandenburgh, H. H.

    1999-01-01

    Murine skeletal muscle cells transduced with foreign genes and tissue engineered in vitro into bioartificial muscles (BAMs) are capable of long-term delivery of soluble growth factors when implanted into syngeneic mice (Vandenburgh et al., 1996b). With the goal of developing a therapeutic cell-based protein delivery system for humans, similar genetic tissue-engineering techniques were designed for human skeletal muscle stem cells. Stem cell myoblasts were isolated, cloned, and expanded in vitro from biopsied healthy adult (mean age, 42 +/- 2 years), and elderly congestive heart failure patient (mean age, 76 +/- 1 years) skeletal muscle. Total cell yield varied widely between biopsies (50 to 672 per 100 mg of tissue, N = 10), but was not significantly different between the two patient groups. Percent myoblasts per biopsy (73 +/- 6%), number of myoblast doublings prior to senescence in vitro (37 +/- 2), and myoblast doubling time (27 +/- 1 hr) were also not significantly different between the two patient groups. Fusion kinetics of the myoblasts were similar for the two groups after 20-22 doublings (74 +/- 2% myoblast fusion) when the biopsy samples had been expanded to 1 to 2 billion muscle cells, a number acceptable for human gene therapy use. The myoblasts from the two groups could be equally transduced ex vivo with replication-deficient retroviral expression vectors to secrete 0.5 to 2 microg of a foreign protein (recombinant human growth hormone, rhGH)/10(6) cells/day, and tissue engineered into human BAMs containing parallel arrays of differentiated, postmitotic myofibers. This work suggests that autologous human skeletal myoblasts from a potential patient population can be isolated, genetically modified to secrete foreign proteins, and tissue engineered into implantable living protein secretory devices for therapeutic use.

  18. HIV-1 cellular and tissue replication patterns in infected humanized mice.

    PubMed

    Araínga, Mariluz; Su, Hang; Poluektova, Larisa Y; Gorantla, Santhi; Gendelman, Howard E

    2016-01-01

    Humanized mice have emerged as a testing platform for HIV-1 pathobiology by reflecting natural human disease processes. Their use to study HIV-1 biology, virology, immunology, pathogenesis and therapeutic development has served as a robust alternative to more-well developed animal models for HIV/AIDS. A critical component in reflecting such human pathobiology rests in defining the tissue and cellular sites for HIV-1 infection. To this end, we examined the tissue sites for viral infection in bone marrow, blood, spleens, liver, gut, brain, kidney and lungs of human CD34+ hematopoietic stem cell engrafted virus-infected NOD.Cg-Prkdc(scid) Il2rg(tm1Wjl)/SzJ mice. Cells were analyzed by flow cytometry and sorted from species mixtures defined as CD34+ lineage negative progenitor cells, CD14+CD16+ monocyte-macrophages and central, stem cell and effector memory T cells. The cell distribution and viral life cycle were found dependent on the tissue compartment and time of infection. Cell subsets contained HIV-1 total and integrated DNA as well as multi-spliced and unspliced RNA in divergent proportions. The data support the idea that humanized mice can provide a means to examine the multifaceted sites of HIV-1 replication including, but not limited to progenitor cells and monocyte-macrophages previously possible only in macaques and human. PMID:26996968

  19. TOPICAL REVIEW: Human soft tissue analysis using x-ray or gamma-ray techniques

    NASA Astrophysics Data System (ADS)

    Theodorakou, C.; Farquharson, M. J.

    2008-06-01

    This topical review is intended to describe the x-ray techniques used for human soft tissue analysis. X-ray techniques have been applied to human soft tissue characterization and interesting results have been presented over the last few decades. The motivation behind such studies is to provide improved patient outcome by using the data obtained to better understand a disease process and improve diagnosis. An overview of theoretical background as well as a complete set of references is presented. For each study, a brief summary of the methodology and results is given. The x-ray techniques include x-ray diffraction, x-ray fluorescence, Compton scattering, Compton to coherent scattering ratio and attenuation measurements. The soft tissues that have been classified using x-rays or gamma rays include brain, breast, colon, fat, kidney, liver, lung, muscle, prostate, skin, thyroid and uterus.

  20. Amyloid structure exhibits polymorphism on multiple length scales in human brain tissue

    PubMed Central

    Liu, Jiliang; Costantino, Isabel; Venugopalan, Nagarajan; Fischetti, Robert F.; Hyman, Bradley T.; Frosch, Matthew P.; Gomez-Isla, Teresa; Makowski, Lee

    2016-01-01

    Aggregation of Aβ amyloid fibrils into plaques in the brain is a universal hallmark of Alzheimer’s Disease (AD), but whether plaques in different individuals are equivalent is unknown. One possibility is that amyloid fibrils exhibit different structures and different structures may contribute differentially to disease, either within an individual brain or between individuals. However, the occurrence and distribution of structural polymorphisms of amyloid in human brain is poorly documented. Here we use X-ray microdiffraction of histological sections of human tissue to map the abundance, orientation and structural heterogeneities of amyloid. Our observations indicate that (i) tissue derived from subjects with different clinical histories may contain different ensembles of fibrillar structures; (ii) plaques harboring distinct amyloid structures can coexist within a single tissue section and (iii) within individual plaques there is a gradient of fibrillar structure from core to margins. These observations have immediate implications for existing theories on the inception and progression of AD. PMID:27629394

  1. The post-analytical phase of histopathology practice: Storage, retention and use of human tissue specimens

    PubMed Central

    Kapila, Supriya Nikita; Boaz, Karen; Natarajan, Srikant

    2016-01-01

    There are several aspects to a histopathology practice besides the acquisition of biopsy specimens and histopathological diagnosis. Pathology Departments are home to an abundant source of knowledge in the form of stored specimens and slides. We attempt to highlight the importance of regulation of storage, retention, and appropriate use of human tissue material in research and ownership rights to the same. We also discuss requirement and waiver of informed consent for scientific work involving the use of such tissues, which in the absence of defined laws come under the purview of Institution Review Boards. Pathology Departments, under the binding of the parent institution, are conceded the responsibility of maintenance and retention of pathology specimens. This communication highlights some of the important aspects in human tissue material handling and research, underscoring the necessity for established regulations regarding the same. PMID:26958513

  2. Human adipose tissue-derived mesenchymal stem cells differentiate into insulin, somatostatin, and glucagon expressing cells

    SciTech Connect

    Timper, Katharina; Seboek, Dalma; Eberhardt, Michael; Linscheid, Philippe; Christ-Crain, Mirjam; Keller, Ulrich; Mueller, Beat; Zulewski, Henryk . E-mail: henryk.zulewski@unibas.ch

    2006-03-24

    Mesenchymal stem cells (MSC) from mouse bone marrow were shown to adopt a pancreatic endocrine phenotype in vitro and to reverse diabetes in an animal model. MSC from human bone marrow and adipose tissue represent very similar cell populations with comparable phenotypes. Adipose tissue is abundant and easily accessible and could thus also harbor cells with the potential to differentiate in insulin producing cells. We isolated human adipose tissue-derived MSC from four healthy donors. During the proliferation period, the cells expressed the stem cell markers nestin, ABCG2, SCF, Thy-1 as well as the pancreatic endocrine transcription factor Isl-1. The cells were induced to differentiate into a pancreatic endocrine phenotype by defined culture conditions within 3 days. Using quantitative PCR a down-regulation of ABCG2 and up-regulation of pancreatic developmental transcription factors Isl-1, Ipf-1, and Ngn3 were observed together with induction of the islet hormones insulin, glucagon, and somatostatin.

  3. Repair of Cartilage Defects in Arthritic Tissue with Differentiated Human Embryonic Stem Cells

    PubMed Central

    Olee, Tsaiwei; Grogan, Shawn P.; Lotz, Martin K.; Colwell, Clifford W.

    2014-01-01

    Chondrocytes have been generated in vitro from a range of progenitor cell types and by a number of strategies. However, achieving reconstitution of actual physiologically relevant, appropriately-laminated cartilage in situ that would be applicable to conditions, such as arthritis and cartilage degeneration remains elusive. This lack of success is multifactorial and includes limited cell source, decreased proliferation rate of mature chondrocytes, lack of maintenance of phenotype, reduced matrix synthesis, and poor integration with host tissue. We report an efficient approach for deriving mesenchymal chondroprogenitor cells from human embryonic stem cells. These cells generated tissue containing cartilage-specific matrix proteins that integrated in situ in a partial-thickness defect in ex vivo articular cartilage harvested from human arthritic joints. Given that stem cells provide a virtually inexhaustible supply of starting material and that our technique is easily scalable, cartilaginous tissue primed and grafted in this manner could be suitable for clinical translation. PMID:24028447

  4. The post-analytical phase of histopathology practice: Storage, retention and use of human tissue specimens.

    PubMed

    Kapila, Supriya Nikita; Boaz, Karen; Natarajan, Srikant

    2016-01-01

    There are several aspects to a histopathology practice besides the acquisition of biopsy specimens and histopathological diagnosis. Pathology Departments are home to an abundant source of knowledge in the form of stored specimens and slides. We attempt to highlight the importance of regulation of storage, retention, and appropriate use of human tissue material in research and ownership rights to the same. We also discuss requirement and waiver of informed consent for scientific work involving the use of such tissues, which in the absence of defined laws come under the purview of Institution Review Boards. Pathology Departments, under the binding of the parent institution, are conceded the responsibility of maintenance and retention of pathology specimens. This communication highlights some of the important aspects in human tissue material handling and research, underscoring the necessity for established regulations regarding the same.

  5. Amyloid structure exhibits polymorphism on multiple length scales in human brain tissue.

    PubMed

    Liu, Jiliang; Costantino, Isabel; Venugopalan, Nagarajan; Fischetti, Robert F; Hyman, Bradley T; Frosch, Matthew P; Gomez-Isla, Teresa; Makowski, Lee

    2016-01-01

    Aggregation of Aβ amyloid fibrils into plaques in the brain is a universal hallmark of Alzheimer's Disease (AD), but whether plaques in different individuals are equivalent is unknown. One possibility is that amyloid fibrils exhibit different structures and different structures may contribute differentially to disease, either within an individual brain or between individuals. However, the occurrence and distribution of structural polymorphisms of amyloid in human brain is poorly documented. Here we use X-ray microdiffraction of histological sections of human tissue to map the abundance, orientation and structural heterogeneities of amyloid. Our observations indicate that (i) tissue derived from subjects with different clinical histories may contain different ensembles of fibrillar structures; (ii) plaques harboring distinct amyloid structures can coexist within a single tissue section and (iii) within individual plaques there is a gradient of fibrillar structure from core to margins. These observations have immediate implications for existing theories on the inception and progression of AD. PMID:27629394

  6. 75 FR 9226 - Agency Information Collection Activities; Proposed Collection; Comment Request; Human Tissue...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-03-01

    ... immunodeficiency virus (HIV), hepatitis B, and hepatitis C through the use of human tissue for transplantation. The... (b) also requires recording and justification of any deviation from the written procedures. Section... excluded from the burden estimates as usual and customary business activities (5 CFR 1320.3(b)(2))....

  7. Regulation of LPS-induced tissue factor expression in human monocytic THP-1 cells by curcumin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Tissue factor (TF) is a transmembrane receptor, which initiates thrombotic episodes associated with various diseases. In addition to membrane-bound TF, we have discovered an alternatively spliced form of human TF mRNA. It was later confirmed that this form of TF mRNA expresses a soluble protein circ...

  8. On the wettability of soft tissues in the human oral cavity.

    PubMed

    van der Mei, Henny C; White, Don J; Busscher, Henk J

    2004-08-01

    In this study, the wettability of gingival surfaces in the human oral cavity was assessed by the measurement of intra-oral water contact angles. Intra-oral water contacts angles were measured in the morning prior to tooth brushing, immediately after tooth brushing and prior to and after lunch in order to reveal the influences of toothpaste and dietary components on the wettability of the gingiva. Within a group of 10 volunteers, gingival surfaces were hydrophobic, with water contact angles ranging from 72 to 79 degrees, which is high as compared with other soft tissues in the human body. Gingival contact angles were not affected by most commercial toothpastes involved in this study, but decreased slightly to 65 degrees after brushing with a hexametaphosphate containing toothpaste. During the day, however, the hydrophobicity readily recovered and after lunch contact angles on the gingival surfaces were higher than early in the morning, now ranging from 76 to 83 degrees. It is generally known that soft tissues in the human body involved in adsorptive and exchange functions and requiring lubrication are more hydrophilic than tissues with more protective functions. This study shows that gingival surfaces classify as the most hydrophobic soft tissue in the human body, attesting to their important protective role in the oral cavity.

  9. Tissue-specific transcriptome sequencing analysis expands the non-human primate reference transcriptome resource (NHPRTR)

    PubMed Central

    Peng, Xinxia; Thierry-Mieg, Jean; Thierry-Mieg, Danielle; Nishida, Andrew; Pipes, Lenore; Bozinoski, Marjan; Thomas, Matthew J.; Kelly, Sara; Weiss, Jeffrey M.; Raveendran, Muthuswamy; Muzny, Donna; Gibbs, Richard A.; Rogers, Jeffrey; Schroth, Gary P.; Katze, Michael G.; Mason, Christopher E.

    2015-01-01