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Sample records for fibrosis cells correlates

  1. Accumulation of Intrahepatic TNF-α-Producing NKp44+ NK Cells Correlates With Liver Fibrosis and Viral Load in Chronic HCV Infection.

    PubMed

    Nel, Isabelle; Lucar, Olivier; Petitdemange, Caroline; Béziat, Vivien; Lapalus, Martine; Bédossa, Pierre; Debré, Patrice; Asselah, Tarik; Marcellin, Patrick; Vieillard, Vincent

    2016-05-01

    In the setting of chronic hepatitis C virus (HCV) infection, changes in natural killer (NK) cells have been shown to reflect activation in response to virus stimulation. The contribution of individual natural cytotoxicity receptors to HCV infection remains to be clarified. NKp44 is the sole specific natural cytotoxicity receptor expressed only on activated NK cells.In this study, peripheral blood and liver NK-cell subsets were purified from 31 patients with chronic C hepatitis or nonalcoholic steatohepatitis, and then characterized by flow cytometry. Their polyfunctional activity was determined by expression of the CD107a degranulation marker, together with intracellular cytokine production.Unlike the patients with nonalcoholic steatohepatitis, patients with chronic HCV infection had a higher frequency of NKp44 NK cells in the liver than in their peripheral blood (P < 0.0001). Intrahepatic NKp44 NK cells from HCV individuals produced higher levels of tumor necrosis factor-α than did NKp44 NK cells (P = 0.0011). Importantly, the frequency of intrahepatic NKp44 NK cells was correlated with both HCV-RNA levels (P = 0.0234) and stage of fibrosis (P = 0.0003).Our findings suggest that the accumulation of intrahepatic tumor necrosis factor-α-producing NKp44 resident NK cells play a role in the liver damage associated with chronic HCV infection.

  2. Role of mast cells in fibrosis of classical Hodgkin lymphoma.

    PubMed

    Nakayama, Shoko; Yokote, Taiji; Hiraoka, Nobuya; Nishiwaki, Uta; Hanafusa, Toshiaki; Nishimura, Yasuichiro; Tsuji, Motomu

    2016-12-01

    The underlying mechanism of fibrosis in classical Hodgkin lymphoma (CHL) remains uncertain. This study aimed to investigate the association of fibrosis in the lymph nodes of patients with CHL through histological examination of the expression of cytokines associated with fibrosis and mast cell proliferation. Additionally, we sought to determine the degree of mast cell infiltration in a nodular sclerosis subtype of CHL (NSCHL) compared with that in non-NSCHL. We analyzed lymph nodes from 22 patients with CHL, of which eight were of the NSCHL and 14 of the non-NSCHL subtype, using immunohistochemical staining of forkhead box P3 (FOXP3), transforming growth factor (TGF)-β, interleukin (IL)-3, IL-13, and stem cell factor (SCF). Mast cells were positive for TGF-β and IL-13, and FOXP3-positive cells were negative for TGF-β. Only the expression of IL-13 in Hodgkin and Reed-Sternberg (HRS) cells was significantly more frequently observed in NSCHL than that in non-NSCHL (P = 0.0028) and was associated with a higher rate of fibrosis (P = 0.0097). The number of mast cells was significantly higher in NSCHL than that in non-NSCHL (P = 0.0001). A significantly positive correlation was observed between the rate of fibrosis and the number of mast cells (correlation coefficient, 0.8524; 95% CI, 0.6725-0.9372) (P <0.0001). The number of mast cells was significantly higher in the group with IL-13-positive HRS cells than that in the group with IL-13-negative HRS cells (P = 0.0157). Based on these findings, we hypothesize that IL-13 production by HRS cells may lead to fibrosis, and furthermore, promote mast cell proliferation and infiltration. This in turn might further produce the fibrotic cytokines IL-13 and TGF-β, resulting in fibrosis typical of NSCHL.

  3. Local Correlation Between Monte-Carlo Dose and Radiation-Induced Fibrosis in Lung Cancer Patients

    SciTech Connect

    Stroian, Gabriela; Martens, Chandra; Souhami, Luis; Collins, D. Louis; Seuntjens, Jan

    2008-03-01

    Purpose: To present a new method of evaluating the correlation between radiotherapy (RT)-induced fibrosis and the local dose delivered to non-small-cell lung cancer patients. Methods and Materials: Treatment plans were generated using the CadPlan treatment planning system (pencil beam, no heterogeneity corrections), and RT delivery was based on these plans. Retrospective Monte-Carlo dose calculations were performed, and the Monte-Carlo distributions of dose to real tissue were calculated using the planning computed tomography (CT) images and the number of monitor units actually delivered. After registration of the follow-up CT images with the planning CT images, different grades of radiologic fibrosis were automatically segmented on the follow-up CT images. Subsequently, patient-specific fibrosis probabilities were studied as a function of the local dose and a function of time after RT completion. Results: A strong patient-specific variation in the fibrosis volumes was found during the follow-up period. For both lungs, the threshold dose for which the probability of fibrosis became significant coincided with the threshold dose at which significant volumes of the lung were exposed. At later stages, only fibrosis localized in the high-dose regions persisted for both lungs. Overall, the Monte-Carlo dose distributions correlated much better with the probability of RT-induced fibrosis than did the CadPlan dose distributions. Conclusion: The presented method allows for an accurate, systematic, patient-specific and post-RT time-dependent numeric study of the relationship between RT-induced fibrosis and the local dose.

  4. Schistosome-induced cholangiocyte proliferation and osteopontin secretion correlate with fibrosis and portal hypertension in human and murine schistosomiasis mansoni.

    PubMed

    Pereira, Thiago A; Syn, Wing-Kin; Machado, Mariana V; Vidigal, Paula V; Resende, Vivian; Voieta, Izabela; Xie, Guanhua; Otoni, Alba; Souza, Márcia M; Santos, Elisângela T; Chan, Isaac S; Trindade, Guilherme V M; Choi, Steve S; Witek, Rafal P; Pereira, Fausto E; Secor, William E; Andrade, Zilton A; Lambertucci, José Roberto; Diehl, Anna Mae

    2015-11-01

    Schistosomiasis is a major cause of portal hypertension worldwide. It associates with portal fibrosis that develops during chronic infection. The mechanisms by which the pathogen evokes these host responses remain unclear. We evaluated the hypothesis that schistosome eggs release factors that directly stimulate liver cells to produce osteopontin (OPN), a pro-fibrogenic protein that stimulates hepatic stellate cells to become myofibroblasts. We also investigated the utility of OPN as a biomarker of fibrosis and/or severity of portal hypertension. Cultured cholangiocytes, Kupffer cells and hepatic stellate cells were treated with soluble egg antigen (SEA); OPN production was quantified by quantitative reverse transcriptase polymerase chain reaction (qRTPCR) and ELISA; cell proliferation was assessed by BrdU (5-bromo-2'-deoxyuridine). Mice were infected with Schistosoma mansoni for 6 or 16 weeks to cause early or advanced fibrosis. Liver OPN was evaluated by qRTPCR and immunohistochemistry (IHC) and correlated with liver fibrosis and serum OPN. Livers from patients with schistosomiasis mansoni (early fibrosis n=15; advanced fibrosis n=72) or healthy adults (n=22) were immunostained for OPN and fibrosis markers. Results were correlated with plasma OPN levels and splenic vein pressures. SEA-induced cholangiocyte proliferation and OPN secretion (P<0.001 compared with controls). Cholangiocytes were OPN (+) in Schistosoma-infected mice and humans. Liver and serum OPN levels correlated with fibrosis stage (mice: r=0.861; human r=0.672, P=0.0001) and myofibroblast accumulation (mice: r=0.800; human: r=0.761, P=0.0001). Numbers of OPN (+) bile ductules strongly correlated with splenic vein pressure (r=0.778; P=0.001). S. mansoni egg antigens stimulate cholangiocyte proliferation and OPN secretion. OPN levels in liver and blood correlate with fibrosis stage and portal hypertension severity.

  5. Congenital hepatic fibrosis, liver cell carcinoma and adult polycystic kidneys.

    PubMed

    Manes, J L; Kissane, J M; Valdes, A J

    1977-06-01

    In reviewing the literature, we found no liver cell carcinoma (LCC) or well-documented adult polycystic kidneys (APK) associated with congenital hepatic fibrosis (CHF). We report a 69-year-old man with CHF, LCC, APK, duplication cyst of distal portion of stomach, two calcified splenic artery aneurysms, myocardial fibrosis and muscular hypertrophy of esophagus. The LCC was grossly predunculated and microscopically showed prominent fibrosis and hyaline intracytoplasmic inclusions in the tumor cells.

  6. Genotype-phenotype correlation for pulmonary function in cystic fibrosis

    PubMed Central

    de Gracia, J; Mata, F; Alvarez, A; Casals, T; Gatner, S; Vendrell, M; de la Rosa, D; Guarner, L; Hermosilla, E

    2005-01-01

    Background: Since the CFTR gene was cloned, more than 1000 mutations have been identified. To date, a clear relationship has not been established between genotype and the progression of lung damage. A study was undertaken of the relationship between genotype, progression of lung disease, and survival in adult patients with cystic fibrosis (CF). Methods: A prospective cohort of adult patients with CF and two CFTR mutations followed up in an adult cystic fibrosis unit was analysed. Patients were classified according to functional effects of classes of CFTR mutations and were grouped based on the CFTR molecular position on the epithelial cell surface (I–II/I–II, I–II/III–V). Spirometric values, progression of lung disease, probability of survival, and clinical characteristics were analysed between groups. Results: Seventy four patients were included in the study. Patients with genotype I–II/I–II had significantly lower current spirometric values (p<0.001), greater loss of pulmonary function (p<0.04), a higher proportion of end-stage lung disease (p<0.001), a higher risk of suffering from moderate to severe lung disease (odds ratio 7.12 (95% CI 1.3 to 40.5)) and a lower probability of survival than patients with genotype I–II/III, I–II/IV and I–II/V (p<0.001). Conclusions: The presence of class I or II mutations on both chromosomes is associated with worse respiratory disease and a lower probability of survival. PMID:15994263

  7. Intrinsic pro-angiogenic status of cystic fibrosis airway epithelial cells

    SciTech Connect

    Verhaeghe, Catherine; Tabruyn, Sebastien P.; Oury, Cecile; Bours, Vincent . E-mail: vbours@ulg.ac.be; Griffioen, Arjan W.

    2007-05-11

    Cystic fibrosis is a common genetic disorder characterized by a severe lung inflammation and fibrosis leading to the patient's death. Enhanced angiogenesis in cystic fibrosis (CF) tissue has been suggested, probably caused by the process of inflammation, as similarly described in asthma and chronic bronchitis. The present study demonstrates an intrinsic pro-angiogenic status of cystic fibrosis airway epithelial cells. Microarray experiments showed that CF airway epithelial cells expressed several angiogenic factors such as VEGF-A, VEGF-C, bFGF, and PLGF at higher levels than control cells. These data were confirmed by real-time quantitative PCR and, at the protein level, by ELISA. Conditioned media of these cystic fibrosis cells were able to induce proliferation, migration and sprouting of cultured primary endothelial cells. This report describes for the first time that cystic fibrosis epithelial cells have an intrinsic angiogenic activity. Since excess of angiogenesis is correlated with more severe pulmonary disease, our results could lead to the development of new therapeutic applications.

  8. Mast Cells: A Pivotal Role in Pulmonary Fibrosis

    PubMed Central

    Veerappan, Arul; O'Connor, Nathan J.; Brazin, Jacqueline; Reid, Alicia C.; Jung, Albert; McGee, David; Summers, Barbara; Branch-Elliman, Dascher; Stiles, Brendon; Worgall, Stefan; Kaner, Robert J.

    2013-01-01

    Pulmonary fibrosis is characterized by an inflammatory response that includes macrophages, neutrophils, lymphocytes, and mast cells. The purpose of this study was to evaluate whether mast cells play a role in initiating pulmonary fibrosis. Pulmonary fibrosis was induced with bleomycin in mast-cell-deficient WBB6F1-W/Wv (MCD) mice and their congenic controls (WBB6F1-+/+). Mast cell deficiency protected against bleomycin-induced pulmonary fibrosis, but protection was reversed with the re-introduction of mast cells to the lungs of MCD mice. Two mast cell mediators were identified as fibrogenic: histamine and renin, via angiotensin (ANG II). Both human and rat lung fibroblasts express the histamine H1 and ANG II AT1 receptor subtypes and when activated, they promote proliferation, transforming growth factor β1 secretion, and collagen synthesis. Mast cells appear to be critical to pulmonary fibrosis. Therapeutic blockade of mast cell degranulation and/or histamine and ANG II receptors should attenuate pulmonary fibrosis. PMID:23570576

  9. Serum cell death biomarkers for prediction of liver fibrosis and poor prognosis in primary biliary cirrhosis.

    PubMed

    Sekiguchi, Tomohiro; Umemura, Takeji; Fujimori, Naoyuki; Shibata, Soichiro; Ichikawa, Yuki; Kimura, Takefumi; Joshita, Satoru; Komatsu, Michiharu; Matsumoto, Akihiro; Tanaka, Eiji; Ota, Masao

    2015-01-01

    The development of simple, noninvasive markers of liver fibrosis is urgently needed for primary biliary cirrhosis (PBC). This study examined the ability of several serum biomarkers of cell death to estimate fibrosis and prognosis in PBC. A cohort of 130 patients with biopsy-proven PBC and 90 healthy subjects were enrolled. We assessed the utility of the M30 ELISA, which detects caspase-cleaved cytokeratin-18 (CK-18) fragments and is representative of apoptotic cell death, as well as the M65 and newly developed M65 Epideath (M65ED) ELISAs, which detect total CK-18 as indicators of overall cell death, in predicting clinically relevant fibrosis stage. All 3 cell death biomarkers were significantly higher in patients with PBC than in healthy controls and were significantly correlated with fibrosis stage. The areas under the receiver operating characteristic curve for the M65 and M65ED assays for differentiation among significant fibrosis, severe fibrosis, and cirrhosis were 0.66 and 0.76, 0.66 and 0.73, and 0.74 and 0.82, respectively. In multivariate analysis, high M65ED (hazard ratio 6.13; 95% confidence interval 1.18-31.69; P = 0.031) and severe fibrosis (hazard ratio 7.45; 95% confidence interval 1.82-30.51; P = 0.005) were independently associated with liver-related death, transplantation, or decompensation. High serum M65ED was also significantly associated with poor outcome in PBC (log-rank test; P = 0.001). Noninvasive cell death biomarkers appear to be clinically useful in predicting fibrosis in PBC. Moreover, the M65ED assay may represent a new surrogate marker of adverse disease outcome.

  10. In hepatic fibrosis, liver sinusoidal endothelial cells acquire enhanced immunogenicity.

    PubMed

    Connolly, Michael K; Bedrosian, Andrea S; Malhotra, Ashim; Henning, Justin R; Ibrahim, Junaid; Vera, Valery; Cieza-Rubio, Napoleon E; Hassan, Burhan U; Pachter, H Leon; Cohen, Steven; Frey, Alan B; Miller, George

    2010-08-15

    The normal liver is characterized by immunologic tolerance. Primary mediators of hepatic immune tolerance are liver sinusoidal endothelial cells (LSECs). LSECs block adaptive immunogenic responses to Ag and induce the generation of T regulatory cells. Hepatic fibrosis is characterized by both intense intrahepatic inflammation and altered hepatic immunity. We postulated that, in liver fibrosis, a reversal of LSEC function from tolerogenic to proinflammatory and immunogenic may contribute to both the heightened inflammatory milieu and altered intrahepatic immunity. We found that, after fibrotic liver injury from hepatotoxins, LSECs become highly proinflammatory and secrete an array of cytokines and chemokines. In addition, LSECs gain enhanced capacity to capture Ag and induce T cell proliferation. Similarly, unlike LSECs in normal livers, in fibrosis, LSECs do not veto dendritic cell priming of T cells. Furthermore, whereas in normal livers, LSECs are active in the generation of T regulatory cells, in hepatic fibrosis LSECs induce an immunogenic T cell phenotype capable of enhancing endogenous CTLs and generating potent de novo CTL responses. Moreover, depletion of LSECs from fibrotic liver cultures mitigates the proinflammatory milieu characteristic of hepatic fibrosis. Our findings offer a critical understanding of the role of LSECs in modulating intrahepatic immunity and inflammation in fibro-inflammatory liver disease.

  11. SECs (Sinusoidal Endothelial Cells), Liver Microenvironment, and Fibrosis

    PubMed Central

    Natarajan, Vaishaali; Harris, Edward N.

    2017-01-01

    Liver fibrosis is a wound-healing response to chronic liver injury such as alcoholic/nonalcoholic fatty liver disease and viral hepatitis with no FDA-approved treatments. Liver fibrosis results in a continual accumulation of extracellular matrix (ECM) proteins and paves the way for replacement of parenchyma with nonfunctional scar tissue. The fibrotic condition results in drastic changes in the local mechanical, chemical, and biological microenvironment of the tissue. Liver parenchyma is supported by an efficient network of vasculature lined by liver sinusoidal endothelial cells (LSECs). These nonparenchymal cells are highly specialized resident endothelial cell type with characteristic morphological and functional features. Alterations in LSECs phenotype including lack of LSEC fenestration, capillarization, and formation of an organized basement membrane have been shown to precede fibrosis and promote hepatic stellate cell activation. Here, we review the interplay of LSECs with the dynamic changes in the fibrotic liver microenvironment such as matrix rigidity, altered ECM protein profile, and cell-cell interactions to provide insight into the pivotal changes in LSEC physiology and the extent to which it mediates the progression of liver fibrosis. Establishing the molecular aspects of LSECs in the light of fibrotic microenvironment is valuable towards development of novel therapeutic and diagnostic targets of liver fibrosis. PMID:28293634

  12. Retroperitoneal fibrosis in three siblings with the sickle cell trait

    PubMed Central

    Phills, James A.; Geggie, Peter; Hidvegi, Robert I.; Oliva, Luis A.

    1973-01-01

    Three West-Indian black siblings with the sickle cell trait developed retroperitoneal fibrosis, a previously unreported association. Other well known renal manifestations associated with the sickle cell trait were also present in some of these cases and included renal medullary necrosis and spontaneous hematuria. It is postulated that the sickling of the erythrocytes in the periureteral vessels resulted in thrombosis, ischemia, reactive scarring and progressive fibrosis indistinguishable from the known histological picture of retroperitoneal fibrosis. The finding of fibrin thrombi in the small veins of the fibrotic tissue of one of these patients would support this explanation. ImagesFIG. 1AFIG. 1BFIG. 2FIG. 3AFIG. 3BFIG. 4AFIG. 4BFIG. 4C PMID:4699274

  13. Synthesis of sulfated oligosaccharides by cystic fibrosis trachea epithelial cells.

    PubMed

    Mendicino, J; Sangadala, S

    1999-11-01

    The mucin glycoproteins in tracheal mucus of patients with cystic fibrosis is more highly sulfated than the corresponding secretions from healthy individuals [16]. In order to further characterize these differences in sulfation and possibly also glycosylation patterns, we compared the structures of sulfated mucin oligosaccharides synthesized by continuously cultured human tracheal cells transformed by simian virus 40. The synthesis of highly sulfated oligosaccharide chains in mucins secreted by normal human epithelial and submucosal cell lines were compared with mucins formed by cystic fibrosis tracheal epithelial and submucosal cell lines. The epithelial cell lines from cystic fibrosis trachea showed a higher rate of sulfate uptake and a significantly higher rate of synthesis and sulfation of high molecular weight chains. Mucins synthesized by each cell line in the presence of 35SO4 were isolated and oligosaccharide chains were released by beta-elimination and separated by ion exchange chromatography and gel filtration. The sulfated high molecular weight chains synthesized by the cystic fibrosis cell lines were characterized by methylation analysis and sequential glycosidase digestion before and after desulfation. Carbohydrate analysis yielded Fuc, Gal and GlcNAc in a ratio of 1:2:2.2 and only one galactosaminitol residue for about every 150-200 sugar residues present. The average molecular size of oligosaccharide chains in these fractions was between 30,000-40,000 daltons. These studies show that increased sulfation of oligosaccharides in mucins synthesized by cells from cystic fibrosis trachea is accompanied by a significant increase in the extension of a basic branched structure present in many of the lower molecular weight oligosaccharides.

  14. Correlation of fibrosis and transforming growth factor-beta type 2 levels in the eye.

    PubMed

    Connor, T B; Roberts, A B; Sporn, M B; Danielpour, D; Dart, L L; Michels, R G; de Bustros, S; Enger, C; Kato, H; Lansing, M

    1989-05-01

    Approximately 1 out of every 10 eyes undergoing surgery for retinal detachment develops excessive intraocular fibrosis that can lead to traction retinal detachment and ultimate blindness. This disease process has been termed proliferative vitreoretinopathy (PVR). The ability to monitor and grade this fibrotic response accurately within the eye as well as the ability to aspirate vitreous cavity fluid bathing the fibrotic tissue makes this an ideal setting in which to investigate the development of fibrosis. Although laboratory studies have recently shown that transforming growth factor-beta (TGF-beta) can enhance fibrosis, little clinical evidence is yet available correlating the level of this or other growth factors with the degree of fibrosis in a clinical setting. We have found that vitreous aspirates from eyes with intraocular fibrosis associated with PVR have more than three times the amount of TGF-beta (1,200 +/- 300 pM [SEM]) found in eyes with uncomplicated retinal detachments without intraocular fibrosis (360 +/- 91 pM [SEM]). Using an in vitro assay, 84-100% of the TGF-beta activity could be blocked with specific antibodies against TGF-beta 2, whereas only 10-21% could be blocked by specific antibodies against TGF-beta 1. TGF-beta 1 was used in an animal model of traction retinal detachment. Since beta 1 and beta 2 have essentially identical biologic effects and only human beta 1 was available in quantities required, beta 1 was chosen for these in vivo studies. The injection of TGF-beta1 plus fibronectin (FN) but not TGF-beta1 alone into the vitreous cavity of rabbits resulted in the increased formation of intraocular fibrosis and traction retinal detachments as compared to control eyes. In previous studies, intravitreal FN levels were also found to be elevated in eyes with intraocular fibrosis.

  15. Inflammatory Leukocyte Phenotypes Correlate with Disease Progression in Idiopathic Pulmonary Fibrosis

    PubMed Central

    Moore, Bethany B.; Fry, Chris; Zhou, Yueren; Murray, Susan; Han, MeiLan K.; Martinez, Fernando J.; Flaherty, Kevin R.

    2014-01-01

    Idiopathic pulmonary fibrosis (IPF) is characterized by progressive deposition of extracellular matrix, worsening dyspnea, and eventual mortality. Pathogenesis of IPF is poorly understood and the role inflammation and activated leukocytes play in the disease process is controversial. Previous studies demonstrated that activated leukocyte subsets characterize IPF patients. We sought to validate this observation in a well-defined cohort of 35 IPF patients and to correlate the observed leukocyte phenotypes with robust parameters of disease progression. We demonstrate that in univariate and multivariate analyses, increases in the CD14hi, CD16hi subset of monocytes measured at baseline correlated with disease progression, with a threshold value >0.5% of the total peripheral blood mononuclear cells being a significant predictor for worse outcome. In addition, several T cell subsets, including CD25 expressing CD4 cells, and CXCR3 expressing CD4 and CD8 subsets correlated with disease progression when found in increased percentages in the peripheral blood of IPF patients when sampled at baseline. Somewhat surprising in comparison to previous literature, the CD4 T cells did not appear to have lost expression of the co-stimulatory molecule, CD28, but the CD8 T cells did. Taken together, these results are consistent with the presence of an inflammatory process in IPF patients who eventually progress. However, when longitudinal measurements of these same markers were examined, there was significant heterogeneity of expression and these biomarkers did not necessarily remain elevated in IPF patients with progressive disease. We interpret this heterogeneity to suggest that IPF patients experience episodic inflammatory events that once triggered, may lead to disease progression. This longitudinal heterogeneity in biomarker analyses may explain why such markers are not consistently measured in all IPF cohorts. PMID:25580363

  16. Cystic fibrosis gene expression is not correlated with rectifying Cl sup minus channels

    SciTech Connect

    Ward, C.L.; Krouse, M.E.; Kopito, R.R.; Wine, J.J. ); Gruenert, D.C. )

    1991-06-15

    Cystic fibrosis (CF) involves a profound reduction of Cl{sup {minus}} permeability in several exocrine tissues. A distinctive, outwardly rectifying, depolarization-induced Cl{sup {minus}} channel (ORDIC channel) has been proposed to account for the Cl{sup {minus}} conductance that is defective in CF. The recently identified CF gene is predicted to code for a 1480-amino acid integral membrane protein termed the CF transmembrane conductance regulator (CFTR). The CFTR shares sequence similarity with a superfamily of ATP-binding membrane transport proteins such as P-glycoprotein and STE6, but it also has features consistent with an ion channel function. It has been proposed that the CFTR might be an ORDIC channel. To determine if CFTR and ORDIC channel expression are correlated, the authors surveyed various cell lines for natural variation in CFTR and ORDIC channel expression. In four human epithelial cell lines (T84, CaCo2, PANC-1, and 9HTEo-/S) that encompass the full observed range of CFTR mRNA levels and ORDIC channel density the authors found no correlation.

  17. Correlation between myocardial fibrosis and restrictive cardiac physiology in patients undergoing retransplantation.

    PubMed

    Kobashigawa, Jon A; Itagaki, Brandon K; Razi, Rabia R; Patel, Jignesh K; Chai, Wanxing; Kawano, Matthew A; Goldstein, Zachary; Kittleson, Michelle M; Fishbein, Michael C

    2013-01-01

    After cardiac transplant, there is often development of restrictive cardiac physiology. Little is known about the factors that contribute to this physiology and its correlation with pathology. Heart retransplantation provides a valuable opportunity to further understand this relationship. In this study, we investigated the correlation of myocardial fibrosis and restrictive physiology, and possible risk factors utilizing data from all retransplants at our center. A retrospective review of the 30 patients who underwent retransplantation at our institution between 1994 and 2004 was performed. Hemodynamic and imaging data were reviewed for the presence of restrictive physiology. Pathology reports were reviewed for the presence of myocardial fibrosis in the explanted hearts. The cohort with restrictive physiology preceding redo heart transplant had significantly more patients exhibiting myocardial fibrosis compared with the non-restrictive physiology group (94.1% vs. 15.4%, p < 0.001). We found no difference in the immunosuppressive regimen, history of rejection, and reason for transplant. In our study, we observed that myocardial fibrosis is an important contributor to the development of restrictive physiology. Further work needs to be done for risk stratification and the mechanism of fibrosis development.

  18. Bone marrow-derived progenitor cells in pulmonary fibrosis.

    PubMed

    Hashimoto, Naozumi; Jin, Hong; Liu, Tianju; Chensue, Stephen W; Phan, Sem H

    2004-01-01

    The origin of fibroblasts in pulmonary fibrosis is assumed to be intrapulmonary, but their extrapulmonary origin and especially derivation from bone marrow (BM) progenitor cells has not been ruled out. To examine this possibility directly, adult mice were durably engrafted with BM isolated from transgenic mice expressing enhanced GFP. Induction of pulmonary fibrosis in such chimera mice by endotracheal bleomycin (BLM) injection caused large numbers of GFP(+) cells to appear in active fibrotic lesions, while only a few GFP(+) cells could be identified in control lungs. Flow-cytometric analysis of lung cells confirmed the BLM-induced increase in GFP(+) cells in chimera mice and revealed a significant increase in GFP(+) cells that also express type I collagen. GFP(+) lung fibroblasts isolated from chimera mice expressed collagen and telomerase reverse transcriptase but not alpha-smooth muscle actin. Treatment of isolated GFP(+) fibroblasts with TGF-beta failed to induce myofibroblast differentiation. Cultured lung fibroblasts expressed the chemokine receptors CXCR4 and CCR7 and responded chemotactically to their cognate ligands, stromal cell-derived factor-1 alpha and secondary lymphoid chemokine, respectively. Thus the collagen-producing lung fibroblasts in pulmonary fibrosis can also be derived from BM progenitor cells.

  19. Correlation analysis between four serum biomarkers of liver fibrosis and liver function in infants with cholestasis

    PubMed Central

    TANG, NING; ZHANG, YAPING; LIU, ZEYU; FU, TAO; LIANG, QINGHONG; AI, XUEMEI

    2016-01-01

    The aim of the present study was to investigate the correlation between four serum biomarkers of liver fibrosis and liver function in infants with cholestasis. A total of 30 infants with cholestasis and 20 healthy infants were included in the study. Biochemical assays based on the initial rate method and colorimetric assays were conducted to determine the levels of liver function markers in the serum [such as alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), direct bilirubin (DBIL), indirect bilirubin (IBIL), γ-glutamyl transferase (γ-GT), cholinesterase (CHE) and total bile acids (TBA)] and four serum biomarkers of liver fibrosis were measured using radioimmunoassays [hyaluronic acid (HA), procollagen type III (PCIII), laminin (LN) and collagen type IV (cIV)]. The serum levels of ALT, AST, TBIL, DBIL, IBIL, γ-GT and TBA in the infants with cholestasis were significantly higher compared to the healthy infants (P<0.01); the serum levels of CHE in the infants with cholestasis were significantly lower compared to the healthy infants (P<0.01). The serum levels of HA, PCIII, and cIV in the infants with cholestasis were significantly higher compared to the healthy infants (P<0.01). Correlation analyses between liver function and the four biomarkers of liver fibrosis showed that HA was positively correlated with AST and γ-GT (P<0.05) and negatively correlated with ALT, CHE and TBA (P<0.05). cIV was positively correlated with γ-GT (P<0.05) and negatively correlated with CHE (P<0.05). In conclusion, statistically significant differences were identified for the liver function markers (ALT, AST, TBIL, DBIL, IBIL, γ-GT and TBA) and the biomarkers HA, PCIII and cIV of liver fibrosis between infants with cholestasis and healthy infants. Thus, the serum levels of HA, cIV, γ-GT and CHE are sensitive markers for cholestatic liver fibrosis in infants. PMID:27347413

  20. Bone marrow–derived progenitor cells in pulmonary fibrosis

    PubMed Central

    Hashimoto, Naozumi; Jin, Hong; Liu, Tianju; Chensue, Stephen W.; Phan, Sem H.

    2004-01-01

    The origin of fibroblasts in pulmonary fibrosis is assumed to be intrapulmonary, but their extrapulmonary origin and especially derivation from bone marrow (BM) progenitor cells has not been ruled out. To examine this possibility directly, adult mice were durably engrafted with BM isolated from transgenic mice expressing enhanced GFP. Induction of pulmonary fibrosis in such chimera mice by endotracheal bleomycin (BLM) injection caused large numbers of GFP+ cells to appear in active fibrotic lesions, while only a few GFP+ cells could be identified in control lungs. Flow-cytometric analysis of lung cells confirmed the BLM-induced increase in GFP+ cells in chimera mice and revealed a significant increase in GFP+ cells that also express type I collagen. GFP+ lung fibroblasts isolated from chimera mice expressed collagen and telomerase reverse transcriptase but not α-smooth muscle actin. Treatment of isolated GFP+ fibroblasts with TGF-β failed to induce myofibroblast differentiation. Cultured lung fibroblasts expressed the chemokine receptors CXCR4 and CCR7 and responded chemotactically to their cognate ligands, stromal cell–derived factor-1α and secondary lymphoid chemokine, respectively. Thus the collagen-producing lung fibroblasts in pulmonary fibrosis can also be derived from BM progenitor cells. PMID:14722616

  1. Expression of cystic fibrosis transmembrane conductance regulator corrects defective chloride channel regulation in cystic fibrosis airway epithelial cells

    NASA Astrophysics Data System (ADS)

    Rich, Devra P.; Anderson, Matthew P.; Gregory, Richard J.; Cheng, Seng H.; Paul, Sucharita; Jefferson, Douglas M.; McCann, John D.; Klinger, Katherine W.; Smith, Alan E.; Welsh, Michael J.

    1990-09-01

    The cystic fibrosis transmembrane conductance regulator (CFTR) was expressed in cultured cystic fibrosis airway epithelial cells and Cl- channel activation assessed in single cells using a fluorescence microscopic assay and the patch-clamp technique. Expression of CFTR, but not of a mutant form of CFTR (ΔF508), corrected the Cl- channel defect. Correction of the phenotypic defect demonstrates a causal relationship between mutations in the CFTR gene and defective Cl- transport which is the hallmark of the disease.

  2. Enhancement of hidden structures of early skin fibrosis using polarization degree patterns and Pearson correlation analysis.

    PubMed

    Sviridov, Alexander P; Chernomordik, Victor; Hassan, Moinuddin; Boccara, Albert C; Russo, Angelo; Smith, Paul; Gandjbakhche, Amir

    2005-01-01

    The skin of athymic nude mice is irradiated with a single dose of x-ray irradiation that initiated fibrosis. Digital photographs of the irradiated mice are taken by illuminating the mouse skin with linearly polarized probe light of 650 nm. The specific pattern of the surface distribution of the degree of polarization enables the detection of initial skin fibrosis structures that were not visually apparent. Data processing of the raw spatial distributions of the degree of polarization based on Fourier filtering of the high-frequency noise improves subjective perception of the revealed structure in the images. In addition, Pearson correlation analysis provides information about skin structural size and directionality.

  3. Baseline ultrasound and clinical correlates in children with cystic fibrosis

    PubMed Central

    Leung, Daniel H.; Ye, Wen; Molleston, Jean P.; Weymann, Alexander; Ling, Simon; Paranjape, Shruti M.; Romero, Rene; Schwarzenberg, Sara Jane; Palermo, Joseph; Alonso, Estella M.; Murray, Karen F.; Marshall, Bruce C.; Sherker, Averell H.; Siegel, Marilyn J.; Krishnamurthy, Rajesh; Harned, Roger; Karmazyn, Boaz; Magee, John C.; Narkewicz, Michael R

    2015-01-01

    Objective To investigate the relationship between abdominal ultrasound (US) findings and demographic, historical and clinical features in children with CF. Study design Children age 3-12 years with CF without known cirrhosis, were enrolled in a prospective, multi-center study of US to predict hepatic fibrosis. Consensus US patterns were assigned by 3 radiologists as normal, heterogeneous, homogeneous, or cirrhosis. Data were derived from direct collection and U.S. or Toronto CF registries. Chi-square or ANOVA were used to compare variables among US groups and between normal and abnormal. Logistic regression was used to study risk factors for having abnormal US. Results Findings in 719 subjects were normal (n=590, 82.1%), heterogeneous (64, 8.9%), homogeneous (41, 5.7%), and cirrhosis (24, 3.3%). Cirrhosis (p=0.0004), homogeneous (p<0.0001) and heterogeneous (p=0.03) were older than normal. More males were heterogeneous (p=0.001). More heterogeneous (15.0%, p=0.009) and cirrhosis (25.0%, p=0.005) had CF-related diabetes or impaired glucose tolerance versus normal (5.4%). Early infection with Pseudomonas aeruginosa (<2 years old) was associated with a lower risk (OR 0.42, p=0.0007) of abnormal. Ursodeoxycholic acid use (OR 3.69, p <0.0001) and CF-related diabetes (OR 2.21, p=0.019) were associated with increased risk of abnormal. Conclusions Unsuspected cirrhosis is seen in 3.3% of young patients with CF, heterogeneous in 8.9%. abnormal US is associated with CF-related diabetes, and early P aeruginosa is associated with normal US. Prospective assessment of these risk factors may identify potential interventional targets. PMID:26254836

  4. Cystic fibrosis mice carrying the missense mutation G551D replicate human genotype-phenotype correlations.

    PubMed Central

    Delaney, S J; Alton, E W; Smith, S N; Lunn, D P; Farley, R; Lovelock, P K; Thomson, S A; Hume, D A; Lamb, D; Porteous, D J; Dorin, J R; Wainwright, B J

    1996-01-01

    We have generated a mouse carrying the human G551D mutation in the cystic fibrosis transmembrane conductance regulator gene (CFTR) by a one-step gene targeting procedure. These mutant mice show cystic fibrosis pathology but have a reduced risk of fatal intestinal blockage compared with 'null' mutants, in keeping with the reduced incidence of meconium ileus in G551D patients. The G551D mutant mice show greatly reduced CFTR-related chloride transport, displaying activity intermediate between that of cftr(mlUNC) replacement ('null') and cftr(mlHGU) insertional (residual activity) mutants and equivalent to approximately 4% of wild-type CFTR activity. The long-term survival of these animals should provide an excellent model with which to study cystic fibrosis, and they illustrate the value of mouse models carrying relevant mutations for examining genotype-phenotype correlations. Images PMID:8605891

  5. Shear wave elastography results correlate with liver fibrosis histology and liver function reserve

    PubMed Central

    Feng, Yan-Hong; Hu, Xiang-Dong; Zhai, Lin; Liu, Ji-Bin; Qiu, Lan-Yan; Zu, Yuan; Liang, Si; Gui, Yu; Qian, Lin-Xue

    2016-01-01

    AIM: To evaluate the correlation of shear wave elastography (SWE) results with liver fibrosis histology and quantitative function reserve. METHODS: Weekly subcutaneous injection of 60% carbon tetrachloride (1.5 mL/kg) was given to 12 canines for 24 wk to induce experimental liver fibrosis, with olive oil given to 2 control canines. At 24 wk, liver condition was evaluated using clinical biochemistry assays, SWE imaging, lidocaine metabolite monoethylglycine-xylidide (MEGX) test, and histologic fibrosis grading. Clinical biochemistry assays were performed at the institutional central laboratory for routine liver function evaluation. Liver stiffness was measured in triplicate from three different intercostal spaces and expressed as mean liver stiffness modulus (LSM). Plasma concentrations of lidocaine and its metabolite MEGX were determined using high-performance liquid chromatography repeated in duplicate. Liver biopsy samples were fixed in 10% formaldehyde, and liver fibrosis was graded using the modified histological activity index Knodell score (F0-F4). Correlations among histologic grading, LSM, and MEGX measures were analyzed with the Pearson linear correlation coefficient. RESULTS: At 24 wk liver fibrosis histologic grading was as follows: F0, n = 2 (control); F1, n = 0; F2, n = 3; F3, n = 7; and F4, n = 2. SWE LSM was positively correlated with histologic grading (r = 0.835, P < 0.001). Specifically, the F4 group had a significantly higher elastic modulus than the F3, F2, and F0 groups (P = 0.002, P = 0.003, and P = 0.006, respectively), and the F3 group also had a significantly higher modulus than the control F0 group (P = 0.039). LSM was negatively associated with plasma MEGX concentrations at 30 min (r = -0.642; P = 0.013) and 60 min (r = -0.651; P = 0.012), time to ½ of the maximum concentration (r = -0.538; P = 0.047), and the area under the curve (r = -0.636; P = 0.014). Multiple comparisons showed identical differences in these three measures

  6. Paricalcitol Reduces Peritoneal Fibrosis in Mice through the Activation of Regulatory T Cells and Reduction in IL-17 Production

    PubMed Central

    González-Mateo, Guadalupe T.; Fernández-Míllara, Vanessa; Bellón, Teresa; Liappas, Georgios; Ruiz-Ortega, Marta; López-Cabrera, Manuel; Selgas, Rafael; Aroeira, Luiz S.

    2014-01-01

    Fibrosis is a significant health problem associated with a chronic inflammatory reaction. The precise mechanisms involved in the fibrotic process are still poorly understood. However, given that inflammation is a major causative factor, immunomodulation is a possible therapeutic approach to reduce fibrosis. The vitamin D receptor (VDR) that is present in all hematopoietic cells has been associated with immunomodulation. We investigated whether the intraperitoneal administration of paricalcitol, a specific activator of the VDR, modulates peritoneal dialysis fluid (PDF)-induced peritoneal fibrosis. We characterized the inflammatory process in the peritoneal cavity of mice treated or not treated with paricalcitol and analyzed the ensuing fibrosis. The treatment reduced peritoneal IL-17 levels, which strongly correlated with a significantly lower peritoneal fibrotic response. In vitro studies demonstrate that both CD4+ and CD8+ regulatory T cells appear to impact the regulation of IL-17. Paricalcitol treatment resulted in a significantly increased frequency of CD8+ T cells showing a regulatory phenotype. The frequency of CD4+ Tregs tends to be increased, but it did not achieve statistical significance. However, paricalcitol treatment increased the number of CD4+ and CD8+ Treg cells in vivo. In conclusion, the activation of immunological regulatory mechanisms by VDR signaling could prevent or reduce fibrosis, as shown in peritoneal fibrosis induced by PDF exposure in mice. PMID:25279459

  7. Renal Medullary and Cortical Correlates in Fibrosis, Epithelial Mass, Microvascularity, and Microanatomy Using Whole Slide Image Analysis Morphometry

    PubMed Central

    Farris, Alton B.; Ellis, Carla L.; Rogers, Thomas E.; Lawson, Diane; Cohen, Cynthia; Rosen, Seymour

    2016-01-01

    Renal tubulointerstitial injury often leads to interstitial fibrosis and tubular atrophy (IF/TA). IF/TA is typically assessed in the renal cortex and can be objectively quantitated with computerized image analysis (IA). However, the human medulla accounts for a substantial proportion of the nephron; therefore, medullary scarring will have important cortical consequences and may parallel overall chronic renal injury. Trichrome, periodic acid–Schiff (PAS), and collagen III immunohistochemistry (IHC) were visually examined and quantitated on scanned whole slide images (WSIs) (N = 67 cases). When tuned to measure fibrosis, IA of trichrome and Trichrome-PAS (T-P) WSIs correlated for all anatomic compartments (among cortex, medulla, and entire tissue, r = 0.84 to 0.89, P all <0.0001); and collagen III deposition correlated between compartments (r = 0.69 to 0.89, P <0.0001 to 0.0002); however, trichrome and T-P measures did not correlate with collagen deposition, suggesting heterogeneous contributions to extracellular matrix deposition. Epithelial cell mass (EPCM) correlated between cortex and medulla when measured with cytokeratin IHC and with the trichrome red portion (r = 0.85 and 0.66, respectively, all P < 0.0001). Visual assessment also correlated between compartments for fibrosis and EPCM. Correlations were found between increasing medullary inner stripe (IS) width and fibrosis in all of the tissue and the medulla by trichrome morphometry (r = 0.56, P < 0.0001, and r = 0.48, P = 0.00008, respectively). Weak correlations were found between increasing IS width and decreasing visual assessment of all tissue EPCM. Microvessel density (MVD) and microvessel area (MVA) measured using a MVD algorithm applied to CD34 IHC correlated significantly between all compartments (r = 0.76 to 0.87 for MVD and 0.71 to 0.87 for MVA, P all < 0.0001). Overall, these findings demonstrate the interrelatedness of the cortex and medulla and the importance of considering the renal

  8. Genetics and epithelial cell dysfunction in cystic fibrosis

    SciTech Connect

    Riordan, J.R.; Buchwald, M.

    1987-01-01

    This book examines the advances being made in the study of the physiology, cell biology, and molecular genetics of cystic fibrosis. Emphasis is placed on various areas of research that involve epithelial cells (e.g., the CF-specific phenotypes exhibited by epithelial cells, abnormalities in epithelium ion transport, chloride channel regulation in CF epithelial.) Coverage is presented on the current status of CF, including data on the incidence of the disease, its mode of inheritance, chromosomal localization, genetic heterogeneity, and screening and management.

  9. Management of Fibrosis: The Mesenchymal Stromal Cells Breakthrough

    PubMed Central

    Usunier, Benoît; Benderitter, Marc; Tamarat, Radia; Chapel, Alain

    2014-01-01

    Fibrosis is the endpoint of many chronic inflammatory diseases and is defined by an abnormal accumulation of extracellular matrix components. Despite its slow progression, it leads to organ malfunction. Fibrosis can affect almost any tissue. Due to its high frequency, in particular in the heart, lungs, liver, and kidneys, many studies have been conducted to find satisfactory treatments. Despite these efforts, current fibrosis management therapies either are insufficiently effective or induce severe adverse effects. In the light of these facts, innovative experimental therapies are being investigated. Among these, cell therapy is regarded as one of the best candidates. In particular, mesenchymal stromal cells (MSCs) have great potential in the treatment of inflammatory diseases. The value of their immunomodulatory effects and their ability to act on profibrotic factors such as oxidative stress, hypoxia, and the transforming growth factor-β1 pathway has already been highlighted in preclinical and clinical studies. Furthermore, their propensity to act depending on the microenvironment surrounding them enhances their curative properties. In this paper, we review a large range of studies addressing the use of MSCs in the treatment of fibrotic diseases. The results reported here suggest that MSCs have antifibrotic potential for several organs. PMID:25132856

  10. Neutrophilic Bronchial Inflammation Correlates with Clinical and Functional Findings in Patients with Noncystic Fibrosis Bronchiectasis

    PubMed Central

    Dente, Federico L.; Bilotta, Marta; Bartoli, Maria Laura; Bacci, Elena; Cianchetti, Silvana; Latorre, Manuela; Malagrinò, Laura; Nieri, Dario; Roggi, Maria Adelaide; Vagaggini, Barbara; Paggiaro, Pierluigi

    2015-01-01

    Background. Neutrophilic bronchial inflammation is a main feature of bronchiectasis, but not much is known about its relationship with other disease features. Aim. To compare airway inflammatory markers with clinical and functional findings in subjects with stable noncystic fibrosis bronchiectasis (NCFB). Methods. 152 NFCB patients (62.6 years; females: 57.2%) underwent clinical and functional cross-sectional evaluation, including microbiologic and inflammatory cell profile in sputum, and exhaled breath condensate malondialdehyde (EBC-MDA). NFCB severity was assessed using BSI and FACED criteria. Results. Sputum neutrophil percentages inversely correlated with FEV1 (P < 0.0001; rho = −0.428), weakly with Leicester Cough Questionnaire score (P = 0.068; rho = −0.58), and directly with duration of the disease (P = 0.004; rho = 0.3) and BSI severity score (P = 0.005; rho = 0.37), but not with FACED. Sputum neutrophilia was higher in colonized subjects, P. aeruginosa colonized subjects showing greater sputum neutrophilia and lower FEV1. Patients with ≥3 exacerbations in the last year showed a significantly greater EBC-MDA than the remaining patients. Conclusions. Sputum neutrophilic inflammation and biomarkers of oxidative stress in EBC can be considered good biomarkers of disease severity in NCFB patients, as confirmed by pulmonary function, disease duration, bacterial colonization, BSI score, and exacerbation rate. PMID:26819500

  11. Gadolinium contrast agent-induced CD163+ ferroportin+ osteogenic cells in nephrogenic systemic fibrosis.

    PubMed

    Swaminathan, Sundararaman; Bose, Chhanda; Shah, Sudhir V; Hall, Kimberly A; Hiatt, Kim M

    2013-09-01

    Gadolinium-based contrast agents are linked to nephrogenic systemic fibrosis in patients with renal insufficiency. The pathology of nephrogenic systemic fibrosis is characterized by abnormal tissue repair: fibrosis and ectopic ossification. The mechanisms by which gadolinium could induce fibrosis and ossification are not known. We examined in vitro the effect of a gadolinium-based contrast agent on human peripheral blood mononuclear cells for phenotype and function relevant to the pathology of nephrogenic systemic fibrosis using immunofluorescence, flow cytometry, real-time PCR, and osteogenic assays. We also examined tissues from patients with nephrogenic systemic fibrosis, using IHC to identify the presence of cells with phenotype induced by gadolinium. Gadolinium contrast induced differentiation of human peripheral blood mononuclear cells into a unique cellular phenotype--CD163(+) cells expressing proteins involved in fibrosis and bone formation. These cells express fibroblast growth factor (FGF)23, osteoblast transcription factors Runt-related transcription factor 2, and osterix, and show an osteogenic phenotype in in vitro assays. We show in vivo the presence of CD163(+)/procollagen-1(+)/osteocalcin(+) cells in the fibrotic and calcified tissues of nephrogenic systemic fibrosis patients. Gadolinium contrast-induced CD163(+)/ferroportin(+)/FGF23(+) cells with osteogenic potential may play a role in systemic fibrosis and ectopic ossification in nephrogenic systemic fibrosis.

  12. Fibrosis in Chronic Hepatitis C: Correlation between Immunohistochemically-Assessed Virus Load with Steatosis and Cellular Iron Content

    PubMed Central

    Akl, Maha; Hindawi, Ali EL; Mosaad, Maha; Montasser, Ahmed; Ray, Ahmed El; Khalil, Heba; Anas, Amgad; Atta, Raffat; Paradis, Valerie; Hadi, Ahmed Abdel; Hammam, Olfat

    2016-01-01

    AIM: We aimed study impact of hepatocytic viral load, steatosis, and iron load on fibrosis in chronic hepatitis C and role of VEGF and VEGFR overexpression in cirrhotic cases in evolving HCC. MATERIAL AND METHODS: Total of 120 cases were included from TBRI and Beaujon Hospital as chronic hepatitis C (CHC), post-hepatitis C cirrhosis, and HCC. Cases of CHC were stained for Sirius red, Prussian blue and immunohistochemically (IHC) for HCV-NS3/NS4. HCC were stained IHC for VEGF and by FISH. RESULTS: Stage of fibrosis was significantly correlated with inflammation in CHC (P < 0.01). Noticed iron load did not correlate with fibrosis. Steatosis was associated with higher inflammation and fibrosis. The cellular viral load did not correlate with inflammation, steatosis or fibrosis. VEGF by IHC was significantly higher in cases of HCC when compared to cirrhotic group (P < 0.001). Amplification of VEGFR2 was confirmed in 40% of cases of HCC. Scoring of VEGF by IHC was the good indicator of VEGFR2 amplification by FISH (P < 0.005). CONCLUSION: Grade of inflammation is the factor affecting fibrosis in CHC. The degree of liver damage is not related to cellular viral load or iron load. Steatosis is associated with higher inflammation and fibrosis. VEGF by IHC is correlated with overexpression of VEGFR2 by FISH. PMID:28028394

  13. Atrophy, fibrosis, and increased PAX7-positive cells in pharyngeal muscles of oculopharyngeal muscular dystrophy patients.

    PubMed

    Gidaro, Teresa; Negroni, Elisa; Perié, Sophie; Mirabella, Massimiliano; Lainé, Jeanne; Lacau St Guily, Jean; Butler-Browne, Gillian; Mouly, Vincent; Trollet, Capucine

    2013-03-01

    Oculopharyngeal muscular dystrophy (OPMD) is a late-onset autosomal dominant inherited dystrophy caused by an abnormal trinucleotide repeat expansion in the poly(A)-binding-protein-nuclear 1 (PABPN1) gene. Primary muscular targets of OPMD are the eyelid elevator and pharyngeal muscles, including the cricopharyngeal muscle (CPM), the progressive involution of which leads to ptosis and dysphagia, respectively. To understand the consequences of PABPN1 polyalanine expansion in OPMD, we studied muscle biopsies from 14 OPMD patients, 3 inclusion body myositis patients, and 9 healthy controls. In OPMD patient CPM (n = 6), there were typical dystrophic features with extensive endomysial fibrosis and marked atrophy of myosin heavy-chain IIa fibers. There were more PAX7-positive cells in all CPM versus other muscles (n = 5, control; n = 3, inclusion body myositis), and they were more numerous in OPMD CPM versus control normal CPM without any sign of muscle regeneration. Intranuclear inclusions were present in all OPMD muscles but unaffected OPMD patient muscles (i.e. sternocleidomastoid, quadriceps, or deltoid; n = 14) did not show evidence of fibrosis, atrophy, or increased PAX7-positive cell numbers. These results suggest that the specific involvement of CPM in OPMD might be caused by failure of the regenerative response with dysfunction of PAX7-positive cells and exacerbated fibrosis that does not correlate with the presence of PABPN1 inclusions.

  14. Pharmacological Intervention in Hepatic Stellate Cell Activation and Hepatic Fibrosis

    PubMed Central

    Schon, Hans-Theo; Bartneck, Matthias; Borkham-Kamphorst, Erawan; Nattermann, Jacob; Lammers, Twan; Tacke, Frank; Weiskirchen, Ralf

    2016-01-01

    The activation and transdifferentiation of hepatic stellate cells (HSCs) into contractile, matrix-producing myofibroblasts (MFBs) are central events in hepatic fibrogenesis. These processes are driven by autocrine- and paracrine-acting soluble factors (i.e., cytokines and chemokines). Proof-of-concept studies of the last decades have shown that both the deactivation and removal of hepatic MFBs as well as antagonizing profibrogenic factors are in principle suitable to attenuate ongoing hepatic fibrosis. Although several drugs show potent antifibrotic activities in experimental models of hepatic fibrosis, there is presently no effective pharmaceutical intervention specifically approved for the treatment of liver fibrosis. Pharmaceutical interventions are generally hampered by insufficient supply of drugs to the diseased liver tissue and/or by adverse effects as a result of affecting non-target cells. Therefore, targeted delivery systems that bind specifically to receptors solely expressed on activated HSCs or transdifferentiated MFBs and delivery systems that can improve drug distribution to the liver in general are urgently needed. In this review, we summarize current strategies for targeted delivery of drugs to the liver and in particular to pro-fibrogenic liver cells. The applicability and efficacy of sequestering molecules, selective protein carriers, lipid-based drug vehicles, viral vectors, transcriptional targeting approaches, therapeutic liver- and HSC-specific nanoparticles, and miRNA-based strategies are discussed. Some of these delivery systems that had already been successfully tested in experimental animal models of ongoing hepatic fibrogenesis are expected to translate into clinically useful therapeutics specifically targeting HSCs. PMID:26941644

  15. Pseudomonas aeruginosa quorum sensing molecules correlate with clinical status in cystic fibrosis.

    PubMed

    Barr, Helen L; Halliday, Nigel; Cámara, Miguel; Barrett, David A; Williams, Paul; Forrester, Douglas L; Simms, Rebecca; Smyth, Alan R; Honeybourne, David; Whitehouse, Joanna L; Nash, Edward F; Dewar, Jane; Clayton, Andrew; Knox, Alan J; Fogarty, Andrew W

    2015-10-01

    Pseudomonas aeruginosa produces quorum sensing signal molecules that are potential biomarkers for infection.A prospective study of 60 cystic fibrosis patients with chronic P. aeruginosa, who required intravenous antibiotics for pulmonary exacerbations, was undertaken. Clinical measurements and biological samples were obtained at the start and end of the treatment period. Additional data were available for 29 of these patients when they were clinically stable.Cross-sectionally, quorum sensing signal molecules were detectable in the sputum, plasma and urine of 86%, 75% and 83% patients, respectively. They were positively correlated between the three biofluids. Positive correlations were observed for most quorum sensing signal molecules in sputum, plasma and urine, with quantitative measures of pulmonary P. aeruginosa load at the start of a pulmonary exacerbation. Plasma concentrations of 2-nonyl-4-hydroxy-quinoline (NHQ) were significantly higher at the start of a pulmonary exacerbation compared to clinical stability (p<0.01). Following the administration of systemic antibiotics, plasma 2-heptyl-4-hydroxyquinoline (p=0.02) and NHQ concentrations (p<0.01) decreased significantly.In conclusion, quorum sensing signal molecules are detectable in cystic fibrosis patients with pulmonary P. aeruginosa infection and are positively correlated with quantitative measures of P. aeruginosa. NHQ correlates with clinical status and has potential as a novel biomarker for P. aeruginosa infection.

  16. Pseudomonas aeruginosa quorum sensing molecules correlate with clinical status in cystic fibrosis

    PubMed Central

    Halliday, Nigel; Cámara, Miguel; Barrett, David A.; Williams, Paul; Forrester, Douglas L.; Simms, Rebecca; Smyth, Alan R.; Honeybourne, David; Whitehouse, Joanna L.; Nash, Edward F.; Dewar, Jane; Clayton, Andrew; Knox, Alan J.; Fogarty, Andrew W.

    2015-01-01

    Pseudomonas aeruginosa produces quorum sensing signal molecules that are potential biomarkers for infection. A prospective study of 60 cystic fibrosis patients with chronic P. aeruginosa, who required intravenous antibiotics for pulmonary exacerbations, was undertaken. Clinical measurements and biological samples were obtained at the start and end of the treatment period. Additional data were available for 29 of these patients when they were clinically stable. Cross-sectionally, quorum sensing signal molecules were detectable in the sputum, plasma and urine of 86%, 75% and 83% patients, respectively. They were positively correlated between the three biofluids. Positive correlations were observed for most quorum sensing signal molecules in sputum, plasma and urine, with quantitative measures of pulmonary P. aeruginosa load at the start of a pulmonary exacerbation. Plasma concentrations of 2-nonyl-4-hydroxy-quinoline (NHQ) were significantly higher at the start of a pulmonary exacerbation compared to clinical stability (p<0.01). Following the administration of systemic antibiotics, plasma 2-heptyl-4-hydroxyquinoline (p=0.02) and NHQ concentrations (p<0.01) decreased significantly. In conclusion, quorum sensing signal molecules are detectable in cystic fibrosis patients with pulmonary P. aeruginosa infection and are positively correlated with quantitative measures of P. aeruginosa. NHQ correlates with clinical status and has potential as a novel biomarker for P. aeruginosa infection. PMID:26022946

  17. The role of circulating mesenchymal progenitor cells (fibrocytes) in the pathogenesis of pulmonary fibrosis.

    PubMed

    Strieter, Robert M; Keeley, Ellen C; Hughes, Molly A; Burdick, Marie D; Mehrad, Borna

    2009-11-01

    Pulmonary fibrosis is associated with a number of disorders that affect the lung. Although there are several cellular types that are involved in the pathogenesis pulmonary fibrosis, the resident lung fibroblast has been viewed traditionally as the primary cell involved in promoting the deposition of ECM that culminates in pulmonary fibrosis. However, recent findings demonstrate that a circulating cell (i.e., the fibrocyte) can contribute to the evolution of pulmonary fibrosis. Fibrocytes are bone marrow-derived mesenchymal progenitor cells that express a variety of cell-surface markers related to leukocytes, hematopoietic progenitor cells, and fibroblasts. Fibrocytes are unique in that they are capable of differentiating into fibroblasts and myofibroblasts, as well as adipocytes. In this review, we present data supporting the critical role these cells play in the pathogenesis of pulmonary fibrosis.

  18. The role of circulating mesenchymal progenitor cells, fibrocytes, in promoting pulmonary fibrosis.

    PubMed

    Strieter, Robert M; Keeley, Ellen C; Burdick, Marie D; Mehrad, Borna

    2009-01-01

    The resident fibroblast has been traditionally viewed as the primary cell involved in promoting pulmonary fibrosis. However, contemporary findings now support the concept of a circulating cell (fibrocyte) that also contributes to pulmonary fibrosis. Fibrocytes are bone marrow-derived mesenchymal progenitor cells that express a variety of cell surface markers related to leukocytes, hematopoietic progenitor cells and fibroblasts. Fibrocytes are unique in that they are capable of differentiating into fibroblasts and myofibroblasts, as well as adipocytes. In this review, we present data supporting the critical role these cells play in the pathogenesis of pulmonary fibrosis.

  19. CD8+ T Cell Response to Gammaherpesvirus Infection Mediates Inflammation and Fibrosis in Interferon Gamma Receptor-Deficient Mice

    PubMed Central

    O’Flaherty, Brigid M.; Matar, Caline G.; Wakeman, Brian S.; Garcia, AnaPatricia; Wilke, Carol A.; Courtney, Cynthia L.; Moore, Bethany B.; Speck, Samuel H.

    2015-01-01

    Idiopathic pulmonary fibrosis (IPF), one of the most severe interstitial lung diseases, is a progressive fibrotic disorder of unknown etiology. However, there is growing appreciation for the role of viral infection in disease induction and/or progression. A small animal model of multi-organ fibrosis, which involves murine gammaherpesvirus (MHV68) infection of interferon gamma receptor deficient (IFNγR-/-) mice, has been utilized to model the association of gammaherpesvirus infections and lung fibrosis. Notably, several MHV68 mutants which fail to induce fibrosis have been identified. Our current study aimed to better define the role of the unique MHV68 gene, M1, in development of pulmonary fibrosis. We have previously shown that the M1 gene encodes a secreted protein which possesses superantigen-like function to drive the expansion and activation of Vβ4+ CD8+ T cells. Here we show that M1-dependent fibrosis is correlated with heightened levels of inflammation in the lung. We observe an M1-dependent cellular infiltrate of innate immune cells with most striking differences at 28 days-post infection. Furthermore, in the absence of M1 protein expression we observed reduced CD8+ T cells and MHV68 epitope specific CD8+ T cells to the lungs—despite equivalent levels of viral replication between M1 null and wild type MHV68. Notably, backcrossing the IFNγR-/- onto the Balb/c background, which has previously been shown to exhibit weak MHV68-driven Vβ4+ CD8+ T cell expansion, eliminated MHV68-induced fibrosis—further implicating the activated Vβ4+ CD8+ T cell population in the induction of fibrosis. We further addressed the role that CD8+ T cells play in the induction of fibrosis by depleting CD8+ T cells, which protected the mice from fibrotic disease. Taken together these findings are consistent with the hypothesized role of Vβ4+ CD8+ T cells as mediators of fibrotic disease in IFNγR-/- mice. PMID:26317335

  20. A network model of correlated growth of tissue stiffening in pulmonary fibrosis

    NASA Astrophysics Data System (ADS)

    Oliveira, Cláudio L. N.; Bates, Jason H. T.; Suki, Béla

    2014-06-01

    During the progression of pulmonary fibrosis, initially isolated regions of high stiffness form and grow in the lung tissue due to collagen deposition by fibroblast cells. We have previously shown that ongoing collagen deposition may not lead to significant increases in the bulk modulus of the lung until these local remodeled regions have become sufficiently numerous and extensive to percolate in a continuous path across the entire tissue (Bates et al 2007 Am. J. Respir. Crit. Care Med. 176 617). This model, however, did not include the possibility of spatially correlated deposition of collagen. In the present study, we investigate whether spatial correlations influence the bulk modulus in a two-dimensional elastic network model of lung tissue. Random collagen deposition at a single site is modeled by increasing the elastic constant of the spring at that site by a factor of 100. By contrast, correlated collagen deposition is represented by stiffening the springs encountered along a random walk starting from some initial spring, the rationale being that excess collagen deposition is more likely in the vicinity of an already stiff region. A combination of random and correlated deposition is modeled by performing random walks of length N from randomly selected initial sites, the balance between the two processes being determined by N. We found that the dependence of bulk modulus, B(N,c), on both N and the fraction of stiff springs, c, can be described by a strikingly simple set of empirical equations. For c<0.3, B(N,c) exhibits exponential growth from its initial value according to B(N,c)\\approx {{B}_{0}}exp (2c)\\left[ 1+{{c}^{\\beta }}ln \\left( {{N}^{{{a}_{I}}}} \\right) \\right], where \\beta =0.994+/- 0.024 and {{a}_{I}}=0.54+/- 0.026. For intermediate concentrations of stiffening, 0.3\\leqslant c\\leqslant 0.8, another exponential rule describes the bulk modulus as B(N,c)=4{{B}_{0}}exp \\left[ {{a}_{II}}\\left( c-{{c}_{c}} \\right) \\right], where {{a

  1. A network model of correlated growth of tissue stiffening in pulmonary fibrosis.

    PubMed

    Oliveira, Cláudio L N; Bates, Jason H T; Suki, Béla

    2014-06-26

    During the progression of pulmonary fibrosis, initially isolated regions of high stiffness form and grow in the lung tissue due to collagen deposition by fibroblast cells. We have previously shown that ongoing collagen deposition may not lead to significant increases in the bulk modulus of the lung until these local remodeled regions have become sufficiently numerous and extensive to percolate in a continuous path across the entire tissue [Bates et al. 2007 Am. J. Respir. Crit. Care Med. 176 617]. This model, however, did not include the possibility of spatially correlated deposition of collagen. In the present study, we investigate whether spatial correlations influence the bulk modulus in a two-dimensional elastic network model of lung tissue. Random collagen deposition at a single site is modeled by increasing the elastic constant of the spring at that site by a factor of 100. By contrast, correlated collagen deposition is represented by stiffening the springs encountered along a random walk starting from some initial spring, the rationale being that excess collagen deposition is more likely in the vicinity of an already stiff region. A combination of random and correlated deposition is modeled by performing random walks of length N from randomly selected initial sites, the balance between the two processes being determined by N. We found that the dependence of bulk modulus, B(N, c), on both N and the fraction of stiff springs, c, can be described by a strikingly simple set of empirical equations. For c < 0.3, B(N, c) exhibits exponential growth from its initial value according to B(N, c) ≈ B0exp(2c)[1 + c(β) ln(N(a)(I))], where β = 0.994 ± 0.024 and aI = 0.54 ± 0.026. For intermediate concentrations of stiffening, 0.3 ≤ c ≤ 0.8, another exponential rule describes the bulk modulus as B(N, c) = 4B0exp[aII (c - cc )], where aII and cc are parameters that depend on N. For c > 0.8, B(N, c) is linear in c and independent of N, such that B(N, c) = 100B0

  2. A network model of correlated growth of tissue stiffening in pulmonary fibrosis

    PubMed Central

    Oliveira, Cláudio L N; Bates, Jason H T; Suki, Béla

    2014-01-01

    During the progression of pulmonary fibrosis, initially isolated regions of high stiffness form and grow in the lung tissue due to collagen deposition by fibroblast cells. We have previously shown that ongoing collagen deposition may not lead to significant increases in the bulk modulus of the lung until these local remodeled regions have become sufficiently numerous and extensive to percolate in a continuous path across the entire tissue [Bates et al. 2007 Am. J. Respir. Crit. Care Med. 176 617]. This model, however, did not include the possibility of spatially correlated deposition of collagen. In the present study, we investigate whether spatial correlations influence the bulk modulus in a two-dimensional elastic network model of lung tissue. Random collagen deposition at a single site is modeled by increasing the elastic constant of the spring at that site by a factor of 100. By contrast, correlated collagen deposition is represented by stiffening the springs encountered along a random walk starting from some initial spring, the rationale being that excess collagen deposition is more likely in the vicinity of an already stiff region. A combination of random and correlated deposition is modeled by performing random walks of length N from randomly selected initial sites, the balance between the two processes being determined by N. We found that the dependence of bulk modulus, B(N, c), on both N and the fraction of stiff springs, c, can be described by a strikingly simple set of empirical equations. For c < 0.3, B(N, c) exhibits exponential growth from its initial value according to B(N, c) ≈ B0exp(2c)[1 + cβ ln(NaI)], where β = 0.994 ± 0.024 and aI = 0.54 ± 0.026. For intermediate concentrations of stiffening, 0.3 ≤ c ≤ 0.8, another exponential rule describes the bulk modulus as B(N, c) = 4B0exp[aII (c − cc)], where aII and cc are parameters that depend on N. For c > 0.8, B(N, c) is linear in c and independent of N, such that B(N, c) = 100B0 − 100a

  3. Dendritic cells maintain dermal adipose–derived stromal cells in skin fibrosis

    PubMed Central

    Chia, Jennifer J.; Zhu, Tong; Chyou, Susan; Dasoveanu, Dragos C.; Carballo, Camila; Tian, Sha; Magro, Cynthia M.; Rodeo, Scott; Spiera, Robert F.; Ruddle, Nancy H.; McGraw, Timothy E.; Browning, Jeffrey L.; Lafyatis, Robert; Gordon, Jessica K.; Lu, Theresa T.

    2016-01-01

    Scleroderma is a group of skin-fibrosing diseases for which there are no effective treatments. A feature of the skin fibrosis typical of scleroderma is atrophy of the dermal white adipose tissue (DWAT). Adipose tissue contains adipose-derived mesenchymal stromal cells (ADSCs) that have regenerative and reparative functions; however, whether DWAT atrophy in fibrosis is accompanied by ADSC loss is poorly understood, as are the mechanisms that might maintain ADSC survival in fibrotic skin. Here, we have shown that DWAT ADSC numbers were reduced, likely because of cell death, in 2 murine models of scleroderma skin fibrosis. The remaining ADSCs showed a partial dependence on dendritic cells (DCs) for survival. Lymphotoxin β (LTβ) expression in DCs maintained ADSC survival in fibrotic skin by activating an LTβ receptor/β1 integrin (LTβR/β1 integrin) pathway on ADSCs. Stimulation of LTβR augmented the engraftment of therapeutically injected ADSCs, which was associated with reductions in skin fibrosis and improved skin function. These findings provide insight into the effects of skin fibrosis on DWAT ADSCs, identify a DC-ADSC survival axis in fibrotic skin, and suggest an approach for improving mesenchymal stromal cell therapy in scleroderma and other diseases. PMID:27721238

  4. Chemokine Receptor Ccr6 Deficiency Alters Hepatic Inflammatory Cell Recruitment and Promotes Liver Inflammation and Fibrosis

    PubMed Central

    Blaya, Delia; Morales-Ibanez, Oriol; Coll, Mar; Millán, Cristina; Altamirano, José; Arroyo, Vicente; Caballería, Joan; Bataller, Ramón; Ginès, Pere; Sancho-Bru, Pau

    2015-01-01

    Chronic liver diseases are characterized by a sustained inflammatory response in which chemokines and chemokine-receptors orchestrate inflammatory cell recruitment. In this study we investigated the role of the chemokine receptor CCR6 in acute and chronic liver injury. In the absence of liver injury Ccr6-/- mice presented a higher number of hepatic macrophages and increased expression of pro-inflammatory cytokines and M1 markers Tnf-α, Il6 and Mcp1. Inflammation and cell recruitment were increased after carbon tetrachloride-induced acute liver injury in Ccr6-/- mice. Moreover, chronic liver injury by carbon tetrachloride in Ccr6-/- mice was associated with enhanced inflammation and fibrosis, altered macrophage recruitment, enhanced CD4+ cells and a reduction in Th17 (CD4+IL17+) and mature dendritic (MHCII+CD11c+) cells recruitment. Clodronate depletion of macrophages in Ccr6-/- mice resulted in a reduction of hepatic pro-inflammatory and pro-fibrogenic markers in the absence and after liver injury. Finally, increased CCR6 hepatic expression in patients with alcoholic hepatitis was found to correlate with liver expression of CCL20 and severity of liver disease. In conclusion, CCR6 deficiency affects hepatic inflammatory cell recruitment resulting in the promotion of hepatic inflammation and fibrosis. PMID:26691857

  5. Chemokine Receptor Ccr6 Deficiency Alters Hepatic Inflammatory Cell Recruitment and Promotes Liver Inflammation and Fibrosis.

    PubMed

    Affò, Silvia; Rodrigo-Torres, Daniel; Blaya, Delia; Morales-Ibanez, Oriol; Coll, Mar; Millán, Cristina; Altamirano, José; Arroyo, Vicente; Caballería, Joan; Bataller, Ramón; Ginès, Pere; Sancho-Bru, Pau

    2015-01-01

    Chronic liver diseases are characterized by a sustained inflammatory response in which chemokines and chemokine-receptors orchestrate inflammatory cell recruitment. In this study we investigated the role of the chemokine receptor CCR6 in acute and chronic liver injury. In the absence of liver injury Ccr6-/- mice presented a higher number of hepatic macrophages and increased expression of pro-inflammatory cytokines and M1 markers Tnf-α, Il6 and Mcp1. Inflammation and cell recruitment were increased after carbon tetrachloride-induced acute liver injury in Ccr6-/- mice. Moreover, chronic liver injury by carbon tetrachloride in Ccr6-/- mice was associated with enhanced inflammation and fibrosis, altered macrophage recruitment, enhanced CD4+ cells and a reduction in Th17 (CD4+IL17+) and mature dendritic (MHCII+CD11c+) cells recruitment. Clodronate depletion of macrophages in Ccr6-/- mice resulted in a reduction of hepatic pro-inflammatory and pro-fibrogenic markers in the absence and after liver injury. Finally, increased CCR6 hepatic expression in patients with alcoholic hepatitis was found to correlate with liver expression of CCL20 and severity of liver disease. In conclusion, CCR6 deficiency affects hepatic inflammatory cell recruitment resulting in the promotion of hepatic inflammation and fibrosis.

  6. Huangqi decoction inhibits apoptosis and fibrosis, but promotes Kupffer cell activation in dimethylnitrosamine-induced rat liver fibrosis

    PubMed Central

    2012-01-01

    Background Previously, Huangqi decoction (HQD) has been found to have a potential therapeutic effect on DMN-induced liver cirrhosis. Here, the mechanisms of HQD action against liver fibrosis were investigated in relation to hepatocyte apoptosis and hepatic inflammation regulation. Methods Liver fibrosis was induced by DMN administration for 2 or 4 weeks. Hepatocyte apoptosis and of Kupffer cells (KC) and hepatic stellate cells (HSC) interaction were investigated using confocal microscopy. The principle cytokines, fibrogenic proteins and apoptotic factors were investigated using western blot analysis. Results Compared with the DMN-water group, HQD showed decreased hepatocyte apoptosis and reduced expression of apoptotic effectors, cleaved-caspase-3, and fibrotic factors, such as smooth muscle α-actin (α-SMA), transforming growth factor beta-1 (TGF-β1). However, the KC marker CD68 increased significantly in DMN-HQD liver. Confocal microscopy demonstrated widespread adhesion of KCs to HSCs in DMN-water and DMN-HQD rats liver. Conclusions HQD exhibited positive protective effects against liver fibrosis; its mechanism of action was associated with protection from hepatocyte apoptosis and the promotion of CD68 expression in the devolopment of liver fibrosis to cirrhosis development. PMID:22531084

  7. Mesothelial cell autoantibodies upregulate transcription factors associated with fibrosis.

    PubMed

    Gilmer, John; Harding, Tanner; Woods, Linda; Black, Brad; Flores, Raja; Pfau, Jean

    2017-01-01

    Amphibole asbestos exposure is associated with the production of mesothelial cell autoantibodies (MCAA). These MCAA have been linked with pleural fibrotic disease in the asbestos exposed community of Libby, Montana, and induce collagen deposition by cultured mesothelial cells. However, the exact intracellular mechanism by which these autoantibodies cause an increase in collagen deposition remains unknown. This study sought to gain insight into the transcription factors involved in the collagen production after human mesothelial cells are exposed to MCAA. In this study, transcription factor activation profiles were generated from human mesothelial cells (Met5A) treated with serum from Libby subjects, and were compared to cells treated with serum cleared of IgG, and therefore containing no MCAA. Analysis of those profiles indicated C/EBP-beta and hypoxia inducible factor 1 alpha (HIF-1α) are significantly increased in the nucleus, indicating activation, due to MCAA exposure compared to controls. Inhibition of either of these transcription factors significantly reduced collagen 1 deposition by these cells following exposure to MCAA. These data suggest autoantibodies are directly involved in type I collagen deposition and may elucidate potential therapeutic targets for autoantibody mediated fibrosis.

  8. Osteopontin promotes fibrosis in dystrophic mouse muscle by modulating immune cell subsets and intramuscular TGF-beta.

    PubMed

    Vetrone, Sylvia A; Montecino-Rodriguez, Encarnacion; Kudryashova, Elena; Kramerova, Irina; Hoffman, Eric P; Liu, Scot D; Miceli, M Carrie; Spencer, Melissa J

    2009-06-01

    Duchenne muscular dystrophy (DMD) is an X-linked, degenerative muscle disease that is exacerbated by secondary inflammation. Here, we characterized the immunological milieu of dystrophic muscle in mdx mice, a model of DMD, to identify potential therapeutic targets. We identified a specific subpopulation of cells expressing the Vbeta8.1/8.2 TCR that is predominant among TCR-beta+ T cells. These cells expressed high levels of osteopontin (OPN), a cytokine that promotes immune cell migration and survival. Elevated OPN levels correlated with the dystrophic process, since OPN was substantially elevated in the serum of mdx mice and muscle biopsies after disease onset. Muscle biopsies from individuals with DMD also had elevated OPN levels. To test the role of OPN in mdx muscle, mice lacking both OPN and dystrophin were generated and termed double-mutant mice (DMM mice). Reduced infiltration of NKT-like cells and neutrophils was observed in the muscle of DMM mice, supporting an immunomodulatory role for OPN in mdx muscle. Concomitantly, an increase in CD4+ and FoxP3+ Tregs was also observed in DMM muscle, which also showed reduced levels of TGF-beta, a known fibrosis mediator. These inflammatory changes correlated with increased strength and reduced diaphragm and cardiac fibrosis. These studies suggest that OPN may be a promising therapeutic target for reducing inflammation and fibrosis in individuals with DMD.

  9. Proteomic Analysis of Nasal Epithelial Cells from Cystic Fibrosis Patients

    PubMed Central

    Papon, Jean-François; Chhuon, Cerina; Zadigue, Patricia; Prulière-Escabasse, Virginie; Amselem, Serge; Escudier, Estelle; Coste, André; Edelman, Aleksander

    2014-01-01

    The pathophysiology of cystic fibrosis (CF) lung disease remains incompletely understood. New explanations for the pathogenesis of CF lung disease may be discovered by studying the patterns of protein expression in cultured human nasal epithelial cells (HNEC). To that aim, we compared the level of protein expressions in primary cultures of HNEC from nasal polyps secondary to CF (CFNP, n = 4), primary nasal polyps (NP, n = 8) and control mucosa (CTRL, n = 4) using isobaric tag for relative and absolute quantification (iTRAQ) labeling coupled with liquid chromatography (LC)-MS-MS. The analysis of the data revealed 42 deregulated protein expressions in CFNP compared to NP and CTRL, suggesting that these alterations are related to CF. Overall, AmiGo analysis highlighted six major pathways important for cell functions that seem to be impaired: metabolism, G protein process, inflammation and oxidative stress response, protein folding, proteolysis and structural proteins. Among them, glucose and fatty acid metabolic pathways could be impaired in CF with nine deregulated proteins. Our proteomic study provides a reproducible set of differentially expressed proteins in airway epithelial cells from CF patients and reveals many novel deregulated proteins that could lead to further studies aiming to clarify the involvement of such proteins in CF pathophysiology. PMID:25268127

  10. Spontaneous in vitro production of rheumatoid factor during infectious exacerbations of cystic fibrosis: correlation with circulating immune complex levels.

    PubMed Central

    Keogan, M T; Callaghan, M; Yanni, G; Mulherin, D; Feighery, C; Brown, D L; Fitzgerald, M X; Bresnihan, B

    1993-01-01

    Rheumatoid factor (RF) production has been demonstrated during infections, including infectious exacerbations of cystic fibrosis (CF). The aim of this study was to evaluate the relationship of RF production to infection, and examine the mechanisms involved. Serial peripheral blood mononuclear cell (PBMC) cultures with measurement of spontaneous production of IgM RF, IgA RF, total IgM and IgA, and measurement of serum levels of immune complexes were carried out during exacerbations of CF. The percentage of B cells expressing CD5 was examined in a second cohort of acutely infected CF patients, and related to IgM RF production. IgM RF production was significantly elevated during acute infection compared with convalescence (P < 0.05), stable CF subjects (P < 0.005) and normal controls (P < 0.05). IgM RF production did not correlate with total IgM production in the majority of patients, but was closely related to circulating immune complex levels in 8/10 subjects. IgA RF production did not increase significantly during infection, and did not correlate with total IgA or IgM RF production, or with circulating immune complex levels. CD5+ B cells were not increased in the CF group, and the percentage of CD5+ B cells did not correlate with IgM RF synthesis. These observations suggest that RF production during infection is specifically induced, possibly by immune complex autoimmunization, and is not simply the result of polyclonal B cell activation. Different patterns of IgM RF and IgA RF synthesis suggest different mechanisms of induction. PMID:7680296

  11. Mutant cystic fibrosis transmembrane conductance regulator inhibits acidification and apoptosis in C127 cells: possible relevance to cystic fibrosis.

    PubMed Central

    Gottlieb, R A; Dosanjh, A

    1996-01-01

    We have shown elsewhere that acidification is an early event in apoptosis, preceding DNA cleavage. Cells expressing the most common mutation (delF508) of the cystic fibrosis transmembrane regulator (CFTR) exhibit a higher resting intracellular pH and are unable to secrete chloride and bicarbonate in response to cAMP. We hypothesized that defective acidification in cells expressing delF508 CFTR would interfere with the acidification that accompanies apoptosis, which in turn, would prevent endonuclease activation and cleavage of DNA. We therefore determined whether the function of the CFTR would affect the process of apoptosis in mouse mammary epithelial C127 cells stably transfected with the wild-type CFTR (C127/wt) or the delF508 mutation of the CFTR (C127/508). C127 cells possessed an acid endonuclease capable of DNA degradation at low pH. Sixteen hours after treatment with cycloheximide, C127/wt cells underwent cytoplasmic acidification. In contrast, C127/508 cells failed to demonstrate acidification. Furthermore, the C127/508 cells did not show nuclear condensation or DNA fragmentation detected by in situ nick-end labeling after treatment with cycloheximide or etoposide, in contrast to the characteristic features of apoptosis demonstrated by the C127/wt cells. Measurement of cell viability indicated a preservation of cell viability in C127/508 cells but not in C127/wt cells. That this resistance to the induction of apoptosis depended upon the loss of CFTR activity is shown by the finding that inhibition of the CFTR with diphenylamine carboxylate in C127/wt cells conferred similar protection. These findings suggest a role for the CFTR in acidification during the initiation of apoptosis in epithelial cells and imply that a failure to undergo programmed cell death could contribute to the pathogenesis of cystic fibrosis. Images Fig. 1 Fig. 2 Fig. 3 PMID:8622979

  12. Calcium channel blockers ameliorate iron overload-associated hepatic fibrosis by altering iron transport and stellate cell apoptosis.

    PubMed

    Zhang, Ying; Zhao, Xin; Chang, Yanzhong; Zhang, Yuanyuan; Chu, Xi; Zhang, Xuan; Liu, Zhenyi; Guo, Hui; Wang, Na; Gao, Yonggang; Zhang, Jianping; Chu, Li

    2016-06-15

    Liver fibrosis is the principal cause of morbidity and mortality in patients with iron overload. Calcium channel blockers (CCBs) can antagonize divalent cation entry into renal and myocardial cells and inhibit fibrogenic gene expression. We investigated the potential of CCBs to resolve iron overload-associated hepatic fibrosis. Kunming mice were assigned to nine groups (n=8 per group): control, iron overload, deferoxamine, high and low dose verapamil, high and low dose nimodipine, and high and low dose diltiazem. Iron deposition and hepatic fibrosis were measured in mouse livers. Expression levels of molecules associated with transmembrane iron transport were determined by molecular biology approaches. In vitro HSC-T6 cells were randomized into nine groups (the same groups as the mice). Changes in proliferation, apoptosis, and metalloproteinase expression in cells were detected to assess the anti-fibrotic effects of CCBs during iron overload conditions. We found that CCBs reduced hepatic iron content, intracellular iron deposition, the number of hepatic fibrotic areas, collagen expression levels, and hydroxyproline content. CCBs rescued abnormal expression of α1C protein in L-type voltage-dependent calcium channel (LVDCC) and down-regulated divalent metal transporter-1 (DMT-1) expression in mouse livers. In iron-overloaded HSC-T6 cells, CCBs reduced iron deposition, inhibited proliferation, induced apoptosis, and elevated expression of matrix metalloproteinase-13 (MMP-13) and tissue inhibitor of metalloproteinase-1 (TIMP-1). CCBs are potential therapeutic agents that can be used to address hepatic fibrosis during iron overload. They resolve hepatic fibrosis probably correlated with regulating transmembrane iron transport and inhibiting HSC growth.

  13. Current status and future prospects of mesenchymal stem cell therapy for liver fibrosis*

    PubMed Central

    Guo, Yang; Chen, Bo; Chen, Li-jun; Zhang, Chun-feng; Xiang, Charlie

    2016-01-01

    Liver fibrosis is the end-stage of many chronic liver diseases and is a significant health threat. The only effective therapy is liver transplantation, which still has many problems, including the lack of donor sources, immunological rejection, and high surgery costs, among others. However, the use of cell therapy is becoming more prevalent, and mesenchymal stem cells (MSCs) seem to be a promising cell type for the treatment of liver fibrosis. MSCs have multiple differentiation abilities, allowing them to migrate directly into injured tissue and differentiate into hepatocyte-like cells. Additionally, MSCs can release various growth factors and cytokines to increase hepatocyte regeneration, regress liver fibrosis, and regulate inflammation and immune responses. In this review, we summarize the current uses of MSC therapies for liver fibrosis and suggest potential future applications. PMID:27819130

  14. B cell activating factor is central to bleomycin- and IL-17-mediated experimental pulmonary fibrosis.

    PubMed

    François, Antoine; Gombault, Aurélie; Villeret, Bérengère; Alsaleh, Ghada; Fanny, Manoussa; Gasse, Paméla; Adam, Sylvain Marchand; Crestani, Bruno; Sibilia, Jean; Schneider, Pascal; Bahram, Seiamak; Quesniaux, Valérie; Ryffel, Bernhard; Wachsmann, Dominique; Gottenberg, Jacques-Eric; Couillin, Isabelle

    2015-01-01

    Idiopathic pulmonary fibrosis (IPF) is a progressive devastating, yet untreatable fibrotic disease of unknown origin. We investigated the contribution of the B-cell activating factor (BAFF), a TNF family member recently implicated in the regulation of pathogenic IL-17-producing cells in autoimmune diseases. The contribution of BAFF was assessed in a murine model of lung fibrosis induced by airway administered bleomycin. We show that murine BAFF levels were strongly increased in the bronchoalveolar space and lungs after bleomycin exposure. We identified Gr1(+) neutrophils as an important source of BAFF upon BLM-induced lung inflammation and fibrosis. Genetic ablation of BAFF or BAFF neutralization by a soluble receptor significantly attenuated pulmonary fibrosis and IL-1β levels. We further demonstrate that bleomycin-induced BAFF expression and lung fibrosis were IL-1β and IL-17A dependent. BAFF was required for rIL-17A-induced lung fibrosis and augmented IL-17A production by CD3(+) T cells from murine fibrotic lungs ex vivo. Finally we report elevated levels of BAFF in bronchoalveolar lavages from IPF patients. Our data therefore support a role for BAFF in the establishment of pulmonary fibrosis and a crosstalk between IL-1β, BAFF and IL-17A.

  15. Liver fibrosis and hepatic stellate cells: Etiology, pathological hallmarks and therapeutic targets

    PubMed Central

    Zhang, Chong-Yang; Yuan, Wei-Gang; He, Pei; Lei, Jia-Hui; Wang, Chun-Xu

    2016-01-01

    Liver fibrosis is a reversible wound-healing process aimed at maintaining organ integrity, and presents as the critical pre-stage of liver cirrhosis, which will eventually progress to hepatocellular carcinoma in the absence of liver transplantation. Fibrosis generally results from chronic hepatic injury caused by various factors, mainly viral infection, schistosomiasis, and alcoholism; however, the exact pathological mechanisms are still unknown. Although numerous drugs have been shown to have antifibrotic activity in vitro and in animal models, none of these drugs have been shown to be efficacious in the clinic. Importantly, hepatic stellate cells (HSCs) play a key role in the initiation, progression, and regression of liver fibrosis by secreting fibrogenic factors that encourage portal fibrocytes, fibroblasts, and bone marrow-derived myofibroblasts to produce collagen and thereby propagate fibrosis. These cells are subject to intricate cross-talk with adjacent cells, resulting in scarring and subsequent liver damage. Thus, an understanding of the molecular mechanisms of liver fibrosis and their relationships with HSCs is essential for the discovery of new therapeutic targets. This comprehensive review outlines the role of HSCs in liver fibrosis and details novel strategies to suppress HSC activity, thereby providing new insights into potential treatments for liver fibrosis. PMID:28082803

  16. Assessment of Correlation between Sweat Chloride Levels and Clinical Features of Cystic Fibrosis Patients

    PubMed Central

    Raina, Manzoor A.; Khan, Mosin S.; Malik, Showkat A.; Raina, AB Hameed; Makhdoomi, Mudassir J.; Bhat, Javed I.

    2016-01-01

    Introduction Cystic Fibrosis (CF) is an autosomal recessive disorder and the incidence of this disease is undermined in Northern India. The distinguishable salty character of the sweat belonging to individuals suffering from CF makes sweat chloride estimation essential for diagnosis of CF disease. Aim The aim of this prospective study was to elucidate the relationship of sweat chloride levels with clinical features and pattern of CF. Materials and Methods A total of 182 patients, with clinical features of CF were included in this study for quantitative measurement of sweat chloride. Sweat stimulation and collection involved pilocarpine iontophoresis based on the Gibson and Cooks methodology. The quantitative estimation of chloride was done by Schales and Schales method with some modifications. Cystic Fibrosis Trans Membrane Conductance Regulator (CFTR) mutation status was recorded in case of patients with borderline sweat chloride levels to correlate the results and for follow-up. Results Out of 182 patients having clinical features consistent with CF, borderline and elevated sweat chloride levels were present in 9 (5%) and 41 (22.5%) subjects respectively. Elevated sweat chloride levels were significantly associated with wheeze, Failure To Thrive (FTT), history of CF in Siblings, product of Consanguineous Marriage (CM), digital clubbing and steatorrhoea on univariate analysis. On multivariate analysis only wheeze, FTT and steatorrhoea were found to be significantly associated with elevated sweat chloride levels (p<0.05). Among the nine borderline cases six cases were positive for at least two CFTR mutations and rest of the three cases were not having any mutation in CFTR gene. Conclusion The diagnosis is often delayed and the disease is advanced in most patients at the time of diagnosis. Sweat testing is a gold standard for diagnosis of CF patients as genetic mutation profile being heterozygous and unlikely to become diagnostic test. PMID:28208841

  17. Cardiac Magnetic Resonance-Verified Myocardial Fibrosis in Chagas Disease: Clinical Correlates and Risk Stratification

    PubMed Central

    Uellendahl, Marly; de Siqueira, Maria Eduarda Menezes; Calado, Eveline Barros; Kalil-Filho, Roberto; Sobral, Dário; Ribeiro, Clébia; Oliveira, Wilson; Martins, Silvia; Narula, Jagat; Rochitte, Carlos Eduardo

    2016-01-01

    Background Chagas disease (CD) is an important cause of heart failure and mortality, mainly in Latin America. This study evaluated the morphological and functional characteristics of the heart as well the extent of myocardial fibrosis (MF) in patients with CD by cardiac magnetic resonance (CMR). The prognostic value of MF evaluated by myocardial-delayed enhancement (MDE) was compared with that via Rassi score. Methods This study assessed 39 patients divided into 2 groups: 28 asymptomatic patients as indeterminate form group (IND); and symptomatic patients as Chagas Heart Disease (CHD) group. All patients underwent CMR using the techniques of cine-MRI and MDE, and the amount of MF was compared with the Rassi score. Results Regarding the morphological and functional analysis, significant differences were observed between both groups (p < 0.001). Furthermore, there was a strong correlation between the extent of MF and the Rassi score (r = 0.76). Conclusions CMR is an important technique for evaluating patients with CD, stressing morphological and functional differences in all clinical presentations. The strong correlation with the Rassi score and the extent of MF detected by CMR emphasizes its role in the prognostic stratification of patients with CD. PMID:27982271

  18. Single-cell RNA sequencing identifies diverse roles of epithelial cells in idiopathic pulmonary fibrosis

    PubMed Central

    Mizuno, Takako; Sridharan, Anusha; Du, Yina; Guo, Minzhe; Wikenheiser-Brokamp, Kathryn A.; Perl, Anne-Karina T.; Funari, Vincent A.; Gokey, Jason J.; Stripp, Barry R.; Whitsett, Jeffrey A.

    2016-01-01

    Idiopathic pulmonary fibrosis (IPF) is a lethal interstitial lung disease characterized by airway remodeling, inflammation, alveolar destruction, and fibrosis. We utilized single-cell RNA sequencing (scRNA-seq) to identify epithelial cell types and associated biological processes involved in the pathogenesis of IPF. Transcriptomic analysis of normal human lung epithelial cells defined gene expression patterns associated with highly differentiated alveolar type 2 (AT2) cells, indicated by enrichment of RNAs critical for surfactant homeostasis. In contrast, scRNA-seq of IPF cells identified 3 distinct subsets of epithelial cell types with characteristics of conducting airway basal and goblet cells and an additional atypical transitional cell that contributes to pathological processes in IPF. Individual IPF cells frequently coexpressed alveolar type 1 (AT1), AT2, and conducting airway selective markers, demonstrating “indeterminate” states of differentiation not seen in normal lung development. Pathway analysis predicted aberrant activation of canonical signaling via TGF-β, HIPPO/YAP, P53, WNT, and AKT/PI3K. Immunofluorescence confocal microscopy identified the disruption of alveolar structure and loss of the normal proximal-peripheral differentiation of pulmonary epithelial cells. scRNA-seq analyses identified loss of normal epithelial cell identities and unique contributions of epithelial cells to the pathogenesis of IPF. The present study provides a rich data source to further explore lung health and disease. PMID:27942595

  19. IL-21 induction of CD4+ T cell differentiation into Th17 cells contributes to bleomycin-induced fibrosis in mice.

    PubMed

    Lei, Ling; Zhong, Xiao-Ning; He, Zhi-Yi; Zhao, Cheng; Sun, Xue-Jiao

    2015-04-01

    Systemic sclerosis (SSc) is a connective tissue disease characterized by fibrosis of the skin and internal organs. Th17 cells and interleukin-17 (also called IL-17A) have been found to be increased in peripheral blood and skin in patients with SSc. IL-21 is a potent inducer of Th17 differentiation that is produced by activated T cells, and whose relationship with Th17 cells in SSc is unclear. Here, using a bleomycin (BLM)-induced mouse model of skin fibrosis, we detected the frequency of CD4+/IL-17+ (Th17) cells, CD4+/IL-21+ T cells and IL-21+ Th17 cells in peripheral blood, skin and lungs, as well as the serum content of IL-17A and IL-21. In addition, we assessed the differentiation of CD4+ T cells cultured from these mice into Th17 cells in response to treatment with IL-21. Compared with the control mice, Th17 cell counts and IL-17A levels were significantly increased and correlated with inflammatory and fibrotic indices in the skin and lungs of the BLM-induced fibrosis mice. Moreover, serum levels of CD4+/IL-21+ T cells, IL-21+ Th17 cells, and IL-21 were significantly increased in these mice, and correlated positively with serum levels of Th17 cells. In vitro experiments showed that IL-21 treated CD4+ T cells derived from BLM-induced mice differentiated into Th17 cells. Our results indicate that Th17 cells and IL-17A contributes to inflammatory and fibrotic processes in the skin and lungs in a BLM-induced mouse model of SSc. Moreover, the expansion of the Th17 cell population may be subsequent to IL-21 promotion of the differentiation of CD4+ T cells in these mice.

  20. CXCR4+ granulocytes reflect fungal cystic fibrosis lung disease.

    PubMed

    Carevic, Melanie; Singh, Anurag; Rieber, Nikolaus; Eickmeier, Olaf; Griese, Matthias; Hector, Andreas; Hartl, Dominik

    2015-08-01

    Cystic fibrosis airways are frequently colonised with fungi. However, the interaction of these fungi with immune cells and the clinical relevance in cystic fibrosis lung disease are incompletely understood.We characterised granulocytes in airway fluids and peripheral blood from cystic fibrosis patients with and without fungal colonisation, non-cystic fibrosis disease controls and healthy control subjects cross-sectionally and longitudinally and correlated these findings with lung function parameters.Cystic fibrosis patients with chronic fungal colonisation by Aspergillus fumigatus were characterised by an accumulation of a distinct granulocyte subset, expressing the HIV coreceptor CXCR4. Percentages of airway CXCR4(+) granulocytes correlated with lung disease severity in patients with cystic fibrosis.These studies demonstrate that chronic fungal colonisation with A. fumigatus in cystic fibrosis patients is associated with CXCR4(+) airway granulocytes, which may serve as a potential biomarker and therapeutic target in fungal cystic fibrosis lung disease.

  1. Free DNA in Cystic Fibrosis Airway Fluids Correlates with Airflow Obstruction

    PubMed Central

    Marcos, Veronica; Zhou-Suckow, Zhe; Önder Yildirim, Ali; Bohla, Alexander; Hector, Andreas; Vitkov, Ljubomir; Krautgartner, Wolf Dietrich; Stoiber, Walter; Griese, Matthias; Eickelberg, Oliver; Mall, Marcus A.; Hartl, Dominik

    2015-01-01

    Chronic obstructive lung disease determines morbidity and mortality of patients with cystic fibrosis (CF). CF airways are characterized by a nonresolving neutrophilic inflammation. After pathogen contact or prolonged activation, neutrophils release DNA fibres decorated with antimicrobial proteins, forming neutrophil extracellular traps (NETs). NETs have been described to act in a beneficial way for innate host defense by bactericidal, fungicidal, and virucidal actions. On the other hand, excessive NET formation has been linked to the pathogenesis of autoinflammatory and autoimmune disease conditions. We quantified free DNA structures characteristic of NETs in airway fluids of CF patients and a mouse model with CF-like lung disease. Free DNA levels correlated with airflow obstruction, fungal colonization, and CXC chemokine levels in CF patients and CF-like mice. When viewed in combination, our results demonstrate that neutrophilic inflammation in CF airways is associated with abundant free DNA characteristic for NETosis, and suggest that free DNA may be implicated in lung function decline in patients with CF. PMID:25918476

  2. Hepatic Stellate Cells and microRNAs in Pathogenesis of Liver Fibrosis

    PubMed Central

    Kitano, Mio; Bloomston, P. Mark

    2016-01-01

    microRNAs (miRNAs) are small non-coding RNAs that regulate gene expression by either blocking translation or inducing degradation of target mRNA. miRNAs play essential roles in diverse biological and pathological processes, including development of hepatic fibrosis. Hepatic stellate cells (HSCs) play a central role in development of hepatic fibrosis and there are intricate regulatory effects of miRNAs on their activation, proliferation, collagen production, migration, and apoptosis. There are multiple differentially expressed miRNAs in activated HSCs, and in this review we aim to summarize current data on miRNAs that participate in the development of hepatic fibrosis. Based on this review, miRNAs may serve as biomarkers for diagnosis of liver disease, as well as markers of disease progression. Most importantly, dysregulated miRNAs may potentially be targeted by novel therapies to treat and reverse progression of hepatic fibrosis. PMID:26999230

  3. Chemokine receptor CXCR6-dependent hepatic NK T Cell accumulation promotes inflammation and liver fibrosis.

    PubMed

    Wehr, Alexander; Baeck, Christer; Heymann, Felix; Niemietz, Patricia Maria; Hammerich, Linda; Martin, Christian; Zimmermann, Henning W; Pack, Oliver; Gassler, Nikolaus; Hittatiya, Kanishka; Ludwig, Andreas; Luedde, Tom; Trautwein, Christian; Tacke, Frank

    2013-05-15

    Chronic liver injury characteristically results in hepatic inflammation, which represents a prerequisite for organ fibrosis. Although NKT cells are abundantly present in liver and involved in hepatic inflammation, molecular mechanisms of their recruitment in liver fibrosis remained elusive. We hypothesized that chemokine receptor CXCR6 and its ligand CXCL16 control NKT cell migration and functionality in liver fibrosis. In patients with chronic liver diseases (n = 58), CXCR6 and CXCL16 expression was intrahepatically upregulated compared with controls. In murine liver, Cxcl16 was strongly expressed by endothelium and macrophages, whereas lymphocyte populations (NKT, NK, CD4 T, CD8 T cells) expressed CXCR6. Intravital two-photon microscopy imaging of Cxcr6(+/gfp) and Cxcr6(gfp/gfp) mice and chemotaxis studies in vitro revealed that CXCR6 specifically controls hepatic NKT cell accumulation during the early response upon experimental liver damage. Hepatic invariant NKT cells expressed distinct proinflammatory cytokines including IFN-γ and IL-4 upon injury. CXCR6-deficient mice were protected from liver fibrosis progression in two independent experimental models. Macrophage infiltration and protein levels of inflammatory cytokines IFN-γ, TNF-α, and IL-4 were also reduced in fibrotic livers of Cxcr6(-/-) mice, corroborating that hepatic NKT cells provide essential cytokine signals perpetuating hepatic inflammation and fibrogenesis. Adoptive transfer of NKT cells, but not CD4 T cells, isolated from wild type livers restored hepatic fibrosis in Cxcr6(-/-) mice upon experimental steatohepatitis. Our results demonstrate that hepatic NKT cells accumulate CXCR6-dependent early upon injury, thereby accentuating the inflammatory response in the liver and promoting hepatic fibrogenesis. Interfering with CXCR6/CXCL16 might therefore bear therapeutic potential in liver fibrosis.

  4. mTOR Overactivation in Mesenchymal cells Aggravates CCl4− Induced liver Fibrosis

    PubMed Central

    Shan, Lanlan; Ding, Yan; Fu, You; Zhou, Ling; Dong, Xiaoying; Chen, Shunzhi; Wu, Hongyuan; Nai, Wenqing; Zheng, Hang; Xu, Wanfu; Bai, Xiaochun; Jia, Chunhong; Dai, Meng

    2016-01-01

    Hepatic stellate cells are of mesenchymal cell type located in the space of Disse. Upon liver injury, HSCs transactivate into myofibroblasts with increase in expression of fibrillar collagen, especially collagen I and III, leading to liver fibrosis. Previous studies have shown mTOR signaling is activated during liver fibrosis. However, there is no direct evidence in vivo. The aim of this study is to examine the effects of conditional deletion of TSC1 in mesenchymal on pathogenesis of liver fibrosis. Crossing mice bearing the floxed TSC1 gene with mice harboring Col1α2-Cre-ER(T) successfully generated progeny with a conditional knockout of TSC1 (TSC1 CKO) in collagen I expressing mesenchymal cells. TSC1 CKO and WT mice were subjected to CCl4, oil or CCl4+ rapamycin treatment for 8 weeks. TSC1 CKO mice developed pronounced liver fibrosis relative to WT mice, as examined by ALT, hydroxyproline, histopathology, and profibrogenic gene. Absence of TSC1 in mesenchymal cells induced proliferation and prevented apoptosis in activated HSCs. However, there were no significant differences in oil-treated TSC1 CKO and WT mice. Rapamycin, restored these phenotypic changes by preventing myofibroblasts proliferation and enhancing their apoptosis. These findings revealed mTOR overactivation in mesenchymal cells aggravates CCl4− induced liver fibrosis and the rapamycin prevent its occurance. PMID:27819329

  5. Cystic fibrosis transmembrane conductance regulator and caveolin-1 regulate epithelial cell internalization of Pseudomonas aeruginosa

    PubMed Central

    Bajmoczi, Milan; Gadjeva, Mihaela; Alper, Seth L.; Pier, Gerald B.; Golan, David E.

    2009-01-01

    Patients with cystic fibrosis (CF) exhibit defective innate immunity and are susceptible to chronic lung infection with Pseudomonas aeruginosa. To investigate the molecular bases for the hypersusceptibility of CF patients to P. aeruginosa, we used the IB3-1 cell line with two defective CF transmembrane conductance regulator (CFTR) genes (ΔF508/W1282X) to generate isogenic stable, clonal lung epithelial cells expressing wild-type (WT)-CFTR with an NH2-terminal green fluorescent protein (GFP) tag. GFP-CFTR exhibited posttranslational modification, subcellular localization, and anion transport function typical of WT-CFTR. P. aeruginosa internalization, a component of effective innate immunity, required functional CFTR and caveolin-1, as shown by: 1) direct correlation between GFP-CFTR expression levels and P. aeruginosa internalization; 2) enhanced P. aeruginosa internalization by aminoglycoside-induced read through of the CFTR W1282X allele in IB3-1 cells; 3) decreased P. aeruginosa internalization following siRNA knockdown of GFP-CFTR or caveolin-1; and 4) spatial association of P. aeruginosa with GFP-CFTR and caveolin-1 at the cell surface. P. aeruginosa internalization also required free lateral diffusion of GFP-CFTR, allowing for bacterial coclustering with GFP-CFTR and caveolin-1 at the plasma membrane. Thus efficient initiation of innate immunity to P. aeruginosa requires formation of an epithelial “internalization platform” involving both caveolin-1 and functional, laterally mobile CFTR. PMID:19386787

  6. Cystic fibrosis transmembrane conductance regulator and caveolin-1 regulate epithelial cell internalization of Pseudomonas aeruginosa.

    PubMed

    Bajmoczi, Milan; Gadjeva, Mihaela; Alper, Seth L; Pier, Gerald B; Golan, David E

    2009-08-01

    Patients with cystic fibrosis (CF) exhibit defective innate immunity and are susceptible to chronic lung infection with Pseudomonas aeruginosa. To investigate the molecular bases for the hypersusceptibility of CF patients to P. aeruginosa, we used the IB3-1 cell line with two defective CF transmembrane conductance regulator (CFTR) genes (DeltaF508/W1282X) to generate isogenic stable, clonal lung epithelial cells expressing wild-type (WT)-CFTR with an NH(2)-terminal green fluorescent protein (GFP) tag. GFP-CFTR exhibited posttranslational modification, subcellular localization, and anion transport function typical of WT-CFTR. P. aeruginosa internalization, a component of effective innate immunity, required functional CFTR and caveolin-1, as shown by: 1) direct correlation between GFP-CFTR expression levels and P. aeruginosa internalization; 2) enhanced P. aeruginosa internalization by aminoglycoside-induced read through of the CFTR W1282X allele in IB3-1 cells; 3) decreased P. aeruginosa internalization following siRNA knockdown of GFP-CFTR or caveolin-1; and 4) spatial association of P. aeruginosa with GFP-CFTR and caveolin-1 at the cell surface. P. aeruginosa internalization also required free lateral diffusion of GFP-CFTR, allowing for bacterial coclustering with GFP-CFTR and caveolin-1 at the plasma membrane. Thus efficient initiation of innate immunity to P. aeruginosa requires formation of an epithelial "internalization platform" involving both caveolin-1 and functional, laterally mobile CFTR.

  7. Effect of adipose tissue-derived stem cell injection in a rat model of urethral fibrosis

    PubMed Central

    Sangkum, Premsant; Yafi, Faysal A.; Kim, Hogyoung; Bouljihad, Mostafa; Ranjan, Manish; Datta, Amrita; Mandava, Sree Harsha; Sikka, Suresh C; Abdel-Mageed, Asim B.; Hellstrom, Wayne J.G.

    2016-01-01

    Introduction: We sought to evaluate the therapeutic effect of adi-pose tissue-derived stem cells (ADSCs) in a rat model of urethral fibrosis. Methods: Eighteen (18) male Sprague-Dawley rats (300‒350 g) were divided into three groups: (1) sham (saline injection); (2) urethral fibrosis group (10 μg transforming growth factor beta 1 (TGF-β1) injection); and (3) ADSCs group (10 μg TGF-β1 injection plus 2 × 105 ADSCs). Rat ADSCs were harvested from rat inguinal fat pads. All study animals were euthanized at two weeks after urethral injection. Following euthanasia, rat urethral tissue was harvested for histologic evaluation. Type I and III collagen levels were quantitated by Western blot analysis. Results: TGF-β1 injection induced significant urethral fibrosis and increased collagen type I and III expression (p<0.05). Significant decrease in submucosal fibrosis and collagen type I and III expression were noted in the ADSCs group compared with the urethral fibrosis group (p<0.05). TGF-β1 induced fibrotic changes were ameliorated by injection of ADSCs. Conclusions: Local injection of ADSCs in a rat model of urethral fibrosis significantly decreased collagen type I and III. These findings suggest that ADSC injection may prevent scar formation and potentially serve as an adjunct treatment to increase the success rate of primary treatment for urethral stricture disease. Further animal and clinical studies are needed to confirm these results. PMID:27790299

  8. Endothelial cell Toll-like receptor 4 regulates fibrosis associated angiogenesis in liver

    PubMed Central

    Jagavelu, K; Routray, C; Shergill, U; O’Hara, SP; Faubion, W; Shah, VH

    2010-01-01

    Angiogenesis defines the growth of new blood vessels from pre-existing vascular endothelial networks and corresponds with the wound healing process that is typified by the process of liver fibrosis. Liver fibrosis is also associated with increased endotoxin within the gut lumen and its associated portal circulation. However, the interrelationship of gut endotoxin and its receptor, Toll-like receptor 4 (TLR4), with liver fibrosis and associated angiogenesis remains incompletely defined. RESULT Here we provide evidence, using complementary genetic, molecular, and pharmacologic approaches that the pattern recognition receptor that recognizes endotoxin, TLR4, expressed on liver endothelial cells (LEC), regulates angiogenic responses both in vitro and in vivo. Mechanistic studies reveal a key role for a cognate TLR4 effector protein, MyD88 in this process which culminates in extracellular protease production that regulates LEC invasive capacity, a key step in angiogenesis. Furthermore TLR4 dependent angiogenesis in vivo corresponds with fibrosis in complementary liver models of fibrosis. CONCLUSION These studies provide evidence that the TLR4 pathway in LEC regulates angiogenesis through its MyD88 effector protein by regulating extracellular protease production and that this process is linked to the development of liver fibrosis. PMID:20564354

  9. Liver fibrosis secondary to bile duct injury: correlation of Smad7 with TGF-β and extracellular matrix proteins

    PubMed Central

    2009-01-01

    Background Liver fibrosis is the result of continuous liver injury stemming from different etiological factors. Bile duct injury induces an altered expression of TGF-β, which has an important role in liver fibrosis because this cytokine induces the expression of target genes such as collagens, PAI-1, TIMPs, and others that lead to extracellular matrix deposition. Smad7 is the principal inhibitor that regulates the target gene transcription of the TGF-β signaling. The aim of the study was to determine whether Smad7 mRNA expression correlates with the gene expression of TGF-β, Col I, Col III, Col IV, or PAI-1 in liver fibrosis secondary to bile duct injury (BDI). Results Serum TGF-β concentration was higher in BDI patients (39 296 pg/ml) than in liver donors (9008 pg/ml). Morphometric analysis of liver sections showed 41.85% of tissue contained fibrotic deposits in BDI patients. mRNA expression of Smad7, Col I, and PAI-1 was also significantly higher (P < 0.05) in patients with BDI than in controls. Smad7 mRNA expression correlated significantly with TGF-β concentration, Col I and Col III expression, and the amount of fibrosis. Conclusion We found augmented serum concentration of TGF-β and an increase in the percentage of fibrotic tissue in the liver of BDI patients. Contrary to expected results, the 6-fold increase in Smad7 expression did not inhibit the expression of TGF-β, collagens, and PAI-1. We also observed greater expression of Col I and Col III mRNA in BDI patients and significant correlations between their expression and TGF-β concentration and Smad7 mRNA expression. PMID:19878580

  10. Umbilical cord-derived mesenchymal stem cells alleviate liver fibrosis in rats

    PubMed Central

    Chai, Ning-Li; Zhang, Xiao-Bin; Chen, Si-Wen; Fan, Ke-Xing; Linghu, En-Qiang

    2016-01-01

    AIM: To evaluate the efficacy of umbilical cord-derived mesenchymal stem cells (UC-MSCs) transplantation in the treatment of liver fibrosis. METHODS: Cultured human UC-MSCs were isolated and transfused into rats with liver fibrosis induced by dimethylnitrosamine (DMN). The effects of UC-MSCs transfusion on liver fibrosis were then evaluated by histopathology; serum interleukin (IL)-4 and IL-10 levels were also measured. Furthermore, Kupffer cells (KCs) in fibrotic livers were isolated and cultured to analyze their phenotype. Moreover, UC-MSCs were co-cultured with KCs in vitro to assess the effects of UC-MSCs on KCs’ phenotype, and IL-4 and IL-10 levels were measured in cell culture supernatants. Finally, UC-MSCs and KCs were cultured in the presence of IL-4 antibodies to block the effects of this cytokine, followed by phenotypical analysis of KCs. RESULTS: UC-MSCs transfused into rats were recruited by the injured liver and alleviated liver fibrosis, increasing serum IL-4 and IL-10 levels. Interestingly, UC-MSCs promoted mobilization of KCs not only in fibrotic livers, but also in vitro. Co-culture of UC-MSCs with KCs resulted in increased production of IL-4 and IL-10. The addition of IL-4 antibodies into the co-culture system resulted in decreased KC mobilization. CONCLUSION: UC-MSCs could increase IL-4 and promote mobilization of KCs both in vitro and in vivo, subsequently alleviating the liver fibrosis induced by DMN. PMID:27468195

  11. IL-21 Promotes Pulmonary Fibrosis through the Induction of Pro-fibrotic CD8+ T Cells

    PubMed Central

    Brodeur, Tia Y.; Robidoux, Tara E.; Weinstein, Jason S.; Craft, Joseph; Swain, Susan L.; Marshak-Rothstein, Ann

    2015-01-01

    Type 2 effector production of IL-13, a demonstrated requirement in models of fibrosis, is routinely ascribed to CD4+ Th2 cells. We now demonstrate a major role for CD8+ T cells in a murine model of sterile lung injury. These pulmonary CD8+ T cells differentiate into IL-13-producing Tc2 and play a major role in a bleomycin-induced model of fibrosis. Differentiation of these Tc2 cells in the lung requires IL-21, and bleomycin treated IL-21- and IL-21R-deficient mice develop inflammation but not fibrosis. Moreover, IL-21R-expressing CD8+ cells are sufficient to reconstitute the fibrotic response in the IL-21R-deficient mice. We further show that the combination of IL-4 and IL-21 skews naïve CD8+ T cells to produce IL-21, which in turn acts in an autocrine manner to support robust IL-13 production. Our data reveal a novel pathway involved in the onset and regulation of pulmonary fibrosis, and identify Tc2 cells as key mediators of fibrogenesis. PMID:26519529

  12. Gas6/Axl pathway is activated in chronic liver disease and its targeting reduces fibrosis via hepatic stellate cell inactivation

    PubMed Central

    Bárcena, Cristina; Stefanovic, Milica; Tutusaus, Anna; Joannas, Leonel; Menéndez, Anghara; García-Ruiz, Carmen; Sancho-Bru, Pau; Marí, Montserrat; Caballeria, Joan; Rothlin, Carla V.; Fernández-Checa, José C.; de Frutos, Pablo García; Morales, Albert

    2015-01-01

    Background & Aims Liver fibrosis, an important health concern associated to chronic liver injury that provides a permissive environment for cancer development, is characterized by accumulation of extracellular matrix components mainly derived from activated hepatic stellate cells (HSCs). Axl, a receptor tyrosine kinase, and its ligand Gas6 are involved in cell differentiation, immune response and carcinogenesis. Methods HSCs were obtained from wild type and Axl−/− mice, treated with recombinant Gas6 protein (rGas6), Axl siRNAs or the Axl inhibitor BGB324, and analyzed by western blot and real-time PCR. Experimental fibrosis was studied in CCl4-treated wild type and Axl−/− mice, and in combination with Axl inhibitor. Gas6 and Axl serum levels were measured in alcoholic liver disease (ALD) and hepatitis C virus (HCV) patients. Results In primary mouse HSCs, Gas6 and Axl levels paralleled HSC activation. rGas6 phosphorylated Axl and AKT prior to HSC phenotypic changes, while Axl siRNA silencing reduced HSC activation. Moreover, BGB324 blocked Axl/AKT phosphorylation and diminished HSC activation. In addition, Axl KO mice displayed decreased HSC activation in vitro and liver fibrogenesis after chronic damage by CCl4 administration. Similarly, BGB324 reduced collagen deposition and CCl4-induced liver fibrosis in mice. Importantly, Gas6 and Axl serum levels increased in ALD and HCV patients, inversely correlating with liver functionality. Conclusions: The Gas6/Axl axis is required for full HSC activation. Gas6 and Axl serum levels increase in parallel to chronic liver disease progression. Axl targeting may be a therapeutic strategy for liver fibrosis management. PMID:25908269

  13. Comparison of Histochemical Stainings in Evaluation of Liver Fibrosis and Correlation with Transient Elastography in Chronic Hepatitis

    PubMed Central

    Cabibi, Daniela; Bronte, Fabrizio; Porcasi, Rossana; Ingrao, Sabrina; Giannone, Antonino Giulio; Maida, Marcello; Grazia Bavetta, Maria; Petta, Salvatore; Di Marco, Vito; Calvaruso, Vincenza

    2015-01-01

    Background and Aim. The best staining to evaluate liver fibrosis in liver hepatitis is still a debated topic. This study aimed to compare Masson's trichrome (MT), Sirius Red (SR), and orcein stainings in evaluating liver fibrosis in chronic HCV hepatitis (CHC) with semiquantitative and quantitative methods (Collagen Proportionate Area (CPA) by Digital Image Analysis (DIA)) and correlate them with transient elastography (TE). Methods. Liver stiffness evaluation of 111 consecutive patients with CHC was performed by TE. Semiquantitative staging by Metavir score system and CPA by DIA were assessed on liver biopsy stained with MT, SR, and orcein. Results. MT, SR, and orcein staining showed concordant results in 89.6% of cases in staging CHC, without significant difference in both semiquantitative and quantitative evaluations of fibrosis. TE values were concordant with orcein levels in 86.5% of the cases and with MT/RS in 77.5% (P < 0.001). No significant correlation between the grade of necroinflammatory activity and TE values was found. Conclusion. In CHC, SR/MT and orcein stainings are almost concordant and when discordant, orcein staining is better related to TE values than MT/RS. This suggests that elastic fibers play a more important role than reticular or collagenous ones in determining stiffness values in CHC. PMID:26665101

  14. Predictive Factors of Late Radiation Fibrosis: A Prospective Study in Non-Small Cell Lung Cancer

    SciTech Connect

    Mazeron, Renaud; Etienne-Mastroianni, Benedicte; Perol, David; Arpin, Dominique; Vincent, Michel; Falchero, Lionel; Martel-Lafay, Isabelle; Carrie, Christian; Claude, Line

    2010-05-01

    Purpose: To determine predictive factors of late radiation fibrosis (RF) after conformal radiotherapy (3D-RT) in non-small cell lung cancer (NSCLC). Methods and Materials: Ninety-six patients with Stage IA-IIIB NSCLC were included in a prospective trial. Clinical evaluation, chest X-ray, and pulmonary functional tests including diffusion parameters were performed before and 6 months after radiotherapy. An independent panel of experts prospectively analyzed RF, using Late Effects in Normal Tissues-Subjective, Objective, Management and Analytic scales classification. Logistic regression analysis was performed to identify relationships between clinical, functional, or treatment parameters and incidence of RF. Variations of circulating serum levels of pro-inflammatory (interleukin-6, tumor necrosis factor alpha, tumor growth factor beta1) and anti-inflammatory (interleukin-10) cytokines during 3D-RT were examined to identify correlations with RF. Results: Of the 96 patients included, 72 were evaluable for RF at 6 months. Thirty-seven (51.4%) developed RF (Grade >=1), including six severe RF (Grades 2-3; 8.3%). In univariate analysis, only poor Karnofsky Performance Status and previous acute radiation pneumonitis were associated with RF (p < 0.05). Dosimetric factors (mean lung dose, percentage of lung volume receiving more than 10, 20, 30, 40, and 50 Gy) were highly correlated with RF (p < 0.001). In multivariate analysis, previous acute radiation pneumonitis and dosimetric parameters were significantly correlated with RF occurrence. It was not significantly correlated either with cytokines at baseline or with their variation during 3D-RT. Conclusions: This study confirms the importance of dosimetric parameters to limit the risk of RF. Contrary to acute radiation pneumonitis, RF was not correlated to cytokine variations during 3D-RT.

  15. Gremlin-1 Overexpression in Mouse Lung Reduces Silica-Induced Lymphocyte Recruitment – A Link to Idiopathic Pulmonary Fibrosis through Negative Correlation with CXCL10 Chemokine

    PubMed Central

    Koli, Katri; Sutinen, Eva; Rönty, Mikko; Rantakari, Pia; Fortino, Vittorio; Pulkkinen, Ville; Greco, Dario; Sipilä, Petra; Myllärniemi, Marjukka

    2016-01-01

    Idiopathic pulmonary fibrosis (IPF) is characterized by activation and injury of epithelial cells, the accumulation of connective tissue and changes in the inflammatory microenvironment. The bone morphogenetic protein (BMP) inhibitor protein gremlin-1 is associated with the progression of fibrosis both in human and mouse lung. We generated a transgenic mouse model expressing gremlin-1 in type II lung epithelial cells using the surfactant protein C (SPC) promoter and the Cre-LoxP system. Gremlin-1 protein expression was detected specifically in the lung after birth and did not result in any signs of respiratory insufficiency. Exposure to silicon dioxide resulted in reduced amounts of lymphocyte aggregates in transgenic lungs while no alteration in the fibrotic response was observed. Microarray gene expression profiling and analyses of bronchoalveolar lavage fluid cytokines indicated a reduced lymphocytic response and a downregulation of interferon-induced gene program. Consistent with reduced Th1 response, there was a downregulation of the mRNA and protein expression of the anti-fibrotic chemokine CXCL10, which has been linked to IPF. In human IPF patient samples we also established a strong negative correlation in the mRNA expression levels of gremlin-1 and CXCL10. Our results suggest that in addition to regulation of epithelial-mesenchymal crosstalk during tissue injury, gremlin-1 modulates inflammatory cell recruitment and anti-fibrotic chemokine production in the lung. PMID:27428020

  16. Deletion of ADORA2B from myeloid cells dampens lung fibrosis and pulmonary hypertension.

    PubMed

    Karmouty-Quintana, Harry; Philip, Kemly; Acero, Luis F; Chen, Ning-Yuan; Weng, Tingting; Molina, Jose G; Luo, Fayong; Davies, Jonathan; Le, Ngoc-Bao; Bunge, Isabelle; Volcik, Kelly A; Le, Thanh-Thuy T; Johnston, Richard A; Xia, Yang; Eltzschig, Holger K; Blackburn, Michael R

    2015-01-01

    Idiopathic pulmonary fibrosis (IPF) is a lethal, fibroproliferative disease. Pulmonary hypertension (PH) can develop secondary to IPF and increase mortality. Alternatively, activated macrophages (AAMs) contribute to the pathogenesis of both IPF and PH. Here we hypothesized that adenosine signaling through the ADORA2B on AAMs impacts the progression of these disorders and that conditional deletion of ADORA2B on myeloid cells would have a beneficial effect in a model of these diseases. Conditional knockout mice lacking ADORA2B on myeloid cells (Adora2B(f/f)-LysM(Cre)) were exposed to the fibrotic agent bleomycin (BLM; 0.035 U/g body weight, i.p.). At 14, 17, 21, 25, or 33 d after exposure, SpO2, bronchoalveolar lavage fluid (BALF), and histologic analyses were performed. On day 33, lung function and cardiovascular analyses were determined. Markers for AAM and mediators of fibrosis and PH were assessed. Adora2B(f/f)-LysM(Cre) mice presented with attenuated fibrosis, improved lung function, and no evidence of PH compared with control mice exposed to BLM. These findings were accompanied by reduced expression of CD206 and arginase-1, markers for AAMs. A 10-fold reduction in IL-6 and a 5-fold decrease in hyaluronan, both linked to lung fibrosis and PH, were also observed. These data suggest that activation of the ADORA2B on macrophages plays an active role in the pathogenesis of lung fibrosis and PH.

  17. Activation of calpain by renin-angiotensin system in pleural mesothelial cells mediates tuberculous pleural fibrosis

    PubMed Central

    Yang, Jie; Xiang, Fei; Cai, Peng-Cheng; Lu, Yu-Zhi; Xu, Xiao-Xiao; Yu, Fan; Li, Feng-Zhi; Greer, Peter A.; Shi, Huan-Zhong; Zhou, Qiong; Xin, Jian-Bao; Ye, Hong; Su, Yunchao

    2016-01-01

    Pleural fibrosis is defined as an excessive deposition of extracellular matrix (ECM) components that results in destruction of the normal pleural tissue architecture. It can result from diverse inflammatory conditions, especially tuberculous pleurisy. Pleural mesothelial cells (PMCs) play a pivotal role in pleural fibrosis. Calpain is a family of calcium-dependent endopeptidases, which plays an important role in ECM remodeling. However, the role of calpain in pleural fibrosis remains unknown. In the present study, we found that tuberculous pleural effusion (TPE) induced calpain activation in PMCs and that inhibition of calpain prevented TPE-induced collagen-I synthesis and cell proliferation of PMCs. Moreover, our data revealed that the levels of angiotensin (ANG)-converting enzyme (ACE) were significantly higher in pleural fluid of patients with TPE than those with malignant pleural effusion, and ACE-ANG II in TPE resulted in activation of calpain and subsequent triggering of the phosphatidylinositol 3-kinase (PI3K)/Akt/NF-κB signaling pathway in PMCs. Finally, calpain activation in PMCs and collagen depositions were confirmed in pleural biopsy specimens from patients with tuberculous pleurisy. Together, these studies demonstrated that calpain is activated by renin-angiotensin system in pleural fibrosis and mediates TPE-induced collagen-I synthesis and proliferation of PMCs via the PI3K/Akt/NF-κB signaling pathway. Calpain in PMCs might be a novel target for intervention in tuberculous pleural fibrosis. PMID:27261452

  18. The Role of Mitochondrial DNA in Mediating Alveolar Epithelial Cell Apoptosis and Pulmonary Fibrosis

    PubMed Central

    Kim, Seok-Jo; Cheresh, Paul; Jablonski, Renea P.; Williams, David B.; Kamp, David W.

    2015-01-01

    Convincing evidence has emerged demonstrating that impairment of mitochondrial function is critically important in regulating alveolar epithelial cell (AEC) programmed cell death (apoptosis) that may contribute to aging-related lung diseases, such as idiopathic pulmonary fibrosis (IPF) and asbestosis (pulmonary fibrosis following asbestos exposure). The mammalian mitochondrial DNA (mtDNA) encodes for 13 proteins, including several essential for oxidative phosphorylation. We review the evidence implicating that oxidative stress-induced mtDNA damage promotes AEC apoptosis and pulmonary fibrosis. We focus on the emerging role for AEC mtDNA damage repair by 8-oxoguanine DNA glycosylase (OGG1) and mitochondrial aconitase (ACO-2) in maintaining mtDNA integrity which is important in preventing AEC apoptosis and asbestos-induced pulmonary fibrosis in a murine model. We then review recent studies linking the sirtuin (SIRT) family members, especially SIRT3, to mitochondrial integrity and mtDNA damage repair and aging. We present a conceptual model of how SIRTs modulate reactive oxygen species (ROS)-driven mitochondrial metabolism that may be important for their tumor suppressor function. The emerging insights into the pathobiology underlying AEC mtDNA damage and apoptosis is suggesting novel therapeutic targets that may prove useful for the management of age-related diseases, including pulmonary fibrosis and lung cancer. PMID:26370974

  19. Amelioration of Murine Schistosoma mansoni Induced Liver Fibrosis by Mesenchymal Stem Cells.

    PubMed

    Abdel Aziz, Mt; Atta, Hm; Roshdy, Nk; Rashed, LA; Sabry, D; Hassouna, Aa; Aboul Fotouh, Gi; Hasan, Nm; Younis, Rh; Chowdhury, Jr

    2012-01-01

    Schistosomiasis is a common chronic helminthic infection of the liver that causes hepatic fibrosis and portal hypertension,contributing to the death of over half a million people a year. Infusion of autologous bone marrow cells into patients with hepatic cirrhosis has been reported to ameliorate symptoms of portal hypertension and improve liver function, either by conversion of the infused mesenchymal stem cells (MSCs) to hepatocytes or by modulating of the hepatic fibrosis. Here,we have investigated the antifibrotic effect of mesenchymal stem cells (MSCs) using S. mansoni-induced liver fibrosis in mice, which causes an intense, stable fibrosis. MSCs derived from bone marrow of male mice were then infused intravenously into female mice that had received intraperitoneal injection of S.mansoni cercariae. Mice were divided into 4 groups: Untreated control; MSCs infusion only; Schistosomiasis only; and Schistosomiasis plus MSCs infusion. Serum alanine aminotransferase (ALT) and liver histopathology were evaluated. Expression of the collagen gene (type I),transforming growth factor (TGF-β), matrix metalloproteinase (MMP2), tissue inhibitor of metalloproteinase (TIMP-1),stromal cell-derived factor-1(SDF-1) and its receptor (CXCR4) were analyzed. MSC infusion resulted in significant decrease in liver collagen and TGF-β gene expression in the Schistosomiasis mice. The ratio of MMP-2 to TIMP-1 expression increased. SDF-1 and CXCR4 mRNA expression also increased. There was overall improvement of liver histology and a statistically significant reduction of serum ALT level. MSCs infusion ameliorated S. mansoni-induced liver fibrosis, probably by modulating the relative expression of MMP and TIMP. The findings support the hypothesis that MSCs participate in liver regeneration and functional improvement by reducing liver fibrosis.

  20. IgG antibodies to Aspergillus fumigatus in cystic fibrosis: a laboratory correlate of disease activity.

    PubMed Central

    Forsyth, K D; Hohmann, A W; Martin, A J; Bradley, J

    1988-01-01

    Serum was collected from 50 patients with cystic fibrosis, and IgG antibodies to Aspergillus fumigatus were measured by enzyme linked immunosorbent assay (ELISA). In addition, total IgE and Aspergillus specific IgE antibodies were measured in 41 of the 50. A close association was found between pulmonary function and clinical state, and IgG antibodies to Aspergillus. There was no association between pulmonary function or clinical state and IgE antibodies. It is postulated that in patients with cystic fibrosis, Aspergillus fumigatus may contribute to deterioration in pulmonary function by local pathogenicity, or by hypersensitivity mechanisms mediated by IgG. PMID:3046514

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

    PubMed

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

    2014-09-01

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

  2. Treatment with 4-Methylpyrazole Modulated Stellate Cells and Natural Killer Cells and Ameliorated Liver Fibrosis in Mice

    PubMed Central

    Lee, Young-Sun; Jung, Ju Yeon; Park, Seol-Hee; Park, Keun-Gyu; Choi, Hueng-Sik; Suh, Jae Myoung; Jeong, Won-Il

    2015-01-01

    Background & Aims Accumulating evidence suggests that retinol and its metabolites are closely associated with liver fibrogenesis. Recently, we demonstrated that genetic ablation of alcohol dehydrogenase 3 (ADH3), a retinol metabolizing gene that is expressed in hepatic stellate cells (HSCs) and natural killer (NK) cells, attenuated liver fibrosis in mice. In the current study, we investigated whether pharmacological ablation of ADH3 has therapeutic effects on experimentally induced liver fibrosis in mice. Methods Liver fibrosis was induced by intraperitoneal injections of carbon tetrachloride (CCl4) or bile duct ligation (BDL) for two weeks. To inhibit ADH3-mediated retinol metabolism, 10 μg 4-methylpyrazole (4-MP)/g of body weight was administered to mice treated with CCl4 or subjected to BDL. The mice were sacrificed at week 2 to evaluate the regression of liver fibrosis. Liver sections were stained for collagen and α-smooth muscle actin (α-SMA). In addition, HSCs and NK cells were isolated from control and treated mice livers for molecular and immunological studies. Results Treatment with 4-MP attenuated CCl4- and BDL-induced liver fibrosis in mice, without any adverse effects. HSCs from 4-MP treated mice depicted decreased levels of retinoic acids and increased retinol content than HSCs from control mice. In addition, the expression of α-SMA, transforming growth factor-β1 (TGF-β1), and type I collagen α1 was significantly reduced in the HSCs of 4-MP treated mice compared to the HSCs from control mice. Furthermore, inhibition of retinol metabolism by 4-MP increased interferon-γ production in NK cells, resulting in increased apoptosis of activated HSCs. Conclusions Based on our data, we conclude that inhibition of retinol metabolism by 4-MP ameliorates liver fibrosis in mice through activation of NK cells and suppression of HSCs. Therefore, retinol and its metabolizing enzyme, ADH3, might be potential targets for therapeutic intervention of liver fibrosis

  3. Th2-Associated Alternative Kupffer Cell Activation Promotes Liver Fibrosis without Inducing Local Inflammation

    PubMed Central

    López-Navarrete, Giuliana; Ramos-Martínez, Espiridión; Suárez-Álvarez, Karina; Aguirre-García, Jesús; Ledezma-Soto, Yadira; León-Cabrera, Sonia; Gudiño-Zayas, Marco; Guzmán, Carolina; Gutiérrez-Reyes, Gabriela; Hernández-Ruíz, Joselín; Camacho-Arroyo, Ignacio; Robles-Díaz, Guillermo; Kershenobich, David; Terrazas, Luis I.; Escobedo, Galileo

    2011-01-01

    Cirrhosis is the final outcome of liver fibrosis. Kupffer cell-mediated hepatic inflammation is considered to aggravate liver injury and fibrosis. Alternatively-activated macrophages are able to control chronic inflammatory events and trigger wound healing processes. Nevertheless, the role of alternative Kupffer cell activation in liver harm is largely unclear. Thus, we evaluated the participation of alternatively-activated Kupffer cells during liver inflammation and fibrosis in the murine model of carbon tetrachloride-induced hepatic damage. To stimulate alternative activation in Kupffer cells, 20 Taenia crassiceps (Tc) larvae were inoculated into BALBc/AnN female mice. Six weeks post-inoculation, carbon tetrachloride or olive oil were orally administered to Tc-inoculated and non-inoculated mice twice per week during other six weeks. The initial exposure of animals to T. crassiceps resulted in high serum concentrations of IL-4 accompanied by a significant increase in the hepatic mRNA levels of Ym-1, with no alteration in iNOS expression. In response to carbon tetrachloride, recruitment of inflammatory cell populations into the hepatic parenchyma was 5-fold higher in non-inoculated animals than Tc-inoculated mice. In contrast, carbon tetrachloride-induced liver fibrosis was significantly less in non-inoculated animals than in the Tc-inoculated group. The latter showed elevated IL-4 serum levels and low IFN-γ concentrations during the whole experiment, associated with hepatic expression of IL-4, TGF-β, desmin and α-sma, as well as increased mRNA levels of Arg-1, Ym-1, FIZZ-1 and MMR in Kupffer cells. These results suggest that alternative Kupffer cell activation is favored in a Th2 microenvironment, whereby such liver resident macrophages could exhibit a dichotomic role during chronic hepatic damage, being involved in attenuation of the inflammatory response but at the same time exacerbation of liver fibrosis. PMID:22110380

  4. Deletion of ADORA2B from myeloid cells dampens lung fibrosis and pulmonary hypertension

    PubMed Central

    Karmouty-Quintana, Harry; Philip, Kemly; Acero, Luis F.; Chen, Ning-Yuan; Weng, Tingting; Molina, Jose G.; Luo, Fayong; Davies, Jonathan; Le, Ngoc-Bao; Bunge, Isabelle; Volcik, Kelly A.; Le, Thanh-Thuy T.; Johnston, Richard A.; Xia, Yang; Eltzschig, Holger K.; Blackburn, Michael R.

    2015-01-01

    Idiopathic pulmonary fibrosis (IPF) is a lethal, fibroproliferative disease. Pulmonary hypertension (PH) can develop secondary to IPF and increase mortality. Alternatively, activated macrophages (AAMs) contribute to the pathogenesis of both IPF and PH. Here we hypothesized that adenosine signaling through the ADORA2B on AAMs impacts the progression of these disorders and that conditional deletion of ADORA2B on myeloid cells would have a beneficial effect in a model of these diseases. Conditional knockout mice lacking ADORA2B on myeloid cells (Adora2Bf/f-LysMCre) were exposed to the fibrotic agent bleomycin (BLM; 0.035 U/g body weight, i.p.). At 14, 17, 21, 25, or 33 d after exposure, SpO2, bronchoalveolar lavage fluid (BALF), and histologic analyses were performed. On day 33, lung function and cardiovascular analyses were determined. Markers for AAM and mediators of fibrosis and PH were assessed. Adora2Bf/f-LysMCre mice presented with attenuated fibrosis, improved lung function, and no evidence of PH compared with control mice exposed to BLM. These findings were accompanied by reduced expression of CD206 and arginase-1, markers for AAMs. A 10-fold reduction in IL-6 and a 5-fold decrease in hyaluronan, both linked to lung fibrosis and PH, were also observed. These data suggest that activation of the ADORA2B on macrophages plays an active role in the pathogenesis of lung fibrosis and PH.—Karmouty-Quintana, H., Philip, K., Acero, L. F., Chen, N.-Y., Weng, T., Molina, J. G., Luo, F., Davies, J., Le, N.-B., Bunge, I., Volcik, K. A., Le, T.-T. T., Johnston, R. A., Xia, Y., Eltzschig, H. K., Blackburn, M. R. Deletion of ADORA2B from myeloid cells dampens lung fibrosis and pulmonary hypertension. PMID:25318478

  5. Amniotic fluid stem cells inhibit the progression of bleomycin-induced pulmonary fibrosis via CCL2 modulation in bronchoalveolar lavage.

    PubMed

    Garcia, Orquidea; Carraro, Gianni; Turcatel, Gianluca; Hall, Marisa; Sedrakyan, Sargis; Roche, Tyler; Buckley, Sue; Driscoll, Barbara; Perin, Laura; Warburton, David

    2013-01-01

    The potential for amniotic fluid stem cell (AFSC) treatment to inhibit the progression of fibrotic lung injury has not been described. We have previously demonstrated that AFSC can attenuate both acute and chronic-fibrotic kidney injury through modification of the cytokine environment. Fibrotic lung injury, such as in Idiopathic Pulmonary Fibrosis (IPF), is mediated through pro-fibrotic and pro-inflammatory cytokine activity. Thus, we hypothesized that AFSC treatment might inhibit the progression of bleomycin-induced pulmonary fibrosis through cytokine modulation. In particular, we aimed to investigate the effect of AFSC treatment on the modulation of the pro-fibrotic cytokine CCL2, which is increased in human IPF patients and is correlated with poor prognoses, advanced disease states and worse fibrotic outcomes. The impacts of intravenous murine AFSC given at acute (day 0) or chronic (day 14) intervention time-points after bleomycin injury were analyzed at either day 3 or day 28 post-injury. Murine AFSC treatment at either day 0 or day 14 post-bleomycin injury significantly inhibited collagen deposition and preserved pulmonary function. CCL2 expression increased in bleomycin-injured bronchoalveolar lavage (BAL), but significantly decreased following AFSC treatment at either day 0 or at day 14. AFSC were observed to localize within fibrotic lesions in the lung, showing preferential targeting of AFSC to the area of fibrosis. We also observed that MMP-2 was transiently increased in BAL following AFSC treatment. Increased MMP-2 activity was further associated with cleavage of CCL2, rendering it a putative antagonist for CCL2/CCR2 signaling, which we surmise is a potential mechanism for CCL2 reduction in BAL following AFSC treatment. Based on this data, we concluded that AFSC have the potential to inhibit the development or progression of fibrosis in a bleomycin injury model during both acute and chronic remodeling events.

  6. A synthetic chloride channel restores chloride conductance in human cystic fibrosis epithelial cells.

    PubMed

    Shen, Bing; Li, Xiang; Wang, Fei; Yao, Xiaoqiang; Yang, Dan

    2012-01-01

    Mutations in the gene-encoding cystic fibrosis transmembrane conductance regulator (CFTR) cause defective transepithelial transport of chloride (Cl(-)) ions and fluid, thereby becoming responsible for the onset of cystic fibrosis (CF). One strategy to reduce the pathophysiology associated with CF is to increase Cl(-) transport through alternative pathways. In this paper, we demonstrate that a small synthetic molecule which forms Cl(-) channels to mediate Cl(-) transport across lipid bilayer membranes is capable of restoring Cl(-) permeability in human CF epithelial cells; as a result, it has the potential to become a lead compound for the treatment of human diseases associated with Cl(-) channel dysfunction.

  7. A Synthetic Chloride Channel Restores Chloride Conductance in Human Cystic Fibrosis Epithelial Cells

    PubMed Central

    Wang, Fei; Yao, Xiaoqiang; Yang, Dan

    2012-01-01

    Mutations in the gene-encoding cystic fibrosis transmembrane conductance regulator (CFTR) cause defective transepithelial transport of chloride (Cl−) ions and fluid, thereby becoming responsible for the onset of cystic fibrosis (CF). One strategy to reduce the pathophysiology associated with CF is to increase Cl− transport through alternative pathways. In this paper, we demonstrate that a small synthetic molecule which forms Cl− channels to mediate Cl− transport across lipid bilayer membranes is capable of restoring Cl− permeability in human CF epithelial cells; as a result, it has the potential to become a lead compound for the treatment of human diseases associated with Cl− channel dysfunction. PMID:22514656

  8. Activated alveolar epithelial cells initiate fibrosis through autocrine and paracrine secretion of connective tissue growth factor.

    PubMed

    Yang, Jibing; Velikoff, Miranda; Canalis, Ernesto; Horowitz, Jeffrey C; Kim, Kevin K

    2014-04-15

    Fibrogenesis involves a pathological accumulation of activated fibroblasts and extensive matrix remodeling. Profibrotic cytokines, such as TGF-β, stimulate fibroblasts to overexpress fibrotic matrix proteins and induce further expression of profibrotic cytokines, resulting in progressive fibrosis. Connective tissue growth factor (CTGF) is a profibrotic cytokine that is indicative of fibroblast activation. Epithelial cells are abundant in the normal lung, but their contribution to fibrogenesis remains poorly defined. Profibrotic cytokines may activate epithelial cells with protein expression and functions that overlap with the functions of active fibroblasts. We found that alveolar epithelial cells undergoing TGF-β-mediated mesenchymal transition in vitro were also capable of activating lung fibroblasts through production of CTGF. Alveolar epithelial cell expression of CTGF was dramatically reduced by inhibition of Rho signaling. CTGF reporter mice demonstrated increased CTGF promoter activity by lung epithelial cells acutely after bleomycin in vivo. Furthermore, mice with lung epithelial cell-specific deletion of CTGF had an attenuated fibrotic response to bleomycin. These studies provide direct evidence that epithelial cell activation initiates a cycle of fibrogenic effector cell activation during progressive fibrosis. Therapy targeted at epithelial cell production of CTGF offers a novel pathway for abrogating this progressive cycle and limiting tissue fibrosis.

  9. The role of club cell phenoconversion and migration in idiopathic pulmonary fibrosis

    PubMed Central

    Fukumoto, Jutaro; Soundararajan, Ramani; Leung, Joseph; Cox, Ruan; Mahendrasah, Sanjay; Muthavarapu, Neha; Herrin, Travis; Czachor, Alexander; Tan, Lee C.; Hosseinian, Nima; Patel, Priyanshi; Gone, Jayanthraj; Breitzig, Mason T.; Cho, Young; Cooke, Andrew J.; Galam, Lakshmi; Narala, Venkata Ramireddy; Pathak, Yashwant; Lockey, Richard F.; Kolliputi, Narasaiah

    2016-01-01

    Idiopathic pulmonary fibrosis (IPF) is an age-related multifactorial disease featuring non-uniform lung fibrosis. The decisive cellular events at early stages of IPF are poorly understood. While the involvement of club cells in IPF pathogenesis is unclear, their migration has been associated with lung fibrosis. In this study, we labeled club cells immunohistochemically in IPF lungs using a club cell marker Claudin-10 (Cldn10), a unique protein based on the recent report which demonstrated that the appearance of Cldn10 in developing and repairing lungs precedes other club cell markers including club cell secretory protein (CCSP). Cldn10-positive cells in IPF lungs displayed marked pleomorphism and were found in varied arrangements, suggesting their phenoconversion. These results were corroborated by immunogold labeling for Cldn10. Further, immunohistochemical double-labeling for Cldn10 and α-smooth muscle actin (α-SMA) demonstrated that aberrant α-SMA signals are frequently encountered near disorganized Cldn10-positive cells in hyperplastic bronchiolar epithelium and thickened interstitium of IPF lungs. Collectively, these data indicate that club cells actively participate in the initiation and progression of IPF through phenoconversion involving the acquisition of proliferative and migratory abilities. Thus, our new findings open the possibility for club cell-targeted therapy to become a strategic option for the treatment of IPF. PMID:27899769

  10. [Correlation of liver stiffness measured by FibroTouch and FibroScan with Ishak fibrosis score in patients with chronic hepatitis B].

    PubMed

    Chen, G F; Ping, J; Gu, H T; Zhao, Z M; Zhou, Y; Xing, F; Tao, Y Y; Mu, Y P; Liu, P; Liu, C H

    2017-02-20

    Objective: To investigate the correlation of liver stiffness measured by FibroTouch (FT) and FibroScan (FS) with Ishak fibrosis score in patients with chronic hepatitis B. Methods: A total of 313 patients with chronic hepatitis B who visited Department of Liver Cirrhosis in Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine from November 2014 to May 2016 were enrolled. All the patients underwent liver biopsy, and FT and FS were used to determine liver stiffness measurement (LSM). Serum biochemical parameters were measured, and the aspartate aminotransferase-to-platelet ratio index (APRI) in a multi-parameter model of liver fibrosis and fibrosis-4 (FIB-4) index were calculated. The consistency between the results of four noninvasive examinations and Ishak fibrosis score was compared. The t-test was used for comparison of LSM determined by FT and FS. Pearson correlation analysis was used investigate the correlation between LSM determined by FT and FS; Spearman correlation analysis was used to investigate the correlation of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels and Knodell score with LSM determined by FT and FS; the correlation between LSM determined by FT and FS and fibrosis stage was analyzed by partial correlation analysis adjusted by Knodell score for liver inflammatory activity; Spearman correlation analysis was used for APRI, FIB-4, and fibrosis stage. Based on the Ishak fibrosis score, the receiver operating characteristic (ROC) curve was used to analyze the values of four noninvasive methods in the diagnosis of liver fibrosis. Results: There was no significant difference in LSM measured by FT and FS in all patients (15.75±9.42 kPa vs 15.42±10.52 kPa, P > 0.05) and Pearson correlation analysis indicated a significant positive correlation between them (r = 0.858, P < 0.01); serum ALT and AST levels and liver inflammatory activity were correlated with LSM determined by FT and FS. There

  11. Human CD133+ Renal Progenitor Cells Induce Erythropoietin Production and Limit Fibrosis After Acute Tubular Injury

    PubMed Central

    Aggarwal, Shikhar; Grange, Cristina; Iampietro, Corinne; Camussi, Giovanni; Bussolati, Benedetta

    2016-01-01

    Persistent alterations of the renal tissue due to maladaptive repair characterize the outcome of acute kidney injury (AKI), despite a clinical recovery. Acute damage may also limit the renal production of erythropoietin, with impairment of the hemopoietic response to ischemia and possible lack of its reno-protective action. We aimed to evaluate the effect of a cell therapy using human CD133+ renal progenitor cells on maladaptive repair and fibrosis following AKI in a model of glycerol-induced rhabdomyolysis. In parallel, we evaluated the effect of CD133+ cells on erythropoietin production. Administration of CD133+ cells promoted the restoration of the renal tissue, limiting the presence of markers of injury and pro-inflammatory molecules. In addition, it promoted angiogenesis and protected against fibrosis up to day 60. No effect of dermal fibroblasts was observed. Treatment with CD133+ cells, but not with PBS or fibroblasts, limited anemia and increased erythropoietin levels both in renal tissue and in circulation. Finally, CD133+ cells contributed to the local production of erythropoietin, as observed by detection of circulating human erythropoietin. CD133+ cells appear therefore an effective source for cell repair, able to restore renal functions, including erythropoietin release, and to limit long term maldifferentiation and fibrosis. PMID:27853265

  12. Chrysin attenuates liver fibrosis and hepatic stellate cell activation through TGF-β/Smad signaling pathway.

    PubMed

    Balta, Cornel; Herman, Hildegard; Boldura, Oana Maria; Gasca, Ionela; Rosu, Marcel; Ardelean, Aurel; Hermenean, Anca

    2015-10-05

    We investigated the protective effect of chrysin on chronic liver fibrosis in mice and the potential mechanism underlying TGF-β1-mediated hepatic stellate cells (HSCs) activation on fibrogenesis. Experimental fibrosis was established by intraperitoneal injection of mice with 20% v/v, 2 ml/kg CCl4 twice a week, for 7 weeks. Mice were orally treated with 3 doses of chrysin (50, 100 and 200 mg/kg) or with vehicle as control. For the assessment of the spontaneous reversion of fibrosis, CCl4 treated animals were investigated after two weeks of recovery time. Silymarin was used as standard hepatoprotective flavonoid. Histopathological investigations showed that hepatic fibrosis grade was markedly reduced in the chrysin groups compared to the fibrotic one. Moreover, CCl4 activated HSCs induced an upregulation of smooth muscle actin (α-SMA), an increased number of TGF-β1 immunopositive cells and marked up-regulation of TGF-β1. α-SMA and TGF-β1 levels were significantly reduced in all chrysin treated groups in a dose-dependent manner, whereas the level of spontaneous reversal of fibrosis was lower compared to all flavonoid treated groups. Liver mRNA levels of Smad 2 in the 50, 100 and 200 mg/kg chrysin treated groups were significantly reduced by about 88.54%, 92.15% and 95.56% of the corresponding levels in the fibrosis mice group. The results were similar for mRNA levels of Smad 3. The protective response to silymarin was almost similar to that seen with the highest doses of chrysin. In this study, we have shown that chrysin has the efficacy to reverse CCl4-stimulated liver fibrosis by inhibition of HSCs activation and proliferation through TGF-β1/Smad pathway. These results suggest that chrysin may be useful in stopping or reversing the progression of liver fibrosis and might offer the possibility to develop a new therapeutic drug, useful in treatment of chronic liver diseases.

  13. S-Nitrosothiols increases cystic fibrosis transmembrane regulator expression and maturation in the cell surface.

    PubMed

    Zaman, Khalequz; Bennett, Deric; Fraser-Butler, Maya; Greenberg, Zivi; Getsy, Paulina; Sattar, Abdus; Smith, Laura; Corey, Deborah; Sun, Fei; Hunt, John; Lewis, Stephen J; Gaston, Benjamin

    2014-01-24

    S-nitrosothiols (SNOs) are endogenous signaling molecules with a broad spectrum of beneficial airway effects. SNOs are normally present in the airway, but levels tend to be low in cystic fibrosis (CF) patients. We and others have demonstrated that S-nitrosoglutathione (GSNO) increases the expression, maturation, and function of wild-type and mutant F508del cystic fibrosis transmembrane conductance regulator (CFTR) in human bronchial airway epithelial (HBAE) cells. We hypothesized that membrane permeable SNOs, such as S-nitrosoglutathione diethyl ester (GNODE) and S-nitroso-N-acetyl cysteine (SNOAC) may be more efficient in increasing the maturation of CFTR. HBAE cells expressing F508del CFTR were exposed to GNODE and SNOAC. The effects of these SNOs on the expression and maturation of F508del CFTR were determined by cell surface biotinylation and Western blot analysis. We also found for the first time that GNODE and SNOAC were effective at increasing CFTR maturation at the cell surface. Furthermore, we found that cells maintained at low temperature increased cell surface stability of F508del CFTR whereas the combination of low temperature and SNO treatment significantly extended the half-life of CFTR. Finally, we showed that SNO decreased the internalization rate of F508del CFTR in HBAE cells. We anticipate identifying the novel mechanisms, optimal SNOs, and lowest effective doses which could benefit cystic fibrosis patients.

  14. Bronchoalveolar lavage in pulmonary fibrosis: comparison of cells obtained with lung biopsy and clinical features

    PubMed Central

    Haslam, P L; Turton, C W G; Heard, B; Lukoszek, A; Collins, J V; Salsbury, A J; Turner-Warwick, M

    1980-01-01

    Bronchoalveolar lavage, open lung biopsy, and cell extraction from the biopsy material have been studied in 21 symptomatic patients with progressive pulmonary fibrosis (18 with cryptogenic fibrosing alveolitis, fulfilling also the criteria for “usual interstitial pneumonia” (UIP), and three with rapidly progressive disease probably related to asbestos exposure). The total and differential cell counts between the three different samples have been compared as well as the influence on them of smoking and their correlation with steroid responsiveness and later progress. There was no correlation between semiquantitative scores of cell types observed within alveolar spaces and in alveolar walls and the differential or total cell counts obtained from extraction or lung lavage samples. There was, however, some correlation between differential counts obtained from lung lavage and extractions (neutrophils p<0·02, eosinophils p<0·07, lymphocytes p<0·08) suggesting that lung lavage reflects the cellularity of the peripheral parts of the lung in patients without overt bronchial disease. Steroid responsiveness related to the percentage of lymphocytes found in extraction samples (p<0·01) and was associated with a complementary fall in the percentage of macrophages (p<0·02). There was no relationship between steroid response and the numbers of neutrophils or eosinophils in extracted samples. There was a trend towards increased numbers of lymphocytes in the lung wash in those patients responding to steroids. Those cases showing more rapid progression before starting treatment tended to have higher percentages of lymphocytes, neutrophils, or eosinophils in the lung lavage than more slowly deteriorating cases (p<0·01). Follow-up studies showed that three cases having predominant lymphocytes in the lung lavage continued to do well while nine cases with predominant neutrophils or eosinophils or both showed a less satisfactory response to steroids and often deteriorated

  15. Early infiltration of p40IL12+CCR7+CD11b+ cells is critical for fibrosis development

    PubMed Central

    Correa‐Costa, Matheus; Azevedo, Hatylas; Silva, Reinaldo Correia; Cruz, Mario Costa; Almeida, Maira Estanislau Soares; Hiyane, Meire Ioshie; Moreira‐Filho, Carlos Alberto; Santos, Marinilce Fagundes; Perez, Katia Regina; Cuccovia, Iolanda Midea; Camara, Niels Olsen Saraiva

    2016-01-01

    Abstract Introduction Macrophages are heterogeneous and thus can be correlated with distinct tissue outcomes after injury. Conflicting data have indicated that the M2‐related phenotype directly triggers fibrosis. Conversely, we hypothesize here that the inflammatory milieu provided by early infiltration of pro‐inflammatory macrophages dictates tissue scarring after injury. Methods and Results We first determined that tissue‐localized macrophages exhibit a pro‐inflammatory phenotype (p40IL12+CCR7+CD11b+) during the early phase of a chronic injury model, in contrast to a pro‐resolving phenotype (Arg1+IL10+CD206+CD11b+) at a later stage. Then, we evaluated the effects of injecting macrophages differentiated in vitro in the presence of IFNγ + LPS or IL4 + IL13 or non‐differentiated macrophages (hereafter, M0) on promoting inflammation and progression of chronic injury in macrophage‐depleted mice. In addition to enhancing the expression of pro‐inflammatory cytokines, the injection of M (IFNγ + LPS), but not M (IL4 + IL13) or M0, accentuated fibrosis while augmenting levels of anti‐inflammatory molecules, increasing collagen deposition and impairing organ function. We observed a similar profile after injection of sorted CCR7+CD11b+ cells and a more pronounced effect of M (IFNγ + LPS) cells originated from Stat6−/− mice. The injection of M (IFNγ + LPS) cells was associated with the up‐regulation of inflammation‐ and fibrosis‐related proteins (Thbs1, Mmp7, Mmp8, and Mmp13). Conclusions Our results suggest that pro‐inflammatory macrophages promote microenvironmental changes that may lead to fibrogenesis by inducing an inflammatory milieu that alters a network of extracellular‐related genes, culminating in tissue fibrosis. PMID:27621813

  16. MALDI imaging reveals NCOA7 as a potential biomarker in oral squamous cell carcinoma arising from oral submucous fibrosis

    PubMed Central

    Wang, Peiqi; Li, Xinyi; Chen, Fangman; Sun, Chongkui; Zhao, Hang; Zeng, Xin; Jiang, Lu; Zhou, Yu; Dan, Hongxia; Feng, Mingye; Liu, Rui; Chen, Qianming

    2016-01-01

    Oral squamous cell carcinoma (OSCC) ranks among the most common cancer worldwide, and is associated with severe morbidity and high mortality. Oral submucous fibrosis (OSF), characterized by fibrosis of the mucosa of the upper digestive tract, is a pre-malignant lesion, but the molecular mechanisms underlying this malignant transformation remains to be elucidated. In this study, matrix-assisted laser desorption ionization imaging mass spectrometry (MALDI-IMS)-based proteomic strategy was employed to profile the differentially expressed peptides/proteins between OSCC tissues and the corresponding adjacent non-cancerous OSF tissues. Sixty-five unique peptide peaks and nine proteins were identified with altered expression levels. Of them, expression of NCOA7 was found to be up-regulated in OSCC tissues by immunohistochemistry staining and western blotting, and correlated with a pan of clinicopathologic parameters, including lesion site, tumor differentiation status and lymph node metastasis. Further, we show that overexpression of NCOA7 promotes OSCC cell proliferation in either in vitro or in vivo models. Mechanistic study demonstrates that NCOA7 induces OSCC cell proliferation probably by activating aryl hydrocarbon receptor (AHR). The present study suggests that NCOA7 is a potential biomarker for early diagnosis of OSF malignant transformation, and leads to a better understanding of the molecular mechanisms responsible for OSCC development. PMID:27509054

  17. Restoration of Chloride Efflux by Azithromycin in Airway Epithelial Cells of Cystic Fibrosis Patients▿

    PubMed Central

    Saint-Criq, Vinciane; Rebeyrol, Carine; Ruffin, Manon; Roque, Telma; Guillot, Loïc; Jacquot, Jacky; Clement, Annick; Tabary, Olivier

    2011-01-01

    Azithromycin (AZM) has shown promising anti-inflammatory properties in chronic obstructive pulmonary diseases, and clinical studies have presented an improvement in the respiratory condition of cystic fibrosis (CF) patients. The aim of this study was to investigate, in human airway cells, the mechanism by which AZM has beneficial effects in CF. We demonstrated that AZM did not have any anti-inflammatory effect on CF airway cells but restored Cl− efflux. PMID:21220528

  18. Strongly correlated perovskite fuel cells

    NASA Astrophysics Data System (ADS)

    Zhou, You; Guan, Xiaofei; Zhou, Hua; Ramadoss, Koushik; Adam, Suhare; Liu, Huajun; Lee, Sungsik; Shi, Jian; Tsuchiya, Masaru; Fong, Dillon D.; Ramanathan, Shriram

    2016-06-01

    Fuel cells convert chemical energy directly into electrical energy with high efficiencies and environmental benefits, as compared with traditional heat engines. Yttria-stabilized zirconia is perhaps the material with the most potential as an electrolyte in solid oxide fuel cells (SOFCs), owing to its stability and near-unity ionic transference number. Although there exist materials with superior ionic conductivity, they are often limited by their ability to suppress electronic leakage when exposed to the reducing environment at the fuel interface. Such electronic leakage reduces fuel cell power output and the associated chemo-mechanical stresses can also lead to catastrophic fracture of electrolyte membranes. Here we depart from traditional electrolyte design that relies on cation substitution to sustain ionic conduction. Instead, we use a perovskite nickelate as an electrolyte with high initial ionic and electronic conductivity. Since many such oxides are also correlated electron systems, we can suppress the electronic conduction through a filling-controlled Mott transition induced by spontaneous hydrogen incorporation. Using such a nickelate as the electrolyte in free-standing membrane geometry, we demonstrate a low-temperature micro-fabricated SOFC with high performance. The ionic conductivity of the nickelate perovskite is comparable to the best-performing solid electrolytes in the same temperature range, with a very low activation energy. The results present a design strategy for high-performance materials exhibiting emergent properties arising from strong electron correlations.

  19. Strongly correlated perovskite fuel cells

    SciTech Connect

    Zhou, You; Guan, Xiaofei; Zhou, Hua; Ramadoss, Koushik; Adam, Suhare; Liu, Huajun; Lee, Sungsik; Shi, Jian; Tsuchiya, Masaru; Fong, Dillon D.; Ramanathan, Shriram

    2016-05-16

    Fuel cells convert chemical energy directly into electrical energy with high efficiencies and environmental benefits, as compared with traditional heat engines1, 2, 3, 4. Yttria-stabilized zirconia is perhaps the material with the most potential as an electrolyte in solid oxide fuel cells (SOFCs), owing to its stability and near-unity ionic transference number5. Although there exist materials with superior ionic conductivity, they are often limited by their ability to suppress electronic leakage when exposed to the reducing environment at the fuel interface. Such electronic leakage reduces fuel cell power output and the associated chemo-mechanical stresses can also lead to catastrophic fracture of electrolyte membranes6. Here we depart from traditional electrolyte design that relies on cation substitution to sustain ionic conduction. Instead, we use a perovskite nickelate as an electrolyte with high initial ionic and electronic conductivity. Since many such oxides are also correlated electron systems, we can suppress the electronic conduction through a filling-controlled Mott transition induced by spontaneous hydrogen incorporation. Using such a nickelate as the electrolyte in free-standing membrane geometry, we demonstrate a low-temperature micro-fabricated SOFC with high performance. The ionic conductivity of the nickelate perovskite is comparable to the best-performing solid electrolytes in the same temperature range, with a very low activation energy. The results present a design strategy for high-performance materials exhibiting emergent properties arising from strong electron correlations.

  20. Strongly correlated perovskite fuel cells.

    PubMed

    Zhou, You; Guan, Xiaofei; Zhou, Hua; Ramadoss, Koushik; Adam, Suhare; Liu, Huajun; Lee, Sungsik; Shi, Jian; Tsuchiya, Masaru; Fong, Dillon D; Ramanathan, Shriram

    2016-06-09

    Fuel cells convert chemical energy directly into electrical energy with high efficiencies and environmental benefits, as compared with traditional heat engines. Yttria-stabilized zirconia is perhaps the material with the most potential as an electrolyte in solid oxide fuel cells (SOFCs), owing to its stability and near-unity ionic transference number. Although there exist materials with superior ionic conductivity, they are often limited by their ability to suppress electronic leakage when exposed to the reducing environment at the fuel interface. Such electronic leakage reduces fuel cell power output and the associated chemo-mechanical stresses can also lead to catastrophic fracture of electrolyte membranes. Here we depart from traditional electrolyte design that relies on cation substitution to sustain ionic conduction. Instead, we use a perovskite nickelate as an electrolyte with high initial ionic and electronic conductivity. Since many such oxides are also correlated electron systems, we can suppress the electronic conduction through a filling-controlled Mott transition induced by spontaneous hydrogen incorporation. Using such a nickelate as the electrolyte in free-standing membrane geometry, we demonstrate a low-temperature micro-fabricated SOFC with high performance. The ionic conductivity of the nickelate perovskite is comparable to the best-performing solid electrolytes in the same temperature range, with a very low activation energy. The results present a design strategy for high-performance materials exhibiting emergent properties arising from strong electron correlations.

  1. Effects of Melatonin on Differentiation Potential of Ito Cells in Mice with Induced Fibrosis of the Liver.

    PubMed

    Nalobin, D S; Suprunenko, E A; Golichenkov, V A

    2016-10-01

    We studied the effects of melatonin on differentiation potential of Ito cells during atypical regeneration of mouse liver under conditions of CCl4-induced fibrosis. The dynamics of fibrosis was traced at the histological level and the effects of melatonin on the differentiation potential of mouse Ito cells were evaluated. Melatonin alleviated fibrotic changes in the liver tissue and reduced differentiation of Ito cells into myofibroblasts under conditions of atypical regeneration of the liver in induced fibrosis. The hepatoprotective role of melatonin was shown.

  2. Hepatitis C virus load in parenchyma cells correlates with hepatic injury in infected patients

    PubMed Central

    Xu, Zhen; Lin, Ji-Zong; Lin, Guo-Li; Wei, Fang-Fang; Liu, Jing; Zhao, Zhi-Xin; Zhang, Ying; Ke, Wei-Ming; Zhang, Xiao-Hong

    2017-01-01

    The association between serum hepatitis C virus (HCV) load and hepatic injury in HCV-infected patients has been extensively investigated. The present study aimed to investigate the association between HCV load in hepatic parenchyma cells and hepatic injury in HCV-infected patients. A total of 56 HCV-infected patients were included in the present retrospective study. The serum HCV mRNA was determined using quantitative polymerase chain reaction, while the hepatic parenchyma cell volume and HCV mRNA in hepatic parenchyma cells were also determined. Hepatic injury was evaluated on the basis of the severity of inflammation and fibrosis. The results demonstrated that there were evident differences in the mean serum HCV RNA levels and the HCV load/parenchyma cell volume among the various grades of hepatic inflammation (G1-G4) when groups with the least and most inflammation were compared (G1 vs. G4; P<0.05). Significant differences in the HCV load existed between groups divided according to the fibrosis grade; in addition, differences existed between fibrosis grades S1 and S2, and S2 and S4 when comparing serum HCV RNA levels (P<0.05). Similarly, differences existed between every two fibrosis stages (S0 vs. S4, S2 vs. S3, and S2 vs. S4; P<0.05) when viral loads and parenchyma cell volumes were compared (F=2.860, P<0.05). Furthermore, the fibrosis staging was correlated with the viral load/parenchyma cell volume (F=2.670, P<0.05). In conclusion, hepatic fibrosis grade was found to be associated with HCV load in parenchyma cells. The results of the present study demonstrated that the viral load in parenchyma cells is a more appropriate index compared with the serum viral load for evaluating HCV replication in hepatocytes, and may function as an important factor in HCV-infected hepatic injury evaluation. PMID:28123484

  3. Hepatitis C virus load in parenchyma cells correlates with hepatic injury in infected patients.

    PubMed

    Xu, Zhen; Lin, Ji-Zong; Lin, Guo-Li; Wei, Fang-Fang; Liu, Jing; Zhao, Zhi-Xin; Zhang, Ying; Ke, Wei-Ming; Zhang, Xiao-Hong

    2017-01-01

    The association between serum hepatitis C virus (HCV) load and hepatic injury in HCV-infected patients has been extensively investigated. The present study aimed to investigate the association between HCV load in hepatic parenchyma cells and hepatic injury in HCV-infected patients. A total of 56 HCV-infected patients were included in the present retrospective study. The serum HCV mRNA was determined using quantitative polymerase chain reaction, while the hepatic parenchyma cell volume and HCV mRNA in hepatic parenchyma cells were also determined. Hepatic injury was evaluated on the basis of the severity of inflammation and fibrosis. The results demonstrated that there were evident differences in the mean serum HCV RNA levels and the HCV load/parenchyma cell volume among the various grades of hepatic inflammation (G1-G4) when groups with the least and most inflammation were compared (G1 vs. G4; P<0.05). Significant differences in the HCV load existed between groups divided according to the fibrosis grade; in addition, differences existed between fibrosis grades S1 and S2, and S2 and S4 when comparing serum HCV RNA levels (P<0.05). Similarly, differences existed between every two fibrosis stages (S0 vs. S4, S2 vs. S3, and S2 vs. S4; P<0.05) when viral loads and parenchyma cell volumes were compared (F=2.860, P<0.05). Furthermore, the fibrosis staging was correlated with the viral load/parenchyma cell volume (F=2.670, P<0.05). In conclusion, hepatic fibrosis grade was found to be associated with HCV load in parenchyma cells. The results of the present study demonstrated that the viral load in parenchyma cells is a more appropriate index compared with the serum viral load for evaluating HCV replication in hepatocytes, and may function as an important factor in HCV-infected hepatic injury evaluation.

  4. HIV-HCV co-infected patients with low CD4+ cell nadirs are at risk for faster fibrosis progression and portal hypertension.

    PubMed

    Reiberger, T; Ferlitsch, A; Sieghart, W; Kreil, A; Breitenecker, F; Rieger, A; Schmied, B; Gangl, A; Peck-Radosavljevic, M

    2010-06-01

    Patients co-infected with the human immunodeficiency virus (HIV) and the hepatitis C virus (HCV) are fraught with a rapid fibrosis progression rate and with complications of portal hypertension (PHT) We aimed to assess the influence of immune function [Centers of Disease Control and Prevention (CDC) stage] on development of PHT and disease progression in HIV-HCV co-infection. Data of 74 interferon-naïve HIV-HCV co-infected patients undergoing liver biopsy, measurement of portal pressure and of liver stiffness and routine laboratory tests (including CD4+ cell count, HIV and HCV viral load) were analysed. Time of initial exposure (risk behaviour) was used to assess fibrosis progression. Fibrosis progression, time to cirrhosis and portal pressure were correlated with HIV status (CDC stage). HIV-HCV patients had rapid progression of fibrosis [0.201 +/- 0.088 METAVIR fibrosis units/year (FU/y)] and accelerated time to cirrhosis (24 +/- 13 years), high HCV viral loads (4.83 x 10(6) IU/mL) and a mean HVPG at the upper limit of normal (5 mmHg). With moderate or severe immunodeficiency, fibrosis progression was even higher (CDC-2 = 0.177 FU/y; CDC-3 = 0.248 FU/y) compared with patients with higher CD4+ nadirs (CDC-1 = 0.120 FU/y; P = 0.0001). An indirect correlation between CD4+ cell count and rate of fibrosis progression (R = -0.6654; P < 0.001) could be demonstrated. Hepatic venous pressure gradient (HVPG) showed early elevation of portal pressure with median values of 4, 8 and 12 mmHg after 10, 15 and 20 years of HCV infection for CDC-3 patients. Patients treated with highly active anti-retroviral therapy (HAART) had similar rates of progression and portal pressure values than patients without HAART. Progression of HCV disease is accelerated in HIV-HCV co-infection, being more pronounced in patients with low CD4+ cell count. A history of a CD4+ cell nadir <200/microL is a risk factor for rapid development of cirrhosis and PHT. Thus, HCV treatment should be considered

  5. The Role of Dendritic Cells in Fibrosis Progression in Nonalcoholic Fatty Liver Disease

    PubMed Central

    Almeda-Valdes, Paloma; Aguilar Olivos, Nancy E.; Barranco-Fragoso, Beatriz; Uribe, Misael; Méndez-Sánchez, Nahum

    2015-01-01

    Nonalcoholic fatty liver disease (NAFLD) is the most frequent cause of chronic liver disease. NAFLD encompasses a wide range of pathologies, from simple steatosis to steatosis with inflammation to fibrosis. The pathogenesis of NAFLD progression has not been completely elucidated, and different liver cells could be implicated. This review focuses on the current evidence of the role of liver dendritic cells (DCs) in the progression from NAFLD to fibrosis. Liver DCs are a heterogeneous population of hepatic antigen-presenting cells; their main function is to induce T-cell mediated immunity by antigen processing and presentation to T cells. During the steady state liver DCs are immature and tolerogenic. However, in an environment of chronic inflammation, DCs are transformed to potent inducers of immune responses. There is evidence about the role of DC in liver fibrosis, but it is not clearly understood. Interestingly, there might be a link between lipid metabolism and DC function, suggesting that immunogenic DCs are associated with liver lipid storage, representing a possible pathophysiological mechanism in NAFLD development. A better understanding of the interaction between inflammatory pathways and the different cell types and the effect on the progression of NAFLD is of great relevance. PMID:26339640

  6. Functional Gene Correction for Cystic Fibrosis in Lung Epithelial Cells Generated From Patient iPSCs

    PubMed Central

    Firth, Amy L; Menon, Tushar; Parker, Gregory S; Qualls, Susan J; Lewis, Benjamin M; Ke, Eugene; Dargitz, Carl T; Wright, Rebecca; Khanna, Ajai; Gage, Fred H; Verma, Inder M

    2015-01-01

    SUMMARY Lung disease is a major cause of death in the USA, with current therapeutic approaches only serving to manage symptoms. The most common chronic and life-threatening genetic disease of the lung is Cystic fibrosis (CF) caused by mutations in the cystic fibrosis transmembrane regulator (CFTR). We have generated induced pluripotent stem cells (iPSC) from CF patients carrying a homozygous deletion of F508 in the CFTR gene, which results in defective processing of CFTR to the cell membrane. This mutation was precisely corrected using CRISPR to target corrective sequences to the endogenous CFTR genomic locus, in combination with a completely excisable selection system which significantly improved the efficiency of this correction. The corrected iPSC were subsequently differentiated to mature airway epithelial cells where recovery of normal CFTR expression and function was demonstrated. This isogenic iPSC-based model system for CF could be adapted for the development of new therapeutic approaches. PMID:26299960

  7. IGF-I induces senescence of hepatic stellate cells and limits fibrosis in a p53-dependent manner

    PubMed Central

    Nishizawa, Hitoshi; Iguchi, Genzo; Fukuoka, Hidenori; Takahashi, Michiko; Suda, Kentaro; Bando, Hironori; Matsumoto, Ryusaku; Yoshida, Kenichi; Odake, Yukiko; Ogawa, Wataru; Takahashi, Yutaka

    2016-01-01

    Hepatic fibrosis in nonalcoholic steatohepatitis (NASH) and cirrhosis determines patient prognosis; however, effective treatment for fibrosis has not been established. Oxidative stress and inflammation activate hepatic stellate cells (HSCs) and promote fibrosis. In contrast, cellular senescence inhibits HSCs’ activity and limits fibrosis. The aim of this study was to explore the effect of IGF-I on NASH and cirrhotic models and to clarify the underlying mechanisms. We demonstrate that IGF-I significantly ameliorated steatosis, inflammation, and fibrosis in a NASH model, methionine-choline-deficient diet-fed db/db mice and ameliorated fibrosis in cirrhotic model, dimethylnitrosamine-treated mice. As the underlying mechanisms, IGF-I improved oxidative stress and mitochondrial function in the liver. In addition, IGF-I receptor was strongly expressed in HSCs and IGF-I induced cellular senescence in HSCs in vitro and in vivo. Furthermore, in mice lacking the key senescence regulator p53, IGF-I did not induce cellular senescence in HSCs or show any effects on fibrosis. Taken together, these results indicate that IGF-I induces senescence of HSCs, inactivates these cells and limits fibrosis in a p53-dependent manner and that IGF-I may be applied to treat NASH and cirrhosis. PMID:27721459

  8. Three distinct cell populations express extracellular matrix proteins and increase in number during skeletal muscle fibrosis.

    PubMed

    Chapman, Mark A; Mukund, Kavitha; Subramaniam, Shankar; Brenner, David; Lieber, Richard L

    2017-02-01

    Tissue extracellular matrix (ECM) provides structural support and creates unique environments for resident cells (Bateman JF, Boot-Handford RP, Lamandé SR. Nat Rev Genet 10: 173-183, 2009; Kjaer M. Physiol Rev 84: 649-98, 2004). However, the identities of cells responsible for creating specific ECM components have not been determined. In striated muscle, the identity of these cells becomes important in disease when ECM changes result in fibrosis and subsequent increased tissue stiffness and dysfunction. Here we describe a novel approach to isolate and identify cells that maintain the ECM in both healthy and fibrotic muscle. Using a collagen I reporter mouse, we show that there are three distinct cell populations that express collagen I in both healthy and fibrotic skeletal muscle. Interestingly, the number of collagen I-expressing cells in all three cell populations increases proportionally in fibrotic muscle, indicating that all cell types participate in the fibrosis process. Furthermore, while some profibrotic ECM and ECM-associated genes are significantly upregulated in fibrotic muscle, the fibrillar collagen gene expression profile is not qualitatively altered. This suggests that muscle fibrosis in this model results from an increased number of collagen I-expressing cells and not the initiation of a specific fibrotic collagen gene expression program. Finally, in fibrotic muscle, we show that these collagen I-expressing cell populations differentially express distinct ECM proteins-fibroblasts express the fibrillar components of ECM, fibro/adipogenic progenitors cells differentially express basal laminar proteins, and skeletal muscle progenitor cells differentially express genes important for the satellite cell.

  9. Reversibility of liver fibrosis.

    PubMed

    Sun, Mengxi; Kisseleva, Tatiana

    2015-09-01

    Liver fibrosis is a serious health problem worldwide, which can be induced by a wide spectrum of chronic liver injuries. However, until today, there is no effective therapy available for liver fibrosis except the removal of underlying etiology or liver transplantation. Recent studies indicate that liver fibrosis is reversible when the causative agent(s) is removed. Understanding of mechanisms of liver fibrosis regression will lead to the identification of new therapeutic targets for liver fibrosis. This review summarizes recent research progress on mechanisms of reversibility of liver fibrosis. While most of the research has been focused on HSCs/myofibroblasts and inflammatory pathways, the crosstalk between different organs, various cell types and multiple signaling pathways should not be overlooked. Future studies that lead to fully understanding of the crosstalk between different cell types and the molecular mechanism underlying the reversibility of liver fibrosis will definitely give rise to new therapeutic strategies to treat liver fibrosis.

  10. Modulation of the Unfolded Protein Response by Tauroursodeoxycholic Acid Counteracts Apoptotic Cell Death and Fibrosis in a Mouse Model for Secondary Biliary Liver Fibrosis

    PubMed Central

    Paridaens, Annelies; Raevens, Sarah; Devisscher, Lindsey; Bogaerts, Eliene; Verhelst, Xavier; Hoorens, Anne; van Vlierberghe, Hans; Van Grunsven, Leo A.; Geerts, Anja; Colle, Isabelle

    2017-01-01

    The role of endoplasmic reticulum stress and the unfolded protein response (UPR) in cholestatic liver disease and fibrosis is not fully unraveled. Tauroursodeoxycholic acid (TUDCA), a hydrophilic bile acid, has been shown to reduce endoplasmic reticulum (ER) stress and counteract apoptosis in different pathologies. We aimed to investigate the therapeutic potential of TUDCA in experimental secondary biliary liver fibrosis in mice, induced by common bile duct ligation. The kinetics of the hepatic UPR and apoptosis during the development of biliary fibrosis was studied by measuring markers at six different timepoints post-surgery by qPCR and Western blot. Next, we investigated the therapeutic potential of TUDCA, 10 mg/kg/day in drinking water, on liver damage (AST/ALT levels) and fibrosis (Sirius red-staining), in both a preventive and therapeutic setting. Common bile duct ligation resulted in the increased protein expression of CCAAT/enhancer-binding protein homologous protein (CHOP) at all timepoints, along with upregulation of pro-apoptotic caspase 3 and 12, tumor necrosis factor receptor superfamily, member 1A (TNFRsf1a) and Fas-Associated protein with Death Domain (FADD) expression. Treatment with TUDCA led to a significant reduction of liver fibrosis, accompanied by a slight reduction of liver damage, decreased hepatic protein expression of CHOP and reduced gene and protein expression of pro-apoptotic markers. These data indicate that TUDCA exerts a beneficial effect on liver fibrosis in a model of cholestatic liver disease, and suggest that this effect might, at least in part, be attributed to decreased hepatic UPR signaling and apoptotic cell death. PMID:28117681

  11. Altered surfactant homeostasis and alveolar epithelial cell stress in amiodarone-induced lung fibrosis.

    PubMed

    Mahavadi, Poornima; Henneke, Ingrid; Ruppert, Clemens; Knudsen, Lars; Venkatesan, Shalini; Liebisch, Gerhard; Chambers, Rachel C; Ochs, Matthias; Schmitz, Gerd; Vancheri, Carlo; Seeger, Werner; Korfei, Martina; Guenther, Andreas

    2014-11-01

    Amiodarone (AD) is a highly efficient antiarrhythmic drug with potentially serious side effects. Severe pulmonary toxicity is reported in patients receiving AD even at low doses and may cause interstitial pneumonia as well as lung fibrosis. Apoptosis of alveolar epithelial type II cells (AECII) has been suggested to play an important role in this disease. In the current study, we aimed to establish a murine model of AD-induced lung fibrosis and analyze surfactant homeostasis, lysosomal, and endoplasmic reticulum (ER) stress in this model. AD/vehicle was instilled intratracheally into C57BL/6 mice, which were sacrificed on days 7, 14, 21, and 28. Extent of lung fibrosis development was assessed by trichrome staining and hydroxyproline measurement. Cytotoxicity was assessed by lactate dehydrogenase assay. Phospholipids (PLs) were analyzed by mass spectrometry. Surfactant proteins (SP) and markers for apoptosis, lysosomal, and ER stress were studied by Western blotting and immunohistochemistry. AECII morphology was evaluated by electron microscopy. Extensive lung fibrosis and AECII hyperplasia were observed in AD-treated mice already at day 7. Surfactant PL and SP accumulated in AECII over time. In parallel, induction of apoptosis, lysosomal, and ER stress was encountered in AECII of mice lungs and in MLE12 cells treated with AD. In vitro, siRNA-mediated knockdown of cathepsin D did not alter the AD-induced apoptotic response. Our data suggest that mice exposed to intratracheal AD develop severe pulmonary fibrosis, exhibit extensive surfactant alterations and cellular stress, but AD-induced AECII apoptosis is not mediated primarily via cathepsin D.

  12. Corona-directed nucleic acid delivery into hepatic stellate cells for liver fibrosis therapy.

    PubMed

    Zhang, Zhengping; Wang, Chunming; Zha, Yinhe; Hu, Wei; Gao, Zhongfei; Zang, Yuhui; Chen, Jiangning; Zhang, Junfeng; Dong, Lei

    2015-03-24

    Strategies to modify nanoparticles with biological ligands for targeted drug delivery in vivo have been widely studied but met with limited clinical success. A possible reason is that, in the blood circulation, serum proteins could rapidly form a layer of protein "corona" on the vehicle surface, which might block the modified ligands and hamper their targeting functions. We speculate that strategies for drug delivery can be designed based upon elegant control of the corona formation on the vehicle surfaces. In this study, we demonstrate a retinol-conjugated polyetherimine (RcP) nanoparticle system that selectively recruited the retinol binding protein 4 (RBP) in its corona components. RBP was found to bind retinol, and direct the antisense oligonucleotide (ASO)-laden RcP carrier to hepatic stellate cells (HSC), which play essential roles in the progression of hepatic fibrosis. In both mouse fibrosis models, induced by carbon tetrachloride (CCl4) and bile duct ligation (BDL), respectively, the ASO-laden RcP particles effectively suppressed the expression of type I collagen (collagen I), and consequently ameliorated hepatic fibrosis. Such findings suggest that this delivery system, designed to exploit the power of corona proteins, can serve as a promising tool for targeted delivery of therapeutic agents for the treatment of hepatic fibrosis.

  13. Deletion of mTORC1 Activity in CD4+ T Cells Is Associated with Lung Fibrosis and Increased γδ T Cells

    PubMed Central

    Vigeland, Christine L.; Collins, Samuel L.; Chan-Li, Yee; Hughes, Andrew H.; Oh, Min-Hee; Powell, Jonathan D.; Horton, Maureen R.

    2016-01-01

    Pulmonary fibrosis is a devastating, incurable disease in which chronic inflammation and dysregulated, excessive wound healing lead to progressive fibrosis, lung dysfunction, and ultimately death. Prior studies have implicated the cytokine IL-17A and Th17 cells in promoting the development of fibrosis. We hypothesized that loss of Th17 cells via CD4-specific deletion of mTORC1 activity would abrogate the development of bleomycin-induced pulmonary fibrosis. However, in actuality loss of Th17 cells led to increased mortality and fibrosis in response to bleomycin. We found that in the absence of Th17 cells, there was continued production of IL-17A by γδ T cells. These IL-17A+ γδ T cells were associated with increased lung neutrophils and M2 macrophages, accelerated development of fibrosis, and increased mortality. These data elucidate the critical role of IL-17A+ γδ T cells in promoting chronic inflammation and fibrosis, and reveal a novel therapeutic target for treatment of pulmonary fibrosis. PMID:27649073

  14. Rare aggressive natural killer cell leukemia presented with bone marrow fibrosis - a diagnostic challenge.

    PubMed

    Soliman, Dina S; Sabbagh, Ahmad Al; Omri, Halima El; Ibrahim, Firyal A; Amer, Aliaa M; Otazu, Ivone B

    2014-01-01

    Aggressive natural killer cell leukemia is an extraordinary rare aggressive malignant neoplasm of natural killer cells. Although its first recognition as a specific entity was approximately 20 years ago, this leukemia has not yet been satisfactorily characterized as fewer than 200 cases have been reported in the literature and up to our knowledge, this is the first case report in Qatar. Reaching a diagnosis of aggressive natural killer leukemia was a challenging experience, because in addition to being a rare entity, the relative scarcity of circulating neoplastic cells, failure to obtain an adequate aspirate sample sufficient to perform flow cytometric analysis, together with the absence of applicable method to prove NK clonality (as it lack specific clonal marker); our case had atypical confusing presentation of striking increase in bone marrow fibrosis that was misleading and complicated the case further. The bone marrow fibrosis encountered may be related to the neoplastic natural killer cells' chemokine profile and it may raise the awareness for considering aggressive natural killer leukemia within the differential diagnosis of leukemia with heightened marrow fibrosis.

  15. Gradually softening hydrogels for modeling hepatic stellate cell behavior during fibrosis regression.

    PubMed

    Caliari, Steven R; Perepelyuk, Maryna; Soulas, Elizabeth M; Lee, Gi Yun; Wells, Rebecca G; Burdick, Jason A

    2016-06-13

    The extracellular matrix (ECM) presents an evolving set of mechanical cues to resident cells. We developed methacrylated hyaluronic acid (MeHA) hydrogels containing both stable and hydrolytically degradable crosslinks to provide cells with a gradually softening (but not fully degradable) milieu, mimicking physiological events such as fibrosis regression. To demonstrate the utility of this cell culture system, we studied the phenotype of rat hepatic stellate cells, the major liver precursors of fibrogenic myofibroblasts, within this softening environment. Stellate cells that were mechanically primed on tissue culture plastic attained a myofibroblast phenotype, which persisted when seeded onto stiff (∼20 kPa) hydrogels. However, mechanically primed stellate cells on stiff-to-soft (∼20 to ∼3 kPa) hydrogels showed reversion of the myofibroblast phenotype over 14 days, with reductions in cell area, expression of the myofibroblast marker alpha-smooth muscle actin (α-SMA), and Yes-associated protein/Transcriptional coactivator with PDZ-binding motif (YAP/TAZ) nuclear localization when compared to stellate cells on stiff hydrogels. Cells on stiff-to-soft hydrogels did not fully revert, however. They displayed reduced expression of glial fibrillary acidic protein (GFAP), and underwent abnormally rapid re-activation to myofibroblasts in response to re-stiffening of the hydrogels through introduction of additional crosslinks. These features are typical of stellate cells with an intermediate phenotype, reported to occur in vivo with fibrosis regression and re-injury. Together, these data suggest that mechanics play an important role in fibrosis regression and that integrating dynamic mechanical cues into model systems helps capture cell behaviors observed in vivo.

  16. Continuous AMD3100 Treatment Worsens Renal Fibrosis through Regulation of Bone Marrow Derived Pro-Angiogenic Cells Homing and T-Cell-Related Inflammation.

    PubMed

    Yang, Juan; Zhu, Fengming; Wang, Xiaohui; Yao, Weiqi; Wang, Meng; Pei, Guangchang; Hu, Zhizhi; Guo, Yujiao; Zhao, Zhi; Wang, Pengge; Mou, Jingyi; Sun, Jie; Zeng, Rui; Xu, Gang; Liao, Wenhui; Yao, Ying

    2016-01-01

    AMD3100 is a small molecule inhibitor of chemokine receptor type 4 (CXCR4), which is located in the cell membranes of CD34+ cells and a variety of inflammatory cells and has been reported to reduce organ fibrosis in the lung, liver and myocardium. However, the effect of AMD3100 on renal fibrosis is unknown. This study investigated the impact of AMD3100 on renal fibrosis. C57bl/6 mice were subjected to unilateral ureteral obstruction (UUO) surgery with or without AMD3100 administration. Tubular injury, collagen deposition and fibrosis were detected and analyzed by histological staining, immunocytochemistry and Western Blot. Bone marrow derived pro-angiogenic cells (CD45+, CD34+ and CD309+ cells) and capillary density (CD31+) were measured by flow cytometry (FACS) and immunofluorescence (IF). Inflammatory cells, chemotactic factors and T cell proliferation were characterized. We found that AMD3100 treatment did not alleviate renal fibrosis but, rather, increased tissue damage and renal fibrosis. Continuous AMD3100 administration did not improve bone marrow derived pro-angiogenic cells mobilization but, rather, inhibited the migration of bone marrow derived pro-angiogenic cells into the fibrotic kidney. Additionally, T cell infiltration was significantly increased in AMD3100-treated kidneys compared to un-treated kidneys. Thus, treatment of UUO mice with AMD3100 led to an increase in T cell infiltration, suggesting that AMD3100 aggravated renal fibrosis.

  17. Dynamic of bone marrow fibrosis regression predicts survival after allogeneic stem cell transplantation for myelofibrosis.

    PubMed

    Kröger, Nicolaus; Zabelina, Tatjana; Alchalby, Haefaa; Stübig, Thomas; Wolschke, Christine; Ayuk, Francis; von Hünerbein, Natascha; Kvasnicka, Hans-Michael; Thiele, Jürgen; Kreipe, Hans-Heinrich; Büsche, Guntram

    2014-06-01

    We correlate regression of bone marrow fibrosis (BMF) on day 30 and 100 after dose- reduced allogeneic stem cell transplantation (allo-SCT) in 57 patients with primary or post-essential thrombocythemia/polycythemia vera myelofibrosis with graft function and survival. The distribution of International Prognostic Scoring System (IPSS) risk score categories was 1 patient with low risk, 5 patients with intermediate-1 risk, 18 patients with intermediate-2 risk, and 33 patients with high risk. Before allo-SCT, 41 patients (72%) were classified as XXX [myclofibrosis (MF)]-3 and 16 (28%) were classified as MF-2 according to the World Health Organization criteria. At postengraftment day +30 (±10 days), 21% of the patients had near-complete or complete regression of BMF (MF-0/-1), and on day +100 (±20 days), 54% were MF-0/-1. The 5-year overall survival rate at day +100 was 96% in patients with MF-0/-1 and 57% for those with MF-2/-3 (P = .04). There was no difference in BMF regression at day +100 between IPSS high-risk and low/intermediate-risk patients. Complete donor cell chimerism at day +100 was seen in 81% of patients with MF-0/-1 and in 31% of those with MF-2/-3. Patients with MF-2/-3 at day +100 were more likely to be transfusion-dependent for either RBCs (P = .014) or platelets (P = .018). Rapid BMF regression after reduced-intensity conditioning allo-SCT resulted in a favorable survival independent of IPSS risk score at transplantation.

  18. A bacterial cell to cell signal in the lungs of cystic fibrosis patients.

    PubMed

    Collier, David N; Anderson, Lisa; McKnight, Susan L; Noah, Terry L; Knowles, Michael; Boucher, Richard; Schwab, Ute; Gilligan, Peter; Pesci, Everett C

    2002-09-24

    Pseudomonas aeruginosa is an opportunistic pathogen that is a major cause of mortality in cystic fibrosis (CF) patients. This bacterium has numerous genes controlled by cell to cell signaling, which occurs through a complex circuitry of interconnected regulatory systems. One of the signals is the Pseudomonas Quinolone Signal (PQS), which was identified as 2-heptyl-3-hydroxy-4-quinolone. This intercellular signal controls the expression of multiple virulence factors and is required for virulence in an insect model of P. aeruginosa infection. Previous studies have implied that the intercellular signals of P. aeruginosa are important for human disease, and our goal was to determine whether PQS was produced during human infections. In this report, three types of samples from CF patients infected with P. aeruginosa were analyzed for the presence of PQS. Sputum, bronchoalveolar lavage fluid, and mucopurulent fluid from distal airways of end-stage lungs removed at transplant, all contained PQS, indicating that this cell to cell signal is produced in vivo by P. aeruginosa infecting the lungs of CF patients.

  19. Combination therapy of mesenchymal stem cells and serelaxin effectively attenuates renal fibrosis in obstructive nephropathy.

    PubMed

    Huuskes, Brooke M; Wise, Andrea F; Cox, Alison J; Lim, Ee X; Payne, Natalie L; Kelly, Darren J; Samuel, Chrishan S; Ricardo, Sharon D

    2015-02-01

    Chronic kidney disease (CKD) results from the development of fibrosis, ultimately leading to end-stage renal disease (ESRD). Although human bone marrow-derived mesenchymal stem cells (MSCs) can accelerate renal repair following acute injury, the establishment of fibrosis during CKD may affect their potential to influence regeneration capacity. Here we tested the novel combination of MSCs with the antifibrotic serelaxin to repair and protect the kidney 7 d post-unilateral ureteral obstruction (UUO), when fibrosis is established. Male C57BL6 mice were sham-operated or UUO-inured (n = 4-6) and received vehicle, MSCs (1 × 10(6)), serelaxin (0.5 mg/kg per d), or the combination of both. In vivo tracing studies with luciferin/enhanced green fluorescent protein (eGFP)-tagged MSCs showed specific localization in the obstructed kidney where they remained for 36 h. Combination therapy conferred significant protection from UUO-induced fibrosis, as indicated by hydroxyproline analysis (P < 0.001 vs. vehicle, P < 0.05 vs. MSC or serelaxin alone). This was accompanied by preserved structural architecture, decreased tubular epithelial injury (P < 0.01 vs. MSCs alone), macrophage infiltration, and myofibroblast localization in the kidney (both P < 0.01 vs. vehicle). Combination therapy also stimulated matrix metalloproteinase (MMP)-2 activity over either treatment alone (P < 0.05 vs. either treatment alone). These results suggest that the presence of an antifibrotic in conjunction with MSCs ameliorates established kidney fibrosis and augments tissue repair to a greater extent than either treatment alone.

  20. Graptopetalum Paraguayense Ameliorates Chemical-Induced Rat Hepatic Fibrosis In Vivo and Inactivates Stellate Cells and Kupffer Cells In Vitro

    PubMed Central

    Su, Li-Jen; Chang, Chia-Chuan; Yang, Chih-Hsueh; Hsieh, Shur-Jong; Wu, Yi-Chin; Lai, Jin-Mei; Tseng, Tzu-Ling; Huang, Chi-Ying F.; Hsu, Shih-Lan

    2013-01-01

    Background Graptopetalum paraguayense (GP) is a folk herbal medicine with hepatoprotective effects that is used in Taiwan. The aim of this study was to evaluate the hepatoprotective and antifibrotic effects of GP on experimental hepatic fibrosis in both dimethylnitrosamine (DMN)- and carbon tetrachloride (CCl4)-induced liver injury rats. Methods Hepatic fibrosis-induced rats were fed with the methanolic extract of GP (MGP) by oral administration every day. Immunohistochemistry, biochemical assays, and Western blot analysis were performed. The effects of MGP on the expression of fibrotic markers and cytokines in the primary cultured hepatic stellate cells (HSCs) and Kupffer cells, respectively, were evaluated. Results Oral administration of MGP significantly alleviated DMN- or CCl4-induced liver inflammation and fibrosis. High levels of alanine transaminase, aspartate transaminase, bilirubin, prothrombin activity and mortality rates also decreased in rats treated with MGP. There were significantly decreased hydroxyproline levels in therapeutic rats compared with those of the liver-damaged rats. Collagen I and alpha smooth muscle actin (α-SMA) expression were all reduced by incubation with MGP in primary cultured rat HSCs. Furthermore, MGP induced apoptotic cell death in activated HSCs. MGP also suppressed lipopolysaccharide-stimulated rat Kupffer cell activation by decreasing nitric oxide, tumor necrosis factor-α and interleukin-6 production, and increasing interleukin-10 expression. Conclusions The results show that the administration of MGP attenuated toxin-induced hepatic damage and fibrosis in vivo and inhibited HSC and Kupffer cell activation in vitro, suggesting that MGP might be a promising complementary or alternative therapeutic agent for liver inflammation and fibrosis. PMID:23335984

  1. Effects of NOX1 on fibroblastic changes of endothelial cells in radiation-induced pulmonary fibrosis

    PubMed Central

    CHOI, SEO-HYUN; KIM, MISEON; LEE, HAE-JUNE; KIM, EUN-HO; KIM, CHUN-HO; LEE, YOON-JIN

    2016-01-01

    Lung fibrosis is a major complication in radiation-induced lung damage following thoracic radiotherapy, while the underlying mechanism has remained to be elucidated. The present study performed immunofluorescence and immunoblot assays on irradiated human pulmonary artery endothelial cells (HPAECs) with or without pre-treatment with VAS2870, a novel NADPH oxidase (NOX) inhibitor, or small hairpin (sh)RNA against NOX1, -2 or -4. VAS2870 reduced the cellular reactive oxygen species content induced by 5 Gy radiation in HPAECs and inhibited phenotypic changes in fibrotic cells, including increased alpha smooth muscle actin and vimentin, and decreased CD31 and vascular endothelial cadherin expression. These fibrotic changes were significantly inhibited by treatment with NOX1 shRNA, but not by NOX2 or NOX4 shRNA. Next, the role of NOX1 in pulmonary fibrosis development was assessed in the lung tissues of C57BL/6J mice following thoracic irradiation using trichrome staining. Administration of an NOX1-specific inhibitor suppressed radiation-induced collagen deposition and fibroblastic changes in the endothelial cells (ECs) of these mice. The results suggested that radiation-induced pulmonary fibrosis may be efficiently reduced by specific inhibition of NOX1, an effect mediated by reduction of fibrotic changes of ECs. PMID:27053172

  2. In liver fibrosis, dendritic cells govern hepatic inflammation in mice via TNF-alpha.

    PubMed

    Connolly, Michael K; Bedrosian, Andrea S; Mallen-St Clair, Jon; Mitchell, Aaron P; Ibrahim, Junaid; Stroud, Andrea; Pachter, H Leon; Bar-Sagi, Dafna; Frey, Alan B; Miller, George

    2009-11-01

    Hepatic fibrosis occurs during most chronic liver diseases and is driven by inflammatory responses to injured tissue. Because DCs are central to modulating liver immunity, we postulated that altered DC function contributes to immunologic changes in hepatic fibrosis and affects the pathologic inflammatory milieu within the fibrotic liver. Using mouse models, we determined the contribution of DCs to altered hepatic immunity in fibrosis and investigated the role of DCs in modulating the inflammatory environment within the fibrotic liver. We found that DC depletion completely abrogated the elevated levels of many inflammatory mediators that are produced in the fibrotic liver. DCs represented approximately 25% of the fibrotic hepatic leukocytes and showed an elevated CD11b+CD8- fraction, a lower B220+ plasmacytoid fraction, and increased expression of MHC II and CD40. Moreover, after liver injury, DCs gained a marked capacity to induce hepatic stellate cells, NK cells, and T cells to mediate inflammation, proliferation, and production of potent immune responses. The proinflammatory and immunogenic effects of fibrotic DCs were contingent on their production of TNF-alpha. Therefore, modulating DC function may be an attractive approach to experimental therapeutics in fibro-inflammatory liver disease.

  3. Use of mesenchymal stem cells to treat liver fibrosis: Current situation and future prospects

    PubMed Central

    Berardis, Silvia; Dwisthi Sattwika, Prenali; Najimi, Mustapha; Sokal, Etienne Marc

    2015-01-01

    Progressive liver fibrosis is a major health issue for which no effective treatment is available, leading to cirrhosis and orthotopic liver transplantation. However, organ shortage is a reality. Hence, there is an urgent need to find alternative therapeutic strategies. Cell-based therapy using mesenchymal stem cells (MSCs) may represent an attractive therapeutic option, based on their immunomodulatory properties, their potential to differentiate into hepatocytes, allowing the replacement of damaged hepatocytes, their potential to promote residual hepatocytes regeneration and their capacity to inhibit hepatic stellate cell activation or induce their apoptosis, particularly via paracrine mechanisms. The current review will highlight recent findings regarding the input of MSC-based therapy for the treatment of liver fibrosis, from in vitro studies to pre-clinical and clinical trials. Several studies have shown the ability of MSCs to reduce liver fibrosis and improve liver function. However, despite these promising results, some limitations need to be considered. Future prospects will also be discussed in this review. PMID:25624709

  4. Knockout of Endothelial Cell-Derived Endothelin-1 Attenuates Skin Fibrosis but Accelerates Cutaneous Wound Healing

    PubMed Central

    Makino, Katsunari; Jinnin, Masatoshi; Aoi, Jun; Kajihara, Ikko; Makino, Takamitsu; Fukushima, Satoshi; Sakai, Keisuke; Nakayama, Kazuhiko; Emoto, Noriaki; Yanagisawa, Masashi; Ihn, Hironobu

    2014-01-01

    Endothelin (ET)-1 is known for the most potent vasoconstrictive peptide that is released mainly from endothelial cells. Several studies have reported ET-1 signaling is involved in the process of wound healing or fibrosis as well as vasodilation. However, little is known about the role of ET-1 in these processes. To clarify its mechanism, we compared skin fibrogenesis and wound repair between vascular endothelial cell-specific ET-1 knockout mice and their wild-type littermates. Bleomycin-injected fibrotic skin of the knockout mice showed significantly decreased skin thickness and collagen content compared to that of wild-type mice, indicating that bleomycin-induced skin fibrosis is attenuated in the knockout mice. The mRNA levels of transforming growth factor (TGF)-β were decreased in the bleomycin-treated skin of ET-1 knockout mice. On the other hand, skin wound healing was accelerated in ET-1 knockout mice, which was indicated by earlier granulation tissue reduction and re-epithelialization in these mice. The mRNA levels of TGF-β, tumor necrosis factor (TNF)-α and connective tissue growth factor (CTGF) were reduced in the wound of ET-1 knockout mice. In endothelial ET-1 knockout mouse, the expression of TNF-α, CTGF and TGF-β was down-regulated. Bosentan, an antagonist of dual ET receptors, is known to attenuate skin fibrosis and accelerate wound healing in systemic sclerosis, and such contradictory effect may be mediated by above molecules. The endothelial cell-derived ET-1 is the potent therapeutic target in fibrosis or wound healing, and investigations of the overall regulatory mechanisms of these pathological conditions by ET-1 may lead to a new therapeutic approach. PMID:24853267

  5. Atrial fibrosis in a chronic murine model of obstructive sleep apnea: mechanisms and prevention by mesenchymal stem cells

    PubMed Central

    2014-01-01

    Background OSA increases atrial fibrillation (AF) risk and is associated with poor AF treatment outcomes. However, a causal association is not firmly established and the mechanisms involved are poorly understood. The aims of this work were to determine whether chronic obstructive sleep apnea (OSA) induces an atrial pro-arrhythmogenic substrate and to explore whether mesenchymal stem cells (MSC) are able to prevent it in a rat model of OSA. Methods A custom-made setup was used to mimic recurrent OSA-like airway obstructions in rats. OSA-rats (n = 16) were subjected to 15-second obstructions, 60 apneas/hour, 6 hours/day during 21 consecutive days. Sham rats (n = 14) were placed in the setup but no obstructions were applied. In a second series of rats, MSC were administered to OSA-rats and saline to Sham-rats. Myocardial collagen deposit was evaluated in Picrosirius-red stained samples. mRNA expression of genes involved in collagen turnover, inflammation and oxidative stress were quantified by real time PCR. MMP-2 protein levels were quantified by Western Blot. Results A 43% greater interstitial collagen fraction was observed in the atria, but not in the ventricles, of OSA-rats compared to Sham-rats (Sham 8.32 ± 0.46% vs OSA 11.90 ± 0.59%, P < 0.01). Angiotensin-I Converting Enzyme (ACE) and Interleukin 6 (IL-6) expression were significantly increased in both atria, while Matrix Metalloproteinase-2 (MMP-2) expression was decreased. MSC administration blunted OSA-induced atrial fibrosis (Sham + Saline 8.39 ± 0.56% vs OSA + MSC 9.57 ± 0.31%, P = 0.11), as well as changes in MMP-2 and IL-6 expression. Interleukin 1-β (IL-1β) plasma concentration correlated to atrial but not ventricular fibrosis. Notably, a 2.5-fold increase in IL-1β plasma levels was observed in the OSA group, which was prevented in rats receiving MSC. Conclusions OSA induces selective atrial fibrosis in a chronic murine model, which can be mediated in part

  6. Interferon-γ production by tubulointerstitial human CD56(bright) natural killer cells contributes to renal fibrosis and chronic kidney disease progression.

    PubMed

    Law, Becker M P; Wilkinson, Ray; Wang, Xiangju; Kildey, Katrina; Lindner, Mae; Rist, Melissa J; Beagley, Kenneth; Healy, Helen; Kassianos, Andrew J

    2017-04-08

    Natural killer (NK) cells are a population of lymphoid cells that play a significant role in mediating innate immune responses. Studies in mice suggest a pathological role for NK cells in models of kidney disease. In this study, we characterized the NK cell subsets present in native kidneys of patients with tubulointerstitial fibrosis, the pathological hallmark of chronic kidney disease. Significantly higher numbers of total NK cells (CD3(-)CD56(+)) were detected in renal biopsies with tubulointerstitial fibrosis compared with diseased biopsies without fibrosis and healthy kidney tissue using multi-color flow cytometry. At a subset level, both the CD56(dim) NK cell subset and particularly the CD56(bright) NK cell subset were elevated in fibrotic kidney tissue. However, only CD56(bright) NK cells significantly correlated with the loss of kidney function. Expression of the tissue-retention and -activation molecule CD69 on CD56(bright) NK cells was significantly increased in fibrotic biopsy specimens compared with non-fibrotic kidney tissue, indicative of a pathogenic phenotype. Further flow cytometric phenotyping revealed selective co-expression of activating receptor CD335 (NKp46) and differentiation marker CD117 (c-kit) on CD56(bright) NK cells. Multi-color immunofluorescent staining of fibrotic kidney tissue localized the accumulation of NK cells within the tubulointerstitium, with CD56(bright) NK cells (NKp46(+) CD117(+)) identified as the source of pro-inflammatory cytokine interferon-γ within the NK cell compartment. Thus, activated interferon-γ-producing CD56(bright) NK cells are positioned to play a key role in the fibrotic process and progression to chronic kidney disease.

  7. MicroRNA-378 limits activation of hepatic stellate cells and liver fibrosis by suppressing Gli3 expression

    PubMed Central

    Hyun, Jeongeun; Wang, Sihyung; Kim, Jieun; Rao, Kummara Madhusudana; Park, Soo Yong; Chung, Ildoo; Ha, Chang-Sik; Kim, Sang-Woo; Yun, Yang H.; Jung, Youngmi

    2016-01-01

    Hedgehog (Hh) signalling regulates hepatic fibrogenesis. MicroRNAs (miRNAs) mediate various cellular processes; however, their role in liver fibrosis is unclear. Here we investigate regulation of miRNAs in chronically damaged fibrotic liver. MiRNA profiling shows that expression of miR-378 family members (miR-378a-3p, miR-378b and miR-378d) declines in carbon tetrachloride (CCl4)-treated compared with corn-oil-treated mice. Overexpression of miR-378a-3p, directly targeting Gli3 in activated hepatic stellate cells (HSCs), reduces expression of Gli3 and profibrotic genes but induces gfap, the inactivation marker of HSCs, in CCl4-treated liver. Smo blocks transcriptional expression of miR-378a-3p by activating the p65 subunit of nuclear factor-κB (NF-κB). The hepatic level of miR-378a-3p is inversely correlated with the expression of Gli3 in tumour and non-tumour tissues in human hepatocellular carcinoma. Our results demonstrate that miR-378a-3p suppresses activation of HSCs by targeting Gli3 and its expression is regulated by Smo-dependent NF-κB signalling, suggesting miR-378a-3p has therapeutic potential for liver fibrosis. PMID:27001906

  8. Streptomycin treatment alters the intestinal microbiome, pulmonary T cell profile and airway hyperresponsiveness in a cystic fibrosis mouse model

    PubMed Central

    Bazett, Mark; Bergeron, Marie-Eve; Haston, Christina K.

    2016-01-01

    Cystic fibrosis transmembrane conductance regulator deficient mouse models develop phenotypes of relevance to clinical cystic fibrosis (CF) including airway hyperresponsiveness, small intestinal bacterial overgrowth and an altered intestinal microbiome. As dysbiosis of the intestinal microbiota has been recognized as an important contributor to many systemic diseases, herein we investigated whether altering the intestinal microbiome of BALB/c Cftrtm1UNC mice and wild-type littermates, through treatment with the antibiotic streptomycin, affects the CF lung, intestinal and bone disease. We demonstrate that streptomycin treatment reduced the intestinal bacterial overgrowth in Cftrtm1UNC mice and altered the intestinal microbiome similarly in Cftrtm1UNC and wild-type mice, principally by affecting Lactobacillus levels. Airway hyperresponsiveness of Cftrtm1UNC mice was ameliorated with streptomycin, and correlated with Lactobacillus abundance in the intestine. Additionally, streptomycin treated Cftrtm1UNC and wild-type mice displayed an increased percentage of pulmonary and mesenteric lymph node Th17, CD8 + IL-17+ and CD8 + IFNγ+ lymphocytes, while the CF-specific increase in respiratory IL-17 producing γδ T cells was decreased in streptomycin treated Cftrtm1UNC mice. Bone disease and intestinal phenotypes were not affected by streptomycin treatment. The airway hyperresponsiveness and lymphocyte profile of BALB/c Cftrtm1UNC mice were affected by streptomycin treatment, revealing a potential intestinal microbiome influence on lung response in BALB/c Cftrtm1UNC mice. PMID:26754178

  9. Correlation of serum liver fibrosis markers with severity of liver dysfunction in liver cirrhosis: a retrospective cross-sectional study

    PubMed Central

    Zhu, Cuihong; Qi, Xingshun; Li, Hongyu; Peng, Ying; Dai, Junna; Chen, Jiang; Xia, Chunlian; Hou, Yue; Zhang, Wenwen; Guo, Xiaozhong

    2015-01-01

    Hyaluronic acid (HA), laminin (LN), amino-terminal pro-peptide of type III pro-collagen (PIIINP), and collagen IV (CIV) are four major serum markers of liver fibrosis. This retrospective cross-sectional study aimed to evaluate the correlations of the four serum markers with the severity of liver dysfunction in cirrhotic patients. Between January 2013 and June 2014, a total of 228 patients with a clinical diagnosis with liver cirrhosis and without malignancy underwent the tests of HA, LN, PIIINP, and CIV levels. Laboratory data were collected. Child-Pugh and model for the end-stage of liver diseases (MELD) scores were calculated. Of them, 32%, 40%, and 18% had Child-Pugh class A, B, and C, respectively. MELD score was 7.58±0.50. HA (coefficient r: 0.1612, P=0.0203), LN (coefficient r: 0.2445, P=0.0004), and CIV (coefficient r: 0.2361, P=0.0006) levels significantly correlated with Child-Pugh score, but not PIIINP level. Additionally, LN (coefficient r: 0.2588, P=0.0002) and CIV (coefficient r: 0.1795, P=0.0108) levels significantly correlated with MELD score, but not HA or PIIINP level. In conclusions, HA, LN, and CIV levels might be positively associated with the severity of liver dysfunction in cirrhotic patients. However, given a relatively weak correlation between them, our findings should be cautiously interpreted and further validated. PMID:26131195

  10. Estimation and correlative study of salivary nitrate and nitrite in tobacco related oral squamous carcinoma and submucous fibrosis

    PubMed Central

    Shende, Vaishali; Biviji, AT; Akarte, N

    2013-01-01

    Oral cancer is one of the ten leading cancers of the world. In India, it is one of the common cancers and is an important public health problem. Tobacco plays significant role in etiology of oral squamous carcinoma. Tobacco which is chewed or smoked contains many alkaloids which are known carcinogens. Oral submucous fibrosis (OSMF) is a disease of the Indian subcontinent, which through immigration has a worldwide distribution. Betel nut chewing plays significant role in etiology of OSMF. The nut alkaloids have been shown experimentally to result in stimulation of collagen synthesis by fibroblasts in vitro, which can induce precancerous conditions. Materials and Methods: The present study was undertaken to detect nitrate and nitrite factor in saliva of cases with oral carcinoma, OSMF and normal individuals without any habits and to determine whether increased salivary nitrate and nitrite level is significant in oral carcinoma and submucous fibrosis using biochemical parameters. Conclusion: We conclude that the major inducer of oral squamous cell carcinoma (OSCC) is exposure to tobacco. Recent studies have demonstrated that oxidative and nitrosative stress contributes to the development of oral carcinogenesis through deoxyribonucleic acid (DNA) damage. Salivary composition of OSCC patients is substantially altered with respect to free radical-involved mechanisms. PMID:24574656

  11. Impaired Cell Volume Regulation in Intestinal Crypt Epithelia of Cystic Fibrosis Mice

    NASA Astrophysics Data System (ADS)

    Valverde, M. A.; O'Brien, J. A.; Sepulveda, F. V.; Ratcliff, R. A.; Evans, M. J.; Colledge, W. H.

    1995-09-01

    Cystic fibrosis is a disease characterized by abnormalities in the epithelia of the lungs, intestine, salivary and sweat glands, liver, and reproductive systems, often as a result of inadequate hydration of their secretions. The primary defect in cystic fibrosis is the altered activity of a cAMP-activated Cl^- channel, the cystic fibrosis transmembrane conductance regulator (CFTR) channel. However, it is not clear how a defect in the CFTR Cl^- channel function leads to the observed pathological changes. Although much is known about the structural properties and regulation of the CFTR, little is known of its relationship to cellular functions other than the cAMP-dependent Cl^- secretion. Here we report that cell volume regulation after hypotonic challenge is also defective in intestinal crypt epithelial cells isolated from CFTR -/- mutant mice. Moreover, the impairment of the regulatory volume decrease in CFTR -/- crypts appears to be related to the inability of a K^+ conductance to provide a pathway for the exit of this cation during the volume adjustments. This provides evidence that the lack of CFTR protein may have additional consequences for the cellular function other than the abnormal cAMP-mediated Cl^- secretion.

  12. BET inhibitors block pancreatic stellate cell collagen I production and attenuate fibrosis in vivo

    PubMed Central

    Kumar, Krishan; DeCant, Brian T.; Grippo, Paul J.; Hwang, Rosa F.; Bentrem, David J.; Ebine, Kazumi

    2017-01-01

    The fibrotic reaction, which can account for over 70%–80% of the tumor mass, is a characteristic feature of human pancreatic ductal adenocarcinoma (PDAC) tumors. It is associated with activation and proliferation of pancreatic stellate cells (PSCs), which are key regulators of collagen I production and fibrosis in vivo. In this report, we show that members of the bromodomain and extraterminal (BET) family of proteins are expressed in primary PSCs isolated from human PDAC tumors, with BRD4 positively regulating, and BRD2 and BRD3 negatively regulating, collagen I expression in primary cancer-associated PSCs. We show that the inhibitory effect of pan-BET inhibitors on collagen I expression in primary cancer-associated PSCs is through blocking of BRD4 function. Importantly, we show that FOSL1 is repressed by BRD4 in primary cancer-associated PSCs and negatively regulates collagen I expression. While BET inhibitors do not affect viability or induce PSC apoptosis or senescence, BET inhibitors induce primary cancer-associated PSCs to become quiescent. Finally, we show that BET inhibitors attenuate stellate cell activation, fibrosis, and collagen I production in the EL-KrasG12D transgenic mouse model of pancreatic tumorigenesis. Our results demonstrate that BET inhibitors regulate fibrosis by modulating the activation and function of cancer-associated PSCs. PMID:28194432

  13. CD133(+) human umbilical cord blood stem cells enhance angiogenesis in experimental chronic hepatic fibrosis.

    PubMed

    Elkhafif, Nagwa; El Baz, Hanan; Hammam, Olfat; Hassan, Salwa; Salah, Faten; Mansour, Wafaa; Mansy, Soheir; Yehia, Hoda; Zaki, Ahmed; Magdy, Ranya

    2011-01-01

    The in vivo angiogenic potential of transplanted human umbilical cord blood (UCB) CD133(+) stem cells in experimental chronic hepatic fibrosis induced by murine schistosomiasis was studied. Enriched cord blood-derived CD133(+) cells were cultured in primary medium for 3 weeks. Twenty-two weeks post-Schistosomiasis infection in mice, after reaching the chronic hepatic fibrotic stage, transplantation of stem cells was performed and mice were sacrificed 3 weeks later. Histopathology and electron microscopy showed an increase in newly formed blood vessels and a decrease in the fibrosis known for this stage of the disease. By immunohistochemical analysis the newly formed blood vessels showed positive expression of the human-specific angiogenic markers CD31, CD34 and von Willebrand factor. Few hepatocyte-like polygonal cells showed positive expression of human vascular endothelial growth factor and inducible nitric oxide synthase. The transplanted CD133(+) human stem cells primarily enhanced hepatic angiogenesis and neovascularization and contributed to repair in a paracrine manner by creating a permissive environment that enabled proliferation and survival of damaged cells rather than by direct differentiation to hepatocytes. A dual advantage of CD133(+) cell therapy in hepatic disease is suggested based on its capability of hematopoietic and endothelial differentiation.

  14. Correlations of Hepatic Hemodynamics, Liver Function, and Fibrosis Markers in Nonalcoholic Fatty Liver Disease: Comparison with Chronic Hepatitis Related to Hepatitis C Virus.

    PubMed

    Shigefuku, Ryuta; Takahashi, Hideaki; Nakano, Hiroyasu; Watanabe, Tsunamasa; Matsunaga, Kotaro; Matsumoto, Nobuyuki; Kato, Masaki; Morita, Ryo; Michikawa, Yousuke; Tamura, Tomohiro; Hiraishi, Tetsuya; Hattori, Nobuhiro; Noguchi, Yohei; Nakahara, Kazunari; Ikeda, Hiroki; Ishii, Toshiya; Okuse, Chiaki; Sase, Shigeru; Itoh, Fumio; Suzuki, Michihiro

    2016-09-14

    The progression of chronic liver disease differs by etiology. The aim of this study was to elucidate the difference in disease progression between chronic hepatitis C (CHC) and nonalcoholic fatty liver disease (NAFLD) by means of fibrosis markers, liver function, and hepatic tissue blood flow (TBF). Xenon computed tomography (Xe-CT) was performed in 139 patients with NAFLD and 152 patients with CHC (including liver cirrhosis (LC)). The cutoff values for fibrosis markers were compared between NAFLD and CHC, and correlations between hepatic TBF and liver function tests were examined at each fibrosis stage. The cutoff values for detection of the advanced fibrosis stage were lower in NAFLD than in CHC. Although portal venous TBF (PVTBF) correlated with liver function tests, PVTBF in initial LC caused by nonalcoholic steatohepatitis (NASH-LC) was significantly lower than that in hepatitis C virus (C-LC) (p = 0.014). Conversely, the liver function tests in NASH-LC were higher than those in C-LC (p < 0.05). It is important to recognize the difference between NAFLD and CHC. We concluded that changes in hepatic blood flow occurred during the earliest stage of hepatic fibrosis in patients with NAFLD; therefore, patients with NAFLD need to be followed carefully.

  15. Correlations of Hepatic Hemodynamics, Liver Function, and Fibrosis Markers in Nonalcoholic Fatty Liver Disease: Comparison with Chronic Hepatitis Related to Hepatitis C Virus

    PubMed Central

    Shigefuku, Ryuta; Takahashi, Hideaki; Nakano, Hiroyasu; Watanabe, Tsunamasa; Matsunaga, Kotaro; Matsumoto, Nobuyuki; Kato, Masaki; Morita, Ryo; Michikawa, Yousuke; Tamura, Tomohiro; Hiraishi, Tetsuya; Hattori, Nobuhiro; Noguchi, Yohei; Nakahara, Kazunari; Ikeda, Hiroki; Ishii, Toshiya; Okuse, Chiaki; Sase, Shigeru; Itoh, Fumio; Suzuki, Michihiro

    2016-01-01

    The progression of chronic liver disease differs by etiology. The aim of this study was to elucidate the difference in disease progression between chronic hepatitis C (CHC) and nonalcoholic fatty liver disease (NAFLD) by means of fibrosis markers, liver function, and hepatic tissue blood flow (TBF). Xenon computed tomography (Xe-CT) was performed in 139 patients with NAFLD and 152 patients with CHC (including liver cirrhosis (LC)). The cutoff values for fibrosis markers were compared between NAFLD and CHC, and correlations between hepatic TBF and liver function tests were examined at each fibrosis stage. The cutoff values for detection of the advanced fibrosis stage were lower in NAFLD than in CHC. Although portal venous TBF (PVTBF) correlated with liver function tests, PVTBF in initial LC caused by nonalcoholic steatohepatitis (NASH-LC) was significantly lower than that in hepatitis C virus (C-LC) (p = 0.014). Conversely, the liver function tests in NASH-LC were higher than those in C-LC (p < 0.05). It is important to recognize the difference between NAFLD and CHC. We concluded that changes in hepatic blood flow occurred during the earliest stage of hepatic fibrosis in patients with NAFLD; therefore, patients with NAFLD need to be followed carefully. PMID:27649152

  16. Regulatory T Cells Promote β-Catenin–Mediated Epithelium-to-Mesenchyme Transition During Radiation-Induced Pulmonary Fibrosis

    SciTech Connect

    Xiong, Shanshan; Pan, Xiujie; Xu, Long; Yang, Zhihua; Guo, Renfeng; Gu, Yongqing; Li, Ruoxi; Wang, Qianjun; Xiao, Fengjun; Du, Li; Zhou, Pingkun; Zhu, Maoxiang

    2015-10-01

    Purpose: Radiation-induced pulmonary fibrosis results from thoracic radiation therapy and severely limits radiation therapy approaches. CD4{sup +}CD25{sup +}FoxP3{sup +} regulatory T cells (Tregs) as well as epithelium-to-mesenchyme transition (EMT) cells are involved in pulmonary fibrosis induced by multiple factors. However, the mechanisms of Tregs and EMT cells in irradiation-induced pulmonary fibrosis remain unclear. In the present study, we investigated the influence of Tregs on EMT in radiation-induced pulmonary fibrosis. Methods and Materials: Mice thoraxes were irradiated (20 Gy), and Tregs were depleted by intraperitoneal injection of a monoclonal anti-CD25 antibody 2 hours after irradiation and every 7 days thereafter. Mice were treated on days 3, 7, and 14 and 1, 3, and 6 months post irradiation. The effectiveness of Treg depletion was assayed via flow cytometry. EMT and β-catenin in lung tissues were detected by immunohistochemistry. Tregs isolated from murine spleens were cultured with mouse lung epithelial (MLE) 12 cells, and short interfering RNA (siRNA) knockdown of β-catenin in MLE 12 cells was used to explore the effects of Tregs on EMT and β-catenin via flow cytometry and Western blotting. Results: Anti-CD25 antibody treatment depleted Tregs efficiently, attenuated the process of radiation-induced pulmonary fibrosis, hindered EMT, and reduced β-catenin accumulation in lung epithelial cells in vivo. The coculture of Tregs with irradiated MLE 12 cells showed that Tregs could promote EMT in MLE 12 cells and that the effect of Tregs on EMT was partially abrogated by β-catenin knockdown in vitro. Conclusions: Tregs can promote EMT in accelerating radiation-induced pulmonary fibrosis. This process is partially mediated through β-catenin. Our study suggests a new mechanism for EMT, promoted by Tregs, that accelerates radiation-induced pulmonary fibrosis.

  17. Therapeutic effect of hepatocyte growth factor-secreting mesenchymal stem cells in a rat model of liver fibrosis

    PubMed Central

    Kim, Myung-Deok; Kim, Sung-Soo; Cha, Hyun-Young; Jang, Seung-Hun; Chang, Da-Young; Kim, Wookhwan; Suh-Kim, Haeyoung; Lee, Jae-Ho

    2014-01-01

    Bone marrow-derived mesenchymal stromal cells (MSCs) have been reported to be beneficial for the treatment of liver fibrosis. Here, we investigated the use of genetically engineered MSCs that overexpress hepatocyte growth factor (HGF) as a means to improve their therapeutic effect in liver fibrosis. Liver fibrosis was induced by intraperitoneal injection of dimethylnitrosamine. HGF-secreting MSCs (MSCs/HGF) were prepared by transducing MSCs with an adenovirus carrying HGF-encoding cDNA. MSCs or MSCs/HGF were injected directly into the spleen of fibrotic rats. Tissue fibrosis was assessed by histological analysis 12 days after stem cell injection. Although treatment with MSCs reduced fibrosis, treatment with MSCs/HGF produced a more significant reduction and was associated with elevated HGF levels in the portal vein. Collagen levels in the liver extract were decreased after MSC/HGF therapy, suggesting recovery from fibrosis. Furthermore, liver function was improved in animals receiving MSCs/HGF, indicating that MSC/HGF therapy resulted not only in reduction of liver fibrosis but also in improvement of hepatocyte function. Assessment of cell and biochemical parameters revealed that mRNA levels of the fibrogenic cytokines PDGF-bb and TGF-β1 were significantly decreased after MSC/HGF therapy. Subsequent to the decrease in collagen, expression of matrix metalloprotease-9 (MMP-9), MMP-13, MMP-14 and urokinase-type plasminogen activator was augmented following MSC/HGF, whereas tissue inhibitor of metalloprotease-1 (TIMP-1) expression was reduced. In conclusion, therapy with MSCs/HGF resulted in an improved therapeutic effect compared with MSCs alone, probably because of the anti-fibrotic activity of HGF. Thus, MSC/HGF represents a promising approach toward a cell therapy for liver fibrosis. PMID:25145391

  18. Modulation of Stem Cell Differentiation and Myostatin as an Approach to Counteract Fibrosis in Muscle Dystrophy and Regeneration after Injury

    DTIC Science & Technology

    2011-03-01

    07-1-0181 TITLE: Modulation of Stem Cell Differentiation and Myostatin as an Approach to Counteract Fibrosis in Muscle Dystrophy and...views, opinions and/or findings contained in this report are those of the author(s) and should not be construed as an official Department of the Army...Myostatin as an Approach to Counteract Fibrosis in Muscle Dystrophy and Regeneration after Injury Dr. Nestor Gonzalez-Cadavid Charles R. Drew

  19. Complement effectors, C5a and C3a, in cystic fibrosis lung fluid correlate with disease severity

    PubMed Central

    Hair, Pamela S.; Sass, Laura A.; Vazifedan, Turaj; Shah, Tushar A.; Krishna, Neel K.

    2017-01-01

    In cystic fibrosis (CF), lung damage is mediated by a cycle of obstruction, infection, inflammation and tissue destruction. The complement system is a major mediator of inflammation for many diseases with the effectors C5a and C3a often playing important roles. We have previously shown in a small pilot study that CF sputum soluble fraction concentrations of C5a and C3a were associated with clinical measures of CF disease. Here we report a much larger study of 34 CF subjects providing 169 testable sputum samples allowing longitudinal evaluation comparing C5a and C3a with clinical markers. Levels of the strongly pro-inflammatory C5a correlated negatively with FEV1% predicted (P < 0.001), whereas the often anti-inflammatory C3a correlated positively with FEV1% predicted (P = 0.01). C5a concentrations correlated negatively with BMI percentile (P = 0.017), positively with worsening of an acute pulmonary exacerbation score (P = 0.007) and positively with P. aeruginosa growth in sputum (P = 0.002). C5a levels also correlated positively with concentrations of other sputum markers associated with worse CF lung disease including neutrophil elastase (P < 0.001), myeloperoxidase activity (P = 0.006) and DNA concentration (P < 0.001). In contrast to C5a, C3a levels correlated negatively with worse acute pulmonary exacerbation score and correlated negatively with sputum concentrations of neutrophil elastase, myeloperoxidase activity and DNA concentration. In summary, these data suggest that in CF sputum, increased C5a is associated with increased inflammation and poorer clinical measures, whereas increased C3a appears to be associated with less inflammation and improved clinical measures. PMID:28278205

  20. Effect of bevacizumab on the expression of fibrosis-related inflammatory mediators in ARPE-19 cells

    PubMed Central

    Chu, San-Jun; Zhang, Zhao-Hua; Wang, Min; Xu, Hai-Feng

    2017-01-01

    AIM To investigate the effect of anti-vascular epithelial growth factor (VEGF) agents on the expression of fibrosis-related inflammatory mediators under normoxic and hypoxic conditions, and to further clarify the mechanism underlying fibrosis after anti-VEGF therapy. METHODS Human retinal pigment epithelial (RPE) cells were incubated under normoxic and hypoxic conditions. For hypoxia treatment, CoCl2 at 200 µmol/L was added to the media. ARPE-19 cells were treated as following: 1) control group: no treatment; 2) bevacizumab group: bevacizumab at 0.25 mg/mL was added to the media; 3) hypoxia group: CoCl2 at 200 µmol/L was added to the media; 4) hypoxia+bevacizumab group: CoCl2 at 200 µmol/L and bevacizumab at 0.25 mg/mL were added to the media. The expression of interleukin (IL)-1β, IL-6, IL-8 and tumor necrosis factor (TNF)-α were evaluated using real-time polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) at 6, 12, 24 and 48h. RESULTS Both mRNA and protein levels of IL-1β, IL-6 and IL-8 were statistically significantly higher in the bevacizumab group than in the control group at each time point, and TNF-α gene and protein expression was only significantly higher only at 24 and 48h (P<0.05). Under hypoxic conditions, bevacizumab significantly increased the expression of IL-1β, IL-6, IL-8 and TNF-α at 6, 12, 24 and 48h (P<0.05). IL-1β, IL-8 and TNF-α peaked at 24h and IL-6 peaked at 12h after the bevacizumab treatment under both normoxic and hypoxic conditions. CONCLUSION Treatment of ARPE-19 cells with bevacizumab can significantly increase the expression of fibrosis-related inflammatory mediators under both normoxic and hypoxic conditions. Inflammatory factors might be involved in the process of fibrosis after anti-VEGF therapy, and the up-regulation of inflammatory factors induced by anti-VEGF drugs might promote the fibrosis process.

  1. Verrucoid Variant of Invasive Squamous Cell Carcinoma in Oral Submucous Fibrosis: A Clinicopathological Challenge

    PubMed Central

    Ramani, Priya; Krithika, C.; Ananthalakshmi, R.; Jagdish, Praveena; Janardhanan, Sunitha; Jeevakarunyam, Sathiyajeeva

    2016-01-01

    Verrucous carcinoma (VC) is an exophytic, low-grade, well-differentiated variant of squamous cell carcinoma. It is described as a lesion appearing in the sixth or seventh decade of life that has minimal aggressive potential and, in long-standing cases, has been shown to transform into squamous cell carcinoma. Oral submucous fibrosis (OSMF) is a potentially malignant disorder, and about one-third of the affected population develop oral squamous cell carcinoma. The histopathological diagnosis of verrucous carcinoma is challenging, and the interpretation of early squamous cell carcinoma requires immense experience. Here we present a rare case of a 24-year-old male with OSMF transforming to verrucous carcinoma with invasive squamous cell carcinoma. Even though the case had a straightforward clinical diagnosis, the serial sectioning done for pathological diagnosis disclosed the squamous cell carcinoma.

  2. A novel exon in the cystic fibrosis transmembrane conductance regulator gene activated by the nonsense mutation E92X in airway epithelial cells of patients with cystic fibrosis.

    PubMed Central

    Will, K; Dörk, T; Stuhrmann, M; Meitinger, T; Bertele-Harms, R; Tümmler, B; Schmidtke, J

    1994-01-01

    Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. We report on a novel nonsense mutation that leads to exon skipping and the activation of a cryptic exon. Screening of genomic DNA from 700 German patients with CF uncovered four cases with the nonsense mutation E92X, a G-->T transversion that creates a termination codon and affects the first base of exon 4 of the CFTR gene. Lymphocyte RNA of two CF patients heterozygous for E92X was found to contain the wild type sequence and a differentially spliced isoform lacking exon 4. In RNA derived from nasal epithelial cells of E92X patients, a third fragment of longer size was observed. Sequencing revealed the presence of E92X and an additional 183-bp fragment, inserted between exons 3 and 4. The 183-bp sequence was mapped to intron 3 of the CFTR gene. It is flanked by acceptor and donor splice sites. We conclude that the 183-bp fragment in intron 3 is a cryptic CFTR exon that can be activated in epithelial cells by the presence of the E92X mutation. E92X abolishes correctly spliced CFTR mRNA and leads to severe cystic fibrosis. Images PMID:7512993

  3. Betaine treatment decreased oxidative stress, inflammation, and stellate cell activation in rats with alcoholic liver fibrosis.

    PubMed

    Bingül, İlknur; Başaran-Küçükgergin, Canan; Aydın, A Fatih; Çoban, Jale; Doğan-Ekici, Işın; Doğru-Abbasoğlu, Semra; Uysal, Müjdat

    2016-07-01

    The aim of this study was to investigate the effect of betaine (BET) on alcoholic liver fibrosis in rats. Fibrosis was experimentally generated with ethanol plus carbon tetrachloride (ETH+CCl4) treatment. Rats were treated with ETH (5% v/v in drinking water) for 14 weeks. CCl4 was administered intraperitoneally (i.p.) 0.2mL/kg twice a week to rats in the last 6 weeks with/without commercial food containing BET (2% w/w). Serum hepatic damage markers, tumor necrosis factor-α, hepatic triglyceride (TG) and hydroxyproline (HYP) levels, and oxidative stress parameters were measured together with histopathologic observations. In addition, α-smooth muscle-actin (α-SMA), transforming growth factor-β1 (TGF-β1) and type I collagen (COL1A1) protein expressions were assayed immunohistochemically to evaluate stellate cell (HSC) activation. mRNA expressions of matrix metalloproteinase-2 (MMP-2) and its inhibitors (TIMP-1 and TIMP-2) were also determined. BET treatment diminished TG and HYP levels; prooxidant status and fibrotic changes; α-SMA, COL1A1 and TGF-β protein expressions; MMP-2, TIMP-1 and TIMP-2 mRNA expressions in the liver of fibrotic rats. In conclusion, these results indicate that the antifibrotic effect of BET may be related to its suppressive effects on oxidant and inflammatory processes together with HSC activation in alcoholic liver fibrosis.

  4. Hepatocellular carcinoma in chronic HBV-HCV co-infection is correlated to fibrosis and disease duration.

    PubMed

    Zampino, Rosa; Pisaturo, Maria A; Cirillo, Grazia; Marrone, Aldo; Macera, Margherita; Rinaldi, Luca; Stanzione, Maria; Durante-Mangoni, Emanuele; Gentile, Ivan; Sagnelli, Evangelista; Signoriello, Giuseppe; Miraglia Del Giudice, Emanuele; Adinolfi, Luigi E; Coppola, Nicola

    2015-01-01

    Hepatocellular carcinoma (HCC) is a development of severe liver disease frequently due to HBV and/or HCV infection. The aim of this retrospective study was to evaluate the development of HCC in patients with HBV-HCV chronic infection compared with patients with single HBV or HCV infection and the viral and host factors correlated to HCC in co-infected patients. We studied 268 patients with histology proven chronic hepatitis: 56 had HBV-HCV co-infection (HBV-HCV group), 46 had HBV infection (HBV group) and 166 had HCV infection (HCV group). Patients were followed up for at least 3 years. Viral and host factors were studied. HCC was more frequent in HBV-HCV group (14%) compared with HBV (2%, p = 0.006) and HCV monoinfected (4%, p = 0.006). The Mantel-Haenszel test used to investigate the relationship between HBV-HCV co-infection and development of HCC indicated an association between development of HCC and HBV-HCV co-infection (p < 0.001). In the HBV-HCV group, patients with HCC were significantly older (p = 0.000), had longer disease duration (p = 0.001), higher blood glucose levels (p = 0.001), lower levels of steatosis (p = 0.02), higher levels of fibrosis (p = 0.000), higher HCV RNA (p = 0.01) than those without HCC. ALT, lipid profile, PNPLA3 variant distribution and HBV viral load did not differ among co-infected patients with or without HCC. In conclusion HCC was more frequent in our patients with HBV-HCV co-infection, than in those with HBV or HCV mono-infection; possible associated risk factors for HCC development seem a long duration of disease, high levels of fibrosis and carbohydrate intolerance.

  5. Exosomes derived from human umbilical cord mesenchymal stem cells alleviate liver fibrosis.

    PubMed

    Li, Tingfen; Yan, Yongmin; Wang, Bingying; Qian, Hui; Zhang, Xu; Shen, Li; Wang, Mei; Zhou, Ying; Zhu, Wei; Li, Wei; Xu, Wenrong

    2013-03-15

    Mesenchymal stem cells (MSCs) have been considered as an attractive tool for the therapy of diseases. Exosomes excreted from MSCs can reduce myocardial ischemia/reperfusion damage and protect against acute tubular injury. However, whether MSC-derived exosomes can relieve liver fibrosis and its mechanism remain unknown. Previous work showed that human umbilical cord-MSCs (hucMSCs) transplanted into acutely injured and fibrotic livers could restore liver function and improve liver fibrosis. In this study, it was found that transplantation of exosomes derived from hucMSC (hucMSC-Ex) reduced the surface fibrous capsules and got their textures soft, alleviated hepatic inflammation and collagen deposition in carbon tetrachloride (CCl4)-induced fibrotic liver. hucMSC-Ex also significantly recovered serum aspartate aminotransferase (AST) activity, decreased collagen type I and III, transforming growth factor (TGF)-β1 and phosphorylation Smad2 expression in vivo. In further experiments, we found that epithelial-to-mesenchymal transition (EMT)-associated markers E-cadherin-positive cells increased and N-cadherin- and vimentin-positive cells decreased after hucMSC-Ex transplantation. Furthermore, the human liver cell line HL7702 underwent typical EMT after induction with recombinant human TGF-β1, and then hucMSC-Ex treatment reversed spindle-shaped and EMT-associated markers expression in vitro. Taken together, these results suggest that hucMSC-Ex could ameliorate CCl4-induced liver fibrosis by inhibiting EMT and protecting hepatocytes. This provides a novel approach for the treatment of fibrotic liver disease.

  6. Radiation-induced lung fibrosis after treatment of small cell carcinoma of the lung with very high-dose cyclophosphamide

    SciTech Connect

    Trask, C.W.; Joannides, T.; Harper, P.G.; Tobias, J.S.; Spiro, S.G.; Geddes, D.M.; Souhami, R.L.; Beverly, P.C.

    1985-01-01

    Twenty-five previously untreated patients with small cell carcinoma of the lung were treated with cyclophosphamide 160 to 200 mg/kg (with autologous bone marrow support) followed by radiotherapy (4000 cGy) to the primary site and mediastinum. No other treatment was given until relapse occurred. Nineteen patients were assessable at least 4 months after radiotherapy; of these, 15 (79%) developed radiologic evidence of fibrosis, which was symptomatic in 14 (74%). The time of onset of fibrosis was related to the volume of lung irradiated. A retrospective analysis was made of 20 consecutive patients treated with multiple-drug chemotherapy and an identical radiotherapy regimen as part of a randomized trial. Radiologic and symptomatic fibrosis was one half as frequent (35%) as in the high-dose cyclophosphamide group. Very high-dose cyclophosphamide appears to sensitize the lung to radiotherapy and promotes the production of fibrosis.

  7. Fibrosis and Subsequent Cytopenias are Associated with bFGF-deficient Pluripotent Mesenchymal Stromal Cells in Large Granular Lymphocyte Leukemia1

    PubMed Central

    Mailloux, Adam W.; Zhang, Ling; Moscinski, Lynn; Bennett, John M.; Yang, Lili; Yoder, Sean J.; Bloom, Gregory; Wei, Codi; Wei, Sheng; Sokol, Lubomir; Loughran, Thomas P.; Epling-Burnette, P.K.

    2015-01-01

    Cytopenias occur frequently in systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), Felty’s Syndrome and Large Granular Lymphocyte (LGL) leukemia, but the bone marrow microenvironment has not been systematically studied. In LGL leukemia (n=24), retrospective analysis of bone marrow (BM) histopathology revealed severe fibrosis in 15 out of 24 patients (63%) in association with the presence of cytopenias, occurrence of autoimmune diseases, and splenomegaly, but was undetectable in control cases with B-cell malignancies (n=11). Fibrosis severity correlated with T-LGL cell numbers in the BM but not in the periphery, suggesting deregulation is limited to the BM microenvironment. To identify fibrosis initiating populations, primary mesenchymal stromal cultures (MSCs) from patients were characterized and found to display proliferation kinetics and overabundant collagen deposition, but displayed normal telomere lengths and osteoblastogenic, chondrogenic, and adipogenic differentiation potentials. To determine the effect of fibrosis on healthy hematopoietic cells (HPCs), bioartificial matrixes from rat-tail or purified human collagen were found to suppress HPC differentiation and proliferation. The ability of patient MSCs to support healthy HSC proliferation was significantly impaired, but could be rescued with collagenase pre-treatment. Clustering analysis confirmed the undifferentiated state of patient MSCs, and pathway analysis revealed an inverse relationship between cell division and pro-fibrotic ontologies associated with reduced basic fibroblast growth factor (FGFb) production, which was confirmed by ELISA. Reconstitution with exogenous FGFb normalized patient MSC proliferation, collagen deposition, and HPC supportive function suggesting LGL BM infiltration and secondary accumulation of MSC-derived collagen is responsible for hematopoietic failure in autoimmune-associated cytopenias in LGL leukemia. PMID:24014875

  8. Functional expression and apical localization of the cystic fibrosis transmembrane conductance regulator in MDCK I cells.

    PubMed Central

    Mohamed, A; Ferguson, D; Seibert, F S; Cai, H M; Kartner, N; Grinstein, S; Riordan, J R; Lukacs, G L

    1997-01-01

    The gene product affected in cystic fibrosis, the cystic fibrosis transmembrane conductance regulator (CFTR), is a chlorideselective ion channel that is regulated by cAMP-dependent protein kinase-mediated phosphorylation, ATP binding and ATP hydrolysis. Mutations in the CFTR gene may result in cystic fibrosis characterized by severe pathology (e.g. recurrent pulmonary infection, male infertility and pancreatic insufficiency) involving organs expressing the CFTR. Interestingly, in the kidney, where expression of the CFTR has been reported, impaired ion transport in patients suffering from cystic fibrosis could not be observed. To understand the role of the CFTR in chloride transport in the kidney, we attempted to identify an epithelial cell line that can serve as a model. We demonstrate that the CFTR is expressed constitutively in Madine-Darby canine kidney (MDCK) type I cells, which are thought to have originated from the distal tubule of the dog nephron. We show expression at the mRNA level, using reverse transcriptase-PCR, and at the protein level, using Western blot analysis with three different monoclonal antibodies. Iodide efflux measurements indicate that CFTR expression confers a plasma membrane anion conductance that is responsive to stimulation by cAMP. The cAMP-stimulated iodide release is sensitive to glybenclamide, diphenylamine carboxylic acid and 5-nitro-2-(3-phenylpropylamino)benzoic acid, but not to 4,4'-di-isothiocyanostilbene-2,2'-disulphonic acid, an inhibitor profile characteristic of the CFTR chloride channel. Finally, the polarized localization of the CFTR to the apical plasma membrane was established by iodide efflux measurements and cell-surface biotinylation on MDCK I monolayers. Interestingly, MDCK type II cells, which are thought to have originated from the proximal tubule of the kidney, lack CFTR protein expression and cAMP-stimulated chloride conductance. In conclusion, we propose that MDCK type I and II cells can serve as convenient

  9. Detection of Hepatic Fibrosis in Ex Vivo Liver Samples Using an Open-Photoacoustic-Cell Method: Feasibility Study

    NASA Astrophysics Data System (ADS)

    Stolik, S.; Fabila, D. A.; de la Rosa, J. M.; Escobedo, G.; Suárez-Álvarez, K.; Tomás, S. A.

    2015-09-01

    Design of non-invasive and accurate novel methods for liver fibrosis diagnosis has gained growing interest. Different stages of liver fibrosis were induced in Wistar rats by intraperitoneally administering different doses of carbon tetrachloride. The liver fibrosis degree was conventionally determined by means of histological examination. An open-photoacoustic-cell (OPC) technique for the assessment of liver fibrosis was developed and is reported here. The OPC technique is based on the fact that the thermal diffusivity can be accurately measured by photoacoustics taking into consideration the photoacoustic signal amplitude versus the modulation frequency. This technique measures directly the heat generated in a sample, due to non-radiative de-excitation processes, following the absorption of light. The thermal diffusivity was measured with a home-made open-photoacoustic-cell system that was specially designed to perform the measurement from ex vivo liver samples. The human liver tissue showed a significant increase in the thermal diffusivity depending on the fibrosis stage. Specifically, liver samples from rats exhibiting hepatic fibrosis showed a significantly higher value of the thermal diffusivity than for control animals.

  10. Inhibition of connective tissue growth factor attenuates paraquat-induced lung fibrosis in a human MRC-5 cell line.

    PubMed

    Huang, Min; Yang, Huifang; Zhu, Lingqin; Li, Honghui; Zhou, Jian; Zhou, Zhijun

    2016-11-01

    Chronic exposure to Paraquat (PQ) may result in progressive pulmonary fibrosis and subsequent chronic obstructive pulmonary malfunction. Connective tissue growth factor (CTGF) has been proposed as a key determinant in the development of lung fibrosis. We investigated thus whether knock down of CTGF can prevent human lung fibroblasts (MRC-5) activation and proliferation with the subsequent inhibition of PQ-induced fibrosis. MRC-5 was transfected with CTGF-siRNAs and exposed to different concentrations of PQ. The siRNA-silencing efficacy was evaluated using western blotting analyses, qRT-PCR and flow cytometry. Next, the viability and migration of MRC-5 was determined. MMP-2, MMP-9, and TIMP-1 accumulation were quantified to evaluate the lung fibrosis exposure to PQ. Over expression of CTGF mRNA was observed in human MRC-5 cell as early as 6 h following PQ stimulation. CTGF gene expression in MRC-5 cells was substantially reduced by RNAi, which significantly suppressed the expression of the lung fibrosis markers such as tissue inhibitor of metalloproteinase-2 (TIMP-2), Matrix metalloproteinase-2 (MMP-2) and Matrix metalloproteinase-9 (MMP-9) that were stimulated by PQ. Inhibition of CTGF expression suppressed impeded the proliferation and migration ability of MRC-5 cells and resulted in cell-extracellular matrix (ECM) protein accumulation in cells. Our results suggest that CTGF promoted the development of PQ-induced lung fibrosis in collaboration with transforming growth factor β1 (TGFβ1). Furthermore, the observed arresting effects of CTGF knock down during this process suggested that CTGF is the potential target site for preventing PQ-induced pulmonary fibrosis. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1620-1626, 2016.

  11. Bone marrow mesenchymal stem cells attenuate silica-induced pulmonary fibrosis via paracrine mechanisms.

    PubMed

    Li, Xiaoli; Wang, Yan; An, Guoliang; Liang, Di; Zhu, Zhonghui; Lian, Ximeng; Niu, Piye; Guo, Caixia; Tian, Lin

    2017-03-15

    The purpose of this study was to investigate the anti-fibrotic effect and possible mechanism of bone marrow mesenchymal stem cells (BMSCs) in silica-induced lung injury and fibrosis in vivo and in vitro. In vivo, rats were exposed to 50mg/ml silica intratracheally. The rats were sacrificed on day 15 or day 30 after intravenous injection of BMSCs. Histopathological examination demonstrated that BMSCs decreased the blue areas of collagen fibers and the number of nodules. Alveolar epithelium was damaged by silica, but it was restored by BMSCs. In vitro, BMSCs co-cultured with RLE-6TN cells in 6-Transwell plates were evaluated to determine the possible mechanism. The results demonstrated that BMSCs downregulated the expression of collagen I and III. BMSCs reversed morphological abnormalities and reduced the proliferation of RLE-6TN cells. These data showed that BMSCs did not give rise to alveolar epithelial cells directly, while the levels of hepatocyte growth factor, keratinocyte growth factor and bone morphogenetic protein -7 increased and expression of tumor necrosis factor-α and transforming growth factor-β1 decreased in the 6TN+Silica+BMSCs group compared with the 6TN+Silica group. Our results revealed that BMSCs exerted anti-fibrotic effects on silica-induced pulmonary fibrosis, which might be associated with paracrine mechanisms rather than differentiation.

  12. NADPH oxidase signal transduces angiotensin II in hepatic stellate cells and is critical in hepatic fibrosis

    PubMed Central

    Bataller, Ramón; Schwabe, Robert F.; Choi, Youkyung H.; Yang, Liu; Paik, Yong Han; Lindquist, Jeffrey; Qian, Ting; Schoonhoven, Robert; Hagedorn, Curt H.; Lemasters, John J.; Brenner, David A.

    2003-01-01

    Angiotensin II (Ang II) is a pro-oxidant and fibrogenic cytokine. We investigated the role of NADPH oxidase in Ang II–induced effects in hepatic stellate cells (HSCs), a fibrogenic cell type. Human HSCs express mRNAs of key components of nonphagocytic NADPH oxidase. Ang II phosphorylated p47phox, a regulatory subunit of NADPH oxidase, and induced reactive oxygen species formation via NADPH oxidase activity. Ang II phosphorylated AKT and MAPKs and increased AP-1 DNA binding in a redox-sensitive manner. Ang II stimulated DNA synthesis, cell migration, procollagen α1(I) mRNA expression, and secretion of TGF-β1 and inflammatory cytokines. These effects were attenuated by N-acetylcysteine and diphenylene iodonium, an NADPH oxidase inhibitor. Moreover, Ang II induced upregulation of genes potentially involved in hepatic wound-healing response in a redox-sensitive manner, as assessed by microarray analysis. HSCs isolated from p47phox–/– mice displayed a blunted response to Ang II compared with WT cells. We also assessed the role of NADPH oxidase in experimental liver fibrosis. After bile duct ligation, p47phox–/– mice showed attenuated liver injury and fibrosis compared with WT counterparts. Moreover, expression of smooth muscle α-actin and expression of TGF-β1 were reduced in p47phox–/– mice. Thus, NADPH oxidase mediates the actions of Ang II on HSCs and plays a critical role in liver fibrogenesis. PMID:14597764

  13. Origin of Matrix-Producing Cells That Contribute to Aortic Fibrosis in Hypertension.

    PubMed

    Wu, Jing; Montaniel, Kim Ramil C; Saleh, Mohamed A; Xiao, Liang; Chen, Wei; Owens, Gary K; Humphrey, Jay D; Majesky, Mark W; Paik, David T; Hatzopoulos, Antonis K; Madhur, Meena S; Harrison, David G

    2016-02-01

    Various hypertensive stimuli lead to exuberant adventitial collagen deposition in large arteries, exacerbating blood pressure elevation and end-organ damage. Collagen production is generally attributed to resident fibroblasts; however, other cells, including resident and bone marrow-derived stem cell antigen positive (Sca-1(+)) cells and endothelial and vascular smooth muscle cells, can produce collagen and contribute to vascular stiffening. Using flow cytometry and immunofluorescence, we found that adventitial Sca-1(+) progenitor cells begin to produce collagen and acquire a fibroblast-like phenotype in hypertension. We also found that bone marrow-derived cells represent more than half of the matrix-producing cells in hypertension, and that one-third of these are Sca-1(+). Cell sorting and lineage-tracing studies showed that cells of endothelial origin contribute to no more than one fourth of adventitial collagen I(+) cells, whereas those of vascular smooth muscle lineage do not contribute. Our findings indicate that Sca-1(+) progenitor cells and bone marrow-derived infiltrating fibrocytes are major sources of arterial fibrosis in hypertension. Endothelial to mesenchymal transition likely also contributes, albeit to a lesser extent and pre-existing resident fibroblasts represent a minority of aortic collagen-producing cells in hypertension. This study shows that vascular stiffening represents a complex process involving recruitment and transformation of multiple cells types that ultimately elaborate adventitial extracellular matrix.

  14. Activated Wnt signaling induces myofibroblast differentiation of mesenchymal stem cells, contributing to pulmonary fibrosis.

    PubMed

    Sun, Zhaorui; Wang, Cong; Shi, Chaowen; Sun, Fangfang; Xu, Xiaomeng; Qian, Weiping; Nie, Shinan; Han, Xiaodong

    2014-05-01

    Acute lung injury may lead to fibrogenesis. However, no treatment is currently available. This study was conducted to determine the effects of bone marrow-derived mesenchymal stem cells (MSCs) in a model of HCl-induced acute lung injury in Sprague-Dawley (SD) rats. Stromal cell-derived factor (SDF)-1 and its receptor CXC chemokine receptor (CXCR)4 have been shown to participate in mobilizing MSCs. Adenovirus carrying the CXCR4 gene was used to transfect MSCs in order to increase the engraftment numbers of MSCs at injured sites. Histological examination data demonstrated that the engraftment of MSCs did not attenuate lung injury and pulmonary fibrosis. The results showed that engraftment of MSCs almost differentiated into myofibroblasts, but rarely differentiated into lung epithelial cells. Additionally, it was demonstrated that activated canonical Wnt/β-catenin signaling in injured lung tissue regulated the myofibroblast differentiation of MSCs in vivo. The in vitro study results demonstrated that activation of the Wnt/β-catenin signaling stimulated MSCs to express myofibroblast markers; however, this process was attenuated by Wnt antagonist DKK1. Therefore, the results demonstrated that the aberrant activation of Wnt signaling induces the myofibroblast differentiation of engrafted MSCs, thus contributing to pulmonary fibrosis following lung injury.

  15. Biphasic recruitment of microchimeric fetal mesenchymal cells in fibrosis following acute kidney injury.

    PubMed

    Roy, Edwige; Seppanen, Elke; Ellis, Rebecca; Lee, Eddy S; Khosrotehrani, Kiarash; Khosroterani, Kiarash; Fisk, Nicholas M; Bou-Gharios, George

    2014-03-01

    Fetal microchimeric cells (FMCs) enter the maternal circulation and persist in tissue for decades. They have capacity to home to injured maternal tissue and differentiate along that tissue's lineage. This raises the question of the origin(s) of cells transferred to the mother during pregnancy. FMCs with a mesenchymal phenotype have been documented in several studies, which makes mesenchymal stem cells an attractive explanation for their broad plasticity. Here we assessed the recruitment and mesenchymal lineage contribution of FMCs in response to acute kidney fibrosis induced by aristolochic acid injection. Serial in vivo bioluminescence imaging revealed a biphasic recruitment of active collagen-producing FMCs during the repair process of injured kidney in post-partum wild-type mothers that had delivered transgenic pups expressing luciferase under the collagen type I-promoter. The presence of FMCs long-term post injury (day 60) was associated with profibrotic molecules (TGF-β/CTGF), serum urea levels, and collagen deposition. Immunostaining confirmed FMCs at short term (day 15) using post-partum wild-type mothers that had delivered green fluorescent protein-positive pups and suggested a mainly hematopoietic phenotype. We conclude that there is biphasic recruitment to, and activity of, FMCs at the injury site. Moreover, we identified five types of FMC, implicating them all in the reparative process at different stages of induced renal interstitial fibrosis.

  16. Correlation of sweat chloride and percent predicted FEV1 in cystic fibrosis patients treated with ivacaftor.

    PubMed

    Fidler, Meredith C; Beusmans, Jack; Panorchan, Paul; Van Goor, Fredrick

    2017-01-01

    Ivacaftor, a CFTR potentiator that enhances chloride transport by acting directly on CFTR to increase its channel gating activity, has been evaluated in patients with different CFTR mutations. Several previous analyses have reported no statistical correlation between change from baseline in ppFEV1 and reduction in sweat chloride levels for individuals treated with ivacaftor. The objective of the post hoc analysis described here was to expand upon previous analyses and evaluate the correlation between sweat chloride levels and absolute ppFEV1 changes across multiple cohorts of patients with different CF-causing mutations who were treated with ivacaftor. The goal of the analysis was to help define the potential value of sweat chloride as a pharmacodynamic biomarker for use in CFTR modulator trials. For any given study, reductions in sweat chloride levels and improvements in absolute ppFEV1 were not correlated for individual patients. However, when the data from all studies were combined, a statistically significant correlation between sweat chloride levels and ppFEV1 changes was observed (p<0.0001). Thus, sweat chloride level changes in response to potentiation of the CFTR protein by ivacaftor appear to be a predictive pharmacodynamic biomarker of lung function changes on a population basis but are unsuitable for the prediction of treatment benefits for individuals.

  17. Fat on sale: role of adipose-derived stem cells as anti-fibrosis agent in regenerative medicine.

    PubMed

    Gupta, Manoj K; Ajay, Amrendra Kumar

    2015-12-01

    The potential use of stem cells for cell-based tissue repair and regeneration offers alternative therapeutic strategies for various diseases. Adipose-derived stem cells (ADSCs) have emerged as a promising source of stem cells suitable for transplantation in regenerative medicine and wound repair. A recent publication in Stem Cell Research & Therapy by Zhang and colleagues reports a new finding about the anti-fibrosis role of ADSCs and conditioned media derived from them on hypertrophic scar formation in vivo.

  18. Liver Fibrosis and Protection Mechanisms Action of Medicinal Plants Targeting Apoptosis of Hepatocytes and Hepatic Stellate Cells

    PubMed Central

    Moreno-Cuevas, Jorge E.; González-Garza, Maria Teresa; Rodríguez-Montalvo, Carlos; Cruz-Vega, Delia Elva

    2014-01-01

    Following chronic liver injury, hepatocytes undergo apoptosis leading to activation of hepatic stellate cells (HSC). Consequently, activated HSC proliferate and produce excessive extracellular matrix, responsible for the scar formation. The pandemic trend of obesity, combined with the high incidence of alcohol intake and viral hepatitis infections, highlights the urgent need to find accessible antifibrotic therapies. Treatment strategies should take into account the versatility of its pathogenesis and act on all the cell lines involved to reduce liver fibrosis. Medicinal plants are achieving popularity as antifibrotic agents, supported by their safety, cost-effectiveness, and versatility. This review will describe the role of hepatocytes and HSC in the pathogenesis of liver fibrosis and detail the mechanisms of modulation of apoptosis of both cell lines by twelve known hepatoprotective plants in order to reduce liver fibrosis. PMID:25505905

  19. Autophagy in hepatic fibrosis.

    PubMed

    Song, Yang; Zhao, Yingying; Wang, Fei; Tao, Lichan; Xiao, Junjie; Yang, Changqing

    2014-01-01

    Hepatic fibrosis is a leading cause of morbidity and mortality worldwide. Hepatic fibrosis is usually associated with chronic liver diseases caused by infection, drugs, metabolic disorders, or autoimmune imbalances. Effective clinical therapies are still lacking. Autophagy is a cellular process that degrades damaged organelles or protein aggregation, which participates in many pathological processes including liver diseases. Autophagy participates in hepatic fibrosis by activating hepatic stellate cells and may participate as well through influencing other fibrogenic cells. Besides that, autophagy can induce some liver diseases to develop while it may play a protective role in hepatocellular abnormal aggregates related liver diseases and reduces fibrosis. With a better understanding of the potential effects of autophagy on hepatic fibrosis, targeting autophagy might be a novel therapeutic strategy for hepatic fibrosis in the near future.

  20. TGF-β1 Induces the Dual Regulation of Hepatic Progenitor Cells with Both Anti- and Proliver Fibrosis.

    PubMed

    Yang, Ai-Ting; Hu, Dou-Dou; Wang, Ping; Cong, Min; Liu, Tian-Hui; Zhang, Dong; Sun, Ya-Meng; Zhao, Wen-Shan; Jia, Ji-Dong; You, Hong

    2016-01-01

    Transforming growth factor-beta 1 (TGF-β1) plays a central role in hepatic progenitor cells- (HPCs-) mediated liver repair and fibrosis. However, different effects of TGF-β1 on progenitor cells have not been described. In this study, both in vitro (HPCs cocultured with hepatic stellate cells (HSCs) in transwells) and in vivo (CCl4-injured liver fibrosis rat) systems were used to evaluate the impacts. We found that HPCs pretreated with TGF-β1 for 12 hours inhibited the activation of HSCs, while sensitization for 48 hours increased the activation of HSCs. Consistent with these in vitro results, the in vivo fibrosis rat model showed the same time-dependent dual effect of TGF-β1. Regression of liver fibrosis as well as normalization of serum aminotransferase and albumin levels was detected in the rats transplanted with HPCs pretreated with TGF-β1 for 12 hours. In contrast, severe liver fibrosis and elevated collagen-1 levels were detected in the rats transplanted with HPCs pretreated with TGF-β1 for 48 hours. Furthermore, the TGF-β1-pretreated HPCs were shown to deactivate HSCs via enhancing SERPINE1 expression. Inhibition of SERPINE1 reversed the deactivation response in a dose-dependent manner.

  1. Mast cells in renal inflammation and fibrosis: lessons learnt from animal studies.

    PubMed

    Madjene, Lydia Celia; Pons, Maguelonne; Danelli, Luca; Claver, Julien; Ali, Liza; Madera-Salcedo, Iris K; Kassas, Asma; Pellefigues, Christophe; Marquet, Florian; Dadah, Albert; Attout, Tarik; El-Ghoneimi, Alaa; Gautier, Gregory; Benhamou, Marc; Charles, Nicolas; Daugas, Eric; Launay, Pierre; Blank, Ulrich

    2015-01-01

    Mast cells are hematopoietic cells involved in inflammation and immunity and have been recognized also as important effector cells in kidney inflammation. In humans, only a few mast cells reside in kidneys constitutively but in progressive renal diseases their numbers increase substantially representing an essential part of the interstitial infiltrate of inflammatory cells. Recent data obtained in experimental animal models have emphasized a complex role of these cells and the mediators they release as they have been shown both to promote, but also to protect from disease and fibrosis development. Sometimes conflicting results have been reported in similar models suggesting a very narrow window between these activities depending on the pathophysiological context. Interestingly in mice, mast cell or mast cell mediator specific actions became also apparent in the absence of significant mast cell kidney infiltration supporting systemic or regional actions via draining lymph nodes or kidney capsules. Many of their activities rely on the capacity of mast cells to release, in a timely controlled manner, a wide range of inflammatory mediators, which can promote anti-inflammatory actions and repair activities that contribute to healing, but in some circumstances or in case of inappropriate regulation may also promote kidney disease.

  2. Mesenchymal stem cells protect against the tissue fibrosis of ketamine-induced cystitis in rat bladder

    PubMed Central

    Kim, Aram; Yu, Hwan Yeul; Heo, Jinbeom; Song, Miho; Shin, Jung-Hyun; Lim, Jisun; Yoon, Soo-Jung; Kim, YongHwan; Lee, Seungun; Kim, Seong Who; Oh, Wonil; Choi, Soo Jin; Shin, Dong-Myung; Choo, Myung-Soo

    2016-01-01

    Abuse of the hallucinogenic drug ketamine promotes the development of lower urinary tract symptoms that resemble interstitial cystitis. The pathophysiology of ketamine-induced cystitis (KC) is largely unknown and effective therapies are lacking. Here, using a KC rat model, we show the therapeutic effects of human umbilical cord-blood (UCB)-derived mesenchymal stem cells (MSCs). Daily injection of ketamine to Sprague-Dawley rats for 2-weeks resulted in defective bladder function, indicated by irregular voiding frequency, increased maximum contraction pressure, and decreased intercontraction intervals and bladder capacity. KC bladders were characterized by severe mast-cell infiltration, tissue fibrosis, apoptosis, upregulation of transforming growth factor-β signaling related genes, and phosphorylation of Smad2 and Smad3 proteins. A single administration of MSCs (1 × 106) into bladder tissue not only significantly ameliorated the aforementioned bladder voiding parameters, but also reversed the characteristic histological and gene-expression alterations of KC bladder. Treatment with the antifibrotic compound N-acetylcysteine also alleviated the symptoms and pathological characteristics of KC bladder, indicating that the antifibrotic capacity of MSC therapy underlies its benefits. Thus, this study for the first-time shows that MSC therapy might help to cure KC by protecting against tissue fibrosis in a KC animal model and provides a foundation for clinical trials of MSC therapy. PMID:27481042

  3. Early Outgrowth Cells Release Soluble Endocrine Antifibrotic Factors That Reduce Progressive Organ Fibrosis

    PubMed Central

    Yuen, Darren A.; Connelly, Kim A.; Zhang, Yanling; Advani, Suzanne L.; Thai, Kerri; Kabir, Golam; Kepecs, David; Spring, Christopher; Smith, Christopher; Batruch, Ihor; Kosanam, Hari; Advani, Andrew; Diamandis, Eleftherios; Marsden, Philip A.; Gilbert, Richard E.

    2017-01-01

    Adult bone marrow-derived cells can improve organ function in chronic disease models, ostensibly by the release of paracrine factors. It has, however, been difficult to reconcile this prevailing paradigm with the lack of cell retention within injured organs and their rapid migration to the reticuloendothelial system. Here, we provide evidence that the salutary antifibrotic effects of bone marrow-derived early outgrowth cells (EOCs) are more consistent with an endocrine mode of action, demonstrating not only the presence of antifibrotic factors in the plasma of EOC-treated rats but also that EOC conditioned medium (EOC-CM) potently attenuates both TGF-β- and angiotensin II-induced fibroblast collagen production in vitro. To examine the therapeutic relevance of these findings in vivo, 5/6 subtotally nephrectomized rats, a model of chronic kidney and heart failure characterized by progressive fibrosis of both organs, were randomized to receive i.v. injections of EOC-CM, unconditioned medium, or 106 EOCs. Rats that received unconditioned medium developed severe kidney injury with cardiac diastolic dysfunction. In comparison, EOC-CM-treated rats demonstrated substantially improved renal and cardiac function and structure, mimicking the changes found in EOC-treated animals. Mass spectrometric analysis of EOC-CM identified proteins that regulate cellular functions implicated in fibrosis. These results indicate that EOCs secrete soluble factor(s) with highly potent antifibrotic activity, that when injected intravenously replicate the salutary effects of the cells themselves. Together, these findings suggest that an endocrine mode of action may underlie the effectiveness of cell therapy in certain settings and portend the possibility for systemic delivery of cell-free therapy. PMID:23922321

  4. Identification of a Cell-of-Origin for Fibroblasts Comprising the Fibrotic Reticulum in Idiopathic Pulmonary Fibrosis

    PubMed Central

    Xia, Hong; Bodempudi, Vidya; Benyumov, Alexey; Hergert, Polla; Tank, Damien; Herrera, Jeremy; Braziunas, Jeff; Larsson, Ola; Parker, Matthew; Rossi, Daniel; Smith, Karen; Peterson, Mark; Limper, Andrew; Jessurun, Jose; Connett, John; Ingbar, David; Phan, Sem; Bitterman, Peter B.; Henke, Craig A.

    2015-01-01

    Idiopathic pulmonary fibrosis (IPF) is a progressive disease of the middle aged and elderly with a prevalence of one million persons worldwide. The fibrosis spreads from affected alveoli into contiguous alveoli, creating a reticular network that leads to death by asphyxiation. Lung fibroblasts from patients with IPF have phenotypic hallmarks, distinguishing them from their normal counterparts: pathologically activated Akt signaling axis, increased collagen and α-smooth muscle actin expression, distinct gene expression profile, and ability to form fibrotic lesions in model organisms. Despite the centrality of these fibroblasts in disease pathogenesis, their origin remains uncertain. Here, we report the identification of cells in the lungs of patients with IPF with the properties of mesenchymal progenitors. In contrast to progenitors isolated from nonfibrotic lungs, IPF mesenchymal progenitor cells produce daughter cells manifesting the full spectrum of IPF hallmarks, including the ability to form fibrotic lesions in zebrafish embryos and mouse lungs, and a transcriptional profile reflecting these properties. Morphological analysis of IPF lung tissue revealed that mesenchymal progenitor cells and cells with the characteristics of their progeny comprised the fibrotic reticulum. These data establish that the lungs of patients with IPF contain pathological mesenchymal progenitor cells that are cells of origin for fibrosis-mediating fibroblasts. These fibrogenic mesenchymal progenitors and their progeny represent an unexplored target for novel therapies to interdict fibrosis. PMID:24631025

  5. Cystic fibrosis transmembrane conductance regulator protein expression rate in healthy spermatozoa is not correlated with ovum fertilisation rate.

    PubMed

    Li, H-G; Xu, C-M; Chen, W-Y; Shi, Q-X; Ni, Y

    2012-05-01

    Our previous studies have shown that the cystic fibrosis transmembrane conductance regulator (CFTR) was important for capacitation and fertilisation in mouse, guinea pig and human spermatozoa. However, it is unclear whether CFTR is correlated with ovum fertilisation rate. The present study was to test the possible relationship between spermatozoa CFTR protein expression rate in healthy men and ovum fertilisation rate during in vitro fertilisation. Ninety-four couples for female factor infertility for IVF-ET treatments were retrospectively studied. All the patients were divided into three groups based on the fertilisation rate of ovum in vitro. It was performed to explore whether there were differences in sperm CFTR protein expression rate among the three groups and the relevance between CFTR protein expression rate and ovum fertilisation rate. Our study showed that there was no significant differences in sperm CFTR protein expression rate among the three groups (F = 0.614, P = 0.544), and the relevance between spermatozoa CFTR protein expression rate and ovum fertilisation rate was not significantly different (r = 0.013, P = 0.904). These results further suggest that CFTR protein expression rate in healthy men spermatozoa was not associated with ovum fertilisation rate and thus we cannot predict ovum fertilisation results by sperm CFTR protein expression rate.

  6. Increased tubulointerstitial recruitment of human CD141(hi) CLEC9A(+) and CD1c(+) myeloid dendritic cell subsets in renal fibrosis and chronic kidney disease.

    PubMed

    Kassianos, Andrew J; Wang, Xiangju; Sampangi, Sandeep; Muczynski, Kimberly; Healy, Helen; Wilkinson, Ray

    2013-11-15

    Dendritic cells (DCs) play critical roles in immune-mediated kidney diseases. Little is known, however, about DC subsets in human chronic kidney disease, with previous studies restricted to a limited set of pathologies and to using immunohistochemical methods. In this study, we developed novel protocols for extracting renal DC subsets from diseased human kidneys and identified, enumerated, and phenotyped them by multicolor flow cytometry. We detected significantly greater numbers of total DCs as well as CD141(hi) and CD1c(+) myeloid DC (mDCs) subsets in diseased biopsies with interstitial fibrosis than diseased biopsies without fibrosis or healthy kidney tissue. In contrast, plasmacytoid DC numbers were significantly higher in the fibrotic group compared with healthy tissue only. Numbers of all DC subsets correlated with loss of kidney function, recorded as estimated glomerular filtration rate. CD141(hi) DCs expressed C-type lectin domain family 9 member A (CLEC9A), whereas the majority of CD1c(+) DCs lacked the expression of CD1a and DC-specific ICAM-3-grabbing nonintegrin (DC-SIGN), suggesting these mDC subsets may be circulating CD141(hi) and CD1c(+) blood DCs infiltrating kidney tissue. Our analysis revealed CLEC9A(+) and CD1c(+) cells were restricted to the tubulointerstitium. Notably, DC expression of the costimulatory and maturation molecule CD86 was significantly increased in both diseased cohorts compared with healthy tissue. Transforming growth factor-β levels in dissociated tissue supernatants were significantly elevated in diseased biopsies with fibrosis compared with nonfibrotic biopsies, with mDCs identified as a major source of this profibrotic cytokine. Collectively, our data indicate that activated mDC subsets, likely recruited into the tubulointerstitium, are positioned to play a role in the development of fibrosis and, thus, progression to chronic kidney disease.

  7. Antisense oligodeoxynucleotide to the cystic fibrosis gene inhibits anion transport in normal cultured sweat duct cells

    SciTech Connect

    Sorscher, E.J.; Kirk, K.L.; Weaver, M.L.; Jilling, T.; Blalock, J.E.; LeBoeuf, R.D. )

    1991-09-01

    The authors have tested the hypothesis that the cystic fibrosis (CF) gene product, called the CF transmembrane conductance regulator (CFTR), mediates anion transport in normal human sweat duct cells. Sweat duct cells in primary culture were treated with oligodeoxynucleotides that were antisense to the CFTR gene transcript in order to block the expression of the wild-type CFTR. Anion transport in CFTR transcript antisense-treated cells was then assessed with a halide-specific dye, 6-methoxy-N-(3-sulfopropryl)quinolinium, and fluorescent digital imaging microscopy to monitor halide influx and efflux from single sweat duct cells. Antisense oligodeoxynucleotide treatment for 24 hr virtually abolished Cl{sup {minus}} transport in sweat duct cells compared with untreated cells or control cells treated with sense oligodeoxynucleotides. Br{sup {minus}} uptake into sweat duct cells was also blocked after a 24-hr CFTR transcript antisense treatments, but not after treatments for only 4 hr. Lower concentrations of antisense oligodeoxynucleotides were less effective at inhibiting Cl{sup {minus}} transport. These results indicate that oligodeoxynucleotides that are antisense to CFTR transcript inhibit sweat duct Cl{sup {minus}} permeability in both a time-dependent and dose-dependent manner. This approach provides evidence that inhibition of the expression of the wild-type CFTR gene in a normal, untransfected epithelial cell results in an inhibition of Cl{sup {minus}} permeability.

  8. Streptomycin treatment alters the intestinal microbiome, pulmonary T cell profile and airway hyperresponsiveness in a cystic fibrosis mouse model.

    PubMed

    Bazett, Mark; Bergeron, Marie-Eve; Haston, Christina K

    2016-01-12

    Cystic fibrosis transmembrane conductance regulator deficient mouse models develop phenotypes of relevance to clinical cystic fibrosis (CF) including airway hyperresponsiveness, small intestinal bacterial overgrowth and an altered intestinal microbiome. As dysbiosis of the intestinal microbiota has been recognized as an important contributor to many systemic diseases, herein we investigated whether altering the intestinal microbiome of BALB/c Cftr(tm1UNC) mice and wild-type littermates, through treatment with the antibiotic streptomycin, affects the CF lung, intestinal and bone disease. We demonstrate that streptomycin treatment reduced the intestinal bacterial overgrowth in Cftr(tm1UNC) mice and altered the intestinal microbiome similarly in Cftr(tm1UNC) and wild-type mice, principally by affecting Lactobacillus levels. Airway hyperresponsiveness of Cftr(tm1UNC) mice was ameliorated with streptomycin, and correlated with Lactobacillus abundance in the intestine. Additionally, streptomycin treated Cftr(tm1UNC) and wild-type mice displayed an increased percentage of pulmonary and mesenteric lymph node Th17, CD8 + IL-17+ and CD8 + IFNγ+ lymphocytes, while the CF-specific increase in respiratory IL-17 producing γδ T cells was decreased in streptomycin treated Cftr(tm1UNC) mice. Bone disease and intestinal phenotypes were not affected by streptomycin treatment. The airway hyperresponsiveness and lymphocyte profile of BALB/c Cftr(tm1UNC) mice were affected by streptomycin treatment, revealing a potential intestinal microbiome influence on lung response in BALB/c Cftr(tm1UNC) mice.

  9. Contribution and Mobilization of Mesenchymal Stem Cells in a mouse model of carbon tetrachloride-induced liver fibrosis.

    PubMed

    Liu, Yan; Yang, Xue; Jing, Yingying; Zhang, Shanshan; Zong, Chen; Jiang, Jinghua; Sun, Kai; Li, Rong; Gao, Lu; Zhao, Xue; Wu, Dong; Shi, Yufang; Han, Zhipeng; Wei, Lixin

    2015-12-08

    Hepatic fibrosis is associated with bone marrow derived mesenchymal stem cells (BM-MSCs). In this study, we aimed to determine what role MSCs play in the process and how they mobilize from bone marrow (BM). We employed a mouse model of carbon tetrachloride(CCl4)-induced liver fibrosis. Frozen section was used to detect MSCs recruited to mice and human fibrotic liver. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) was detected to assess liver function. It was found that MSCs of both exogenous and endogenous origin could aggravate liver fibrosis and attenuate liver damage as indicated by lower serum ALT and AST levels. Stromal cell-derived factor-1 (SDF-1α)/ CXCR4 was the most important chemotactic axis regulating MSCs migration from BM to fibrotic liver. Frozen section results showed that the migration did not start from the beginning of liver injury but occurred when the expression balance of SDF-1α between liver and BM was disrupted, where SDF-1α expression in liver was higher than that in BM. Our findings provide further evidence to show the role of BM-MSCs in liver fibrosis and to elucidate the mechanism underlying MSCs mobilization in our early liver fibrosis mice model induced by CCl4.

  10. Bone marrow mesenchymal stem cells protect against bleomycin-induced pulmonary fibrosis in rat by activating Nrf2 signaling

    PubMed Central

    Ni, Shirong; Wang, Dexuan; Qiu, Xiaoxiao; Pang, Lingxia; Song, Zhangjuan; Guo, Kunyuan

    2015-01-01

    Pulmonary fibrosis is a progressive and lethal disorder. Although the precise mechanisms of pulmonary fibrosis are not fully understood, oxidant/antioxidant may play an important role in many of the processes of inflammation and fibrosis. Keap1-Nrf2-ARE pathway represents one of the most important cellular defense mechanisms against oxidative stress. Mesenchymal stem cells (MSC) are in clinical trials for widespread indications including musculoskeletal, neurological, cardiac and haematological disorders. One emerging concept is that MSCs may have paracrine, rather than a functional, roles in lung injury repair and regeneration. In the present study, we investigated bone marrow mesenchymal stem cells (BMSCs) for the treatment of bleomycin-induced pulmonary fibrosis. Our results showed that BMSCs administration significantly ameliorated the bleomycin mediated histological alterations and blocked collagen deposition with parallel reduction in the hydroxyproline level. The gene expression levels of NAD(P)H: quinine oxidoreductase 1 (NQO1), gama-glutamylcysteine synthetase (γ-GCS), heme oxygenase-1 (HO-1) and nuclear factor erythroid 2-related factor 2 (Nrf2), attenuated by bleomycin, were increased up to basal levels after BMSCs transplantation. BMSCs significantly increased superoxide dismutase (SOD) activity and inhibited malondialdehyde (MDA) production in the injured lung. The present study provides evidence that BMSCs may be a potential therapeutic reagent for the treatment of lung fibrosis. PMID:26339340

  11. Tissue culture of normal and cystic fibrosis sweat gland duct cells. I. Alterations in dome formation.

    PubMed

    Hazen-Martin, D J; Spicer, S S; Sens, M A; Jenkins, M Q; Westphal, M C; Sens, D A

    1987-01-01

    The elucidation of the underlying defect in fluid secretion by cystic fibrosis (CF) sweat glands is hindered by the unavailability of an experimental model for investigating this disease. As a potential model system, a serum-free growth medium was developed that supports the explant growth of epithelial cells from fragments of human skin. Immunohistochemical analysis demonstrated that these epithelial cell outgrowths originated from the duct of the sweat gland. By electron microscopy, the cells were demonstrated to possess keratinocyte-like morphology as noted by the presence of a multilayered outgrowth of cells containing well-defined keratin bundles. Identical outgrowths from skin biopsies of CF patients were compared to normal outgrowths and alterations were noted to occur in dome formation and in the number of intercellular spaces between cells. Doming alterations were also noted to occur in the CF heterozygous state. No differences in cell fine structure or in growth factor requirements for cell proliferation were noted between normal and CF cells. The potential use of this system as a model for CF research is discussed.

  12. Developmental Reprogramming in Mesenchymal Stromal Cells of Human Subjects with Idiopathic Pulmonary Fibrosis

    PubMed Central

    Chanda, Diptiman; Kurundkar, Ashish; Rangarajan, Sunad; Locy, Morgan; Bernard, Karen; Sharma, Nirmal S.; Logsdon, Naomi J.; Liu, Hui; Crossman, David K.; Horowitz, Jeffrey C.; De Langhe, Stijn; Thannickal, Victor J.

    2016-01-01

    Cellular plasticity and de-differentiation are hallmarks of tissue/organ regenerative capacity in diverse species. Despite a more restricted capacity for regeneration, humans with age-related chronic diseases, such as cancer and fibrosis, show evidence of a recapitulation of developmental gene programs. We have previously identified a resident population of mesenchymal stromal cells (MSCs) in the terminal airways-alveoli by bronchoalveolar lavage (BAL) of human adult lungs. In this study, we characterized MSCs from BAL of patients with stable and progressive idiopathic pulmonary fibrosis (IPF), defined as <5% and ≥10% decline, respectively, in forced vital capacity over the preceding 6-month period. Gene expression profiles of MSCs from IPF subjects with progressive disease were enriched for genes regulating lung development. Most notably, genes regulating early tissue patterning and branching morphogenesis were differentially regulated. Network interactive modeling of a set of these genes indicated central roles for TGF-β and SHH signaling. Importantly, fibroblast growth factor-10 (FGF-10) was markedly suppressed in IPF subjects with progressive disease, and both TGF-β1 and SHH signaling were identified as critical mediators of this effect in MSCs. These findings support the concept of developmental gene re-activation in IPF, and FGF-10 deficiency as a potentially critical factor in disease progression. PMID:27869174

  13. Detection of KI WU and Merkel cell polyomavirus in respiratory tract of cystic fibrosis patients.

    PubMed

    Iaria, M; Caccuri, F; Apostoli, P; Giagulli, C; Pelucchi, F; Padoan, R F; Caruso, A; Fiorentini, S

    2015-06-01

    In the last few years, many reports have confirmed the presence of WU, KI and Merkel cell (MC) polyomaviruses (PyV) in respiratory samples wordwide, but their pathogenic role in patients with underlying conditions such as cystic fibrosis is still debated. To determine the prevalence of MCPyV, WUPyV and KIPyV, we conducted a 1-year-long microbiological testing of respiratory specimens from 93 patients with cystic fibrosis in Brescia, Italy. We detected PyV DNA in 94 out of 337 analysed specimens. KIPyV was the most common virus detected (12.1%), followed by WUPyV (8.9%) and MCPyV (6.8%). We found an intriguing association between the presence of MCPyV and the concurrent isolation of Pseudomonas aeruginosa, as well as with the patient status, classified as chronically colonized with P. aeruginosa. Our study adds perspective on the prevalence and the potential pathogenic role of PyV infections.

  14. Quantitative computed tomography analysis of the airways in patients with cystic fibrosis using automated software: correlation with spirometry in the evaluation of severity*

    PubMed Central

    Santos, Marcel Koenigkam; Cruvinel, Danilo Lemos; de Menezes, Marcelo Bezerra; Teixeira, Sara Reis; Vianna, Elcio de Oliveira; Elias Júnior, Jorge; Martinez, José Antonio Baddini

    2016-01-01

    Objective To perform a quantitative analysis of the airways using automated software, in computed tomography images of patients with cystic fibrosis, correlating the results with spirometric findings. Materials and Methods Thirty-four patients with cystic fibrosis were studied-20 males and 14 females; mean age 18 ± 9 years-divided into two groups according to the spirometry findings: group I (n = 21), without severe airflow obstruction (forced expiratory volume in first second [FEV1] > 50% predicted), and group II (n = 13), with severe obstruction (FEV1 ≤ 50% predicted). The following tracheobronchial tree parameters were obtained automatically: bronchial diameter, area, thickness, and wall attenuation. Results On average, 52 bronchi per patient were studied. The number of bronchi analyzed was higher in group II. The correlation with spirometry findings, especially between the relative wall thickness of third to eighth bronchial generation and predicted FEV1, was better in group I. Conclusion Quantitative analysis of the airways by computed tomography can be useful for assessing disease severity in cystic fibrosis patients. In patients with severe airflow obstruction, the number of bronchi studied by the method is higher, indicating more bronchiectasis. In patients without severe obstruction, the relative bronchial wall thickness showed a good correlation with the predicted FEV1. PMID:28100929

  15. Iron overload correlates with serum liver fibrotic markers and liver dysfunction: Potential new methods to predict iron overload-related liver fibrosis in thalassemia patients

    PubMed Central

    Wang, Man; Liu, Rongrong; Liang, Yuzhen; Yang, Gaohui; Huang, Yumei; Yu, Chunlan; Sun, Kaiqi; Xia, Yang

    2016-01-01

    Background Early detection of liver fibrosis in thalassemia patients and rapid initiation of treatment to interfere with its progression are extremely important. Objective This study aimed to find a sensitive, easy-to-detect and noninvasive method other than liver biopsy for early detection of liver fibrosis in thalassemia patients. Methods A total of 244 Chinese Thalassemia patients with non-transfusion-dependent thalassemia (NTDT, n = 105) or thalassemia major (TM, n = 139) and 120 healthy individuals were recruited into the present study, and blood collagen type IV (C IV), precollagen type III (PIIINPC) and hyaluronic acid (HA), aspartate aminotransferase (AST), alanine aminotransferase (ALT) and ferritin were measured. Liver iron concentration was determined by MRI. The correlation of serum markers with liver iron load and liver function was evaluated. Results Serum C IV, PIIINPC and HA were significantly elevated in Chinese patients with NTDT and further elevated in TM patients. Moreover, C IV, PIIINPC and HA were also positively correlated to serum ferritin and liver iron concentration and further elevated during the progression to multi-organ damage in NTDT patients. Finally, serum ferritin and liver iron concentration were significantly correlated with liver dysfunction determined by AST and ALT. Conclusion Taken together, our results indicate that monitoring serum C IV, PIIINPC and HA is a potentially sensitive method to predict the risks for iron overload-related liver fibrosis in Chinese thalassemia patients.

  16. Flagellin induces myeloid-derived suppressor cells: implications for Pseudomonas aeruginosa infection in cystic fibrosis lung disease.

    PubMed

    Rieber, Nikolaus; Brand, Alina; Hector, Andreas; Graepler-Mainka, Ute; Ost, Michael; Schäfer, Iris; Wecker, Irene; Neri, Davide; Wirth, Andreas; Mays, Lauren; Zundel, Sabine; Fuchs, Jörg; Handgretinger, Rupert; Stern, Martin; Hogardt, Michael; Döring, Gerd; Riethmüller, Joachim; Kormann, Michael; Hartl, Dominik

    2013-02-01

    Pseudomonas aeruginosa persists in patients with cystic fibrosis (CF) and drives CF lung disease progression. P. aeruginosa potently activates the innate immune system, mainly mediated through pathogen-associated molecular patterns, such as flagellin. However, the host is unable to eradicate this flagellated bacterium efficiently. The underlying immunological mechanisms are incompletely understood. Myeloid-derived suppressor cells (MDSCs) are innate immune cells generated in cancer and proinflammatory microenvironments and are capable of suppressing T cell responses. We hypothesized that P. aeruginosa induces MDSCs to escape T cell immunity. In this article, we demonstrate that granulocytic MDSCs accumulate in CF patients chronically infected with P. aeruginosa and correlate with CF lung disease activity. Flagellated P. aeruginosa culture supernatants induced the generation of MDSCs, an effect that was 1) dose-dependently mimicked by purified flagellin protein, 2) significantly reduced using flagellin-deficient P. aeruginosa bacteria, and 3) corresponded to TLR5 expression on MDSCs in vitro and in vivo. Both purified flagellin and flagellated P. aeruginosa induced an MDSC phenotype distinct from that of the previously described MDSC-inducing cytokine GM-CSF, characterized by an upregulation of the chemokine receptor CXCR4 on the surface of MDSCs. Functionally, P. aeruginosa-infected CF patient ex vivo-isolated as well as flagellin or P. aeruginosa in vitro-generated MDSCs efficiently suppressed polyclonal T cell proliferation in a dose-dependent manner and modulated Th17 responses. These studies demonstrate that flagellin induces the generation of MDSCs and suggest that P. aeruginosa uses this mechanism to undermine T cell-mediated host defense in CF and other P. aeruginosa-associated chronic lung diseases.

  17. Effect of taurine on acinar cell apoptosis and pancreatic fibrosis in dibutyltin dichloride-induced chronic pancreatitis.

    PubMed

    Matsushita, Koki; Mizushima, Takaaki; Shirahige, Akinori; Tanioka, Hiroaki; Sawa, Kiminari; Ochi, Koji; Tanimoto, Mitsune; Koide, Norio

    2012-01-01

    The relationship between pancreatic fibrosis and apoptosis of pancreatic acinar cells has not been fully elucidated. We reported that taurine had an anti-fibrotic effect in a dibutyltin dichloride (DBTC)-chronic pancreatitis model. However, the effect of taurine on apoptosis of pancreatic acinar cells is still unclear. Therefore, we examined apoptosis in DBTC-chronic pancreatitis and in the AR42J pancreatic acinar cell line with/without taurine. Pancreatic fibrosis was induced by a single administration of DBTC. Rats were fed a taurine-containing diet or a normal diet and were sacrificed at day 5. The AR42J pancreatic acinar cell line was incubated with/without DBTC with taurine chloramines. Apoptosis was determined by using terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) assay. The expression of Bad and Bcl-2 proteins in the AR42J cells lysates was detected by Western blot analysis. The apoptotic index of pancreatic acinar cells in DBTC-administered rats was significantly increased. Taurine treatment inhibited pancreatic fibrosis and apoptosis of acinar cells induced by DBTC. The number of TUNEL-positive cells in the AR42J pancreatic acinar cell lines was significantly increased by the addition of DBTC. Incubation with taurine chloramines ameliorated these changes. In conclusion, taurine inhibits apoptosis of pancreatic acinar cells and pancreatitis in experimental chronic pancreatitis.

  18. Oncoprotein mdig contributes to silica-induced pulmonary fibrosis by altering balance between Th17 and Treg T cells

    PubMed Central

    Sun, Jiaying; Zhang, Yadong; Lu, Yongju; Battelli, Lori; Porter, Dale W.; Chen, Fei

    2015-01-01

    Mineral dust-induced gene (mdig, also named Mina53) was first identified from alveolar macrophages of the coal miners with chronic lung inflammation or fibrosis, but how this gene is involved in lung diseases is poorly understood. Here we show that heterozygotic knockout of mdig (mdig+/−) ameliorates silica-induced lung fibrosis by altering the balance between Th17 cells and Treg cells. Relative to the wild type (WT) mice, infiltration of the macrophages and Th17 cells was reduced in lungs from silica-exposed mdig+/− mice. In contrast, an increased infiltration of the T regulatory (Treg) cells to the lung intestitium was observed in the mdig+/− mice treated with silica. Both the number of Th17 cells in the lung lymph nodes and the level of IL-17 in the bronchoalveolar lavage fluids were decreased in the mdig+/− mice in response to silica. Thus, these results suggest that mdig may contribute to silica-induced lung fibrosis by altering the balance between Th17 and Treg cells. Genetic deficiency of mdig impairs Th17 cell infiltration and function, but favors infiltration of the Treg cells, the immune suppressive T cells that are able to limit the inflammatory responses by repressing the Th17 cells and macrophages. PMID:25669985

  19. Targeting dexamethasone to Kupffer cells: effects on liver inflammation and fibrosis in rats.

    PubMed

    Melgert, B N; Olinga, P; Van Der Laan, J M; Weert, B; Cho, J; Schuppan, D; Groothuis, G M; Meijer, D K; Poelstra, K

    2001-10-01

    Kupffer cells (KC) play an important role in the pathogenesis of inflammatory liver diseases leading to fibrosis. Anti-inflammatory drugs are only effective when administered at high doses that may cause side effects. Therefore, dexamethasone coupled to mannosylated albumin (Dexa(5)-Man(10)-HSA) was designed by us to selectively deliver this anti-inflammatory drug to the KC. The effectiveness of Dexa(5)-Man(10)-HSA was studied both in organ cultures and fibrosis induced by bile duct ligation (BDL) in rats. Dexa(5)-Man(10)-HSA accumulated in livers of both healthy and fibrotic rats (67% +/- 5% and 70% +/- 9% of the dose, respectively) and uptake was found almost exclusively in KC. Active dexamethasone was liberated from its carrier, because Dexa(5)-Man(10)-HSA could effectively inhibit nitric oxide (NO) and tumor necrosis factor alpha (TNF-alpha) release in endotoxin-activated liver slices. In vivo, however, this was associated with increased collagen I and III depositions and enhanced tissue inhibitor of metalloproteinase-1 (TIMP-1) mRNA expression. This was accompanied by a decreased influx of reactive oxygen species (ROS) producing cells in the livers of BDL animals treated with Dexa(5)-Man(10)-HSA as compared with untreated BDL rats. Dexa(5)-Man(10)-HSA treatment also replenished the depleted glycogen stores in hepatocytes of BDL livers. In conclusion, our studies showed selective delivery of dexamethasone to KC with Dexa(5)-Man(10)-HSA. This conjugate reduced intrahepatic ROS in vivo and TNF-alpha production in vitro and prevented glycogen depletion in vivo, indicating effective pharmacologic targeting. Dexa(5)-Man(10)-HSA, however, also accelerated fibrogenesis, which was paralleled by TIMP-1 mRNA induction. Targeting of dexamethasone to KC provides evidence for a dual role of this cell type in fibrogenesis of BDL rats.

  20. Upregulation of Steroidogenic Acute Regulatory Protein by Hypoxia Stimulates Aldosterone Synthesis in Pulmonary Artery Endothelial Cells to Promote Pulmonary Vascular Fibrosis

    PubMed Central

    Maron, Bradley A.; Oldham, William M.; Chan, Stephen Y.; Vargas, Sara O.; Arons, Elena; Zhang, Ying-Yi; Loscalzo, Joseph; Leopold, Jane A.

    2014-01-01

    Background The molecular mechanism(s) regulating hypoxia-induced vascular fibrosis are unresolved. Hyperaldosteronism correlates positively with vascular remodeling in pulmonary arterial hypertension (PAH), suggesting that aldosterone may contribute to the pulmonary vasculopathy of hypoxia. The hypoxia-sensitive transcription factors c-Fos/c-Jun regulate steroidogenic acute regulatory protein (StAR), which facilitates the rate-limiting step of aldosterone steroidogenesis. We hypothesized that c-Fos/c-Jun upregulation by hypoxia activates StAR-dependent aldosterone synthesis in human pulmonary artery endothelial cells (HPAECs) to promote vascular fibrosis in PAH. Methods and Results Patients with PAH, rats with Sugen/hypoxia-PAH, and mice exposed to chronic hypoxia expressed increased StAR in remodeled pulmonary arterioles, providing a basis for investigating hypoxia-StAR signaling in HPAECs. Hypoxia (2.0% FiO2) increased aldosterone levels selectively in HPAECs, which was confirmed by liquid chromatography-mass spectrometry. Increased aldosterone by hypoxia resulted from enhanced c-Fos/c-Jun binding to the proximal activator protein (AP-1) site of the StAR promoter in HPAECs, which increased StAR expression and activity. In HPAECs transfected with StAR-siRNA or treated with the AP-1 inhibitor, SR-11302, hypoxia failed to increase aldosterone, confirming that aldosterone biosynthesis required StAR activation by c-Fos/c-Jun. The functional consequences of aldosterone were confirmed by pharmacological inhibition of the mineralocorticoid receptor with spironolactone or eplerenone, which attenuated hypoxia-induced upregulation of the fibrogenic protein connective tissue growth factor and collagen III in vitro, and decreased pulmonary vascular fibrosis to improve pulmonary hypertension in Conclusions Our findings identify autonomous aldosterone synthesis in HPAECs due to hypoxia-mediated upregulation of StAR as a novel molecular mechanism that promotes pulmonary vascular

  1. Contribution and Mobilization of Mesenchymal Stem Cells in a mouse model of carbon tetrachloride-induced liver fibrosis

    PubMed Central

    Liu, Yan; Yang, Xue; Jing, Yingying; Zhang, Shanshan; Zong, Chen; Jiang, Jinghua; Sun, Kai; Li, Rong; Gao, Lu; Zhao, Xue; Wu, Dong; Shi, Yufang; Han, Zhipeng; Wei, Lixin

    2015-01-01

    Hepatic fibrosis is associated with bone marrow derived mesenchymal stem cells (BM-MSCs). In this study, we aimed to determine what role MSCs play in the process and how they mobilize from bone marrow (BM). We employed a mouse model of carbon tetrachloride(CCl4)-induced liver fibrosis. Frozen section was used to detect MSCs recruited to mice and human fibrotic liver. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) was detected to assess liver function. It was found that MSCs of both exogenous and endogenous origin could aggravate liver fibrosis and attenuate liver damage as indicated by lower serum ALT and AST levels. Stromal cell–derived factor-1 (SDF-1α)/ CXCR4 was the most important chemotactic axis regulating MSCs migration from BM to fibrotic liver. Frozen section results showed that the migration did not start from the beginning of liver injury but occured when the expression balance of SDF-1α between liver and BM was disrupted, where SDF-1α expression in liver was higher than that in BM. Our findings provide further evidence to show the role of BM-MSCs in liver fibrosis and to elucidate the mechanism underlying MSCs mobilization in our early liver fibrosis mice model induced by CCl4. PMID:26643997

  2. Modified citrus pectin stops progression of liver fibrosis by inhibiting galectin-3 and inducing apoptosis of stellate cells.

    PubMed

    Abu-Elsaad, Nashwa M; Elkashef, Wagdi Fawzi

    2016-05-01

    Modified citrus pectin (MCP) is a pH modified form of the dietary soluble citrus peel fiber known as pectin. The current study aims at testing its effect on liver fibrosis progression. Rats were injected with CCl4 (1 mL/kg, 40% v/v, i.p., twice a week for 8 weeks). Concurrently, MCP (400 or 1200 mg/kg) was administered daily in drinking water from the first week in groups I and II (prophylactic model) and in the beginning of week 5 in groups III and IV (therapeutic model). Liver function biomarkers (ATL, AST, and ALP), fibrosis markers (laminin and hyaluronic acid), and antioxidant biomarkers (reduced glutathione (GSH) and superoxide dismutase (SOD)) were measured. Stained liver sections were scored for fibrosis and necroinflammation. Additionally, expression of galectin-3 (Gal-3), α-smooth muscle actin (SMA), tissue inhibitor metalloproteinase (TIMP)-1, collagen (Col)1A1, caspase (Cas)-3, and apoptosis related factor (FAS) were assigned. Modified pectin late administration significantly (p < 0.05) decreased malondialdehyde (MDA), TIMP-1, Col1A1, α-SMA, and Gal-3 levels and increased levels of FAS, Cas-3, GSH, and SOD. It also decreased percentage of fibrosis and necroinflammation significantly (p < 0.05). It can be concluded that MCP can attenuate liver fibrosis through an antioxidant effect, inhibition of Gal-3 mediated hepatic stellate cells activation, and induction of apoptosis.

  3. Dysfunction of mitochondria Ca2+ uptake in cystic fibrosis airway epithelial cells.

    PubMed

    Antigny, Fabrice; Girardin, Nathalie; Raveau, Dorothée; Frieden, Maud; Becq, Frédéric; Vandebrouck, Clarisse

    2009-07-01

    In the genetic disease cystic fibrosis (CF), the most common mutation F508del promotes the endoplasmic reticulum (ER) retention of misfolded CF proteins. Furthermore, in homozygous F508del-CFTR airway epithelial cells, the histamine Ca(2+) mobilization is abnormally increased. Because the uptake of Ca(2+) by mitochondria during Ca(2+) influx or Ca(2+) release from ER stores may be crucial for maintaining a normal Ca(2+) homeostasis, we compared the mitochondria morphology and distribution by transmission electron microscopy technique and the mitochondria membrane potential variation (DeltaPsi(mit)) using a fluorescent probe (TMRE) on human CF (CF-KM4) and non-CF (MM39) tracheal serous gland cell lines. Confocal imaging of Rhod-2-AM-loaded or of the mitochondrial targeted cameleon 4mtD3cpv-transfected human CF and non-CF cells, were used to examine the ability of mitochondria to sequester intracellular Ca(2+). The present study reveals that (i) the mitochondria network is fragmented in F508del-CFTR cells, (ii) the DeltaPsi(mit) of CF mitochondria is depolarized compared non-CF mitochondria, and (iii) the CF mitochondria Ca(2+) uptake is reduced compared non-CF cells. We propose that these defects in airway epithelial F508del-CFTR cells are the consequence of mitochondrial membrane depolarization leading to a deficient mitochondrial Ca(2+) uptake.

  4. Increased sulfation of glycoconjugates by cultured nasal epithelial cells from patients with cystic fibrosis.

    PubMed Central

    Cheng, P W; Boat, T F; Cranfill, K; Yankaskas, J R; Boucher, R C

    1989-01-01

    Cystic fibrosis (CF) respiratory epithelia exhibit abnormal anion transport that may be linked to abnormal lung defense. In these studies, we investigated whether primary cultures of CF respiratory epithelial cells regulate abnormally the sulfate content of high molecular weight glycoconjugates (HMG) participating in airways' mucosal defense. HMG, including glycosaminoglycans and mucin-type glycoproteins released spontaneously into medium and HMG released from cell surfaces by trypsin, were metabolically labeled with 35SO4- and [6-3H]-glucosamine (GlcN) or 35SO4- and [3H]serine. All three classes of HMG from CF cells exhibited 35S/3H labeling ratios 1.5-4-fold greater than HMG from normal or disease control cells. Differences for labeling ratios of HMG from CF cells were shown to be the consequence of increased 35SO4- incorporation rather than decreased peptide synthesis and release or HMG glycosylation. The buoyant density of CF mucin-type HMG also was increased, consistent with increased sulfation. These observations suggest that oversulfation of a spectrum of HMG is a genetically determined characteristic of CF epithelial cells and may play an important pathophysiological role by altering the properties of mucous secretions and/or the interactions between selected bacteria and HMG at the airways' surface. Images PMID:2738159

  5. Genetic ablation of mast cells redefines the role of mast cells in skin wound healing and bleomycin-induced fibrosis.

    PubMed

    Willenborg, Sebastian; Eckes, Beate; Brinckmann, Jürgen; Krieg, Thomas; Waisman, Ari; Hartmann, Karin; Roers, Axel; Eming, Sabine A

    2014-07-01

    Conclusive evidence for the impact of mast cells (MCs) in skin repair is still lacking. Studies in mice examining the role of MC function in the physiology and pathology of skin regenerative processes have obtained contradictory results. To clarify the specific role of MCs in regenerative conditions, here we used a recently developed genetic mouse model that allows conditional MC ablation to examine MC-specific functions in skin. This mouse model is based on the cell type-specific expression of Cre recombinase in connective tissue-type MCs under control of the Mcpt5 promoter and the Cre-inducible diphtheria toxin receptor-mediated cell lineage ablation by diphtheria toxin. In response to excisional skin injury, genetic ablation of MCs did not affect the kinetics of reepithelialization, the formation of vascularized granulation tissue, or scar formation. Furthermore, genetic ablation of MCs failed to prevent the development of skin fibrosis upon bleomycin challenge. The amount of deposited collagen and the biochemistry of collagen fibril crosslinks within fibrotic lesions were comparable in MC-depleted and control mice. Collectively, our findings strongly suggest that significant reduction of MC numbers does not affect skin wound healing and bleomycin-induced fibrosis in mice, and provide to our knowledge previously unreported insight in the long-debated contribution of MCs in skin regenerative processes.

  6. Oxidative stress and hepatic stellate cell activation are key events in arsenic induced liver fibrosis in mice

    SciTech Connect

    Ghatak, Subhadip; Biswas, Ayan; Dhali, Gopal Krishna; Chowdhury, Abhijit; Boyer, James L.; Santra, Amal

    2011-02-15

    Arsenic is an environmental toxicant and carcinogen. Exposure to arsenic is associated with development of liver fibrosis and portal hypertension through ill defined mechanisms. We evaluated hepatic fibrogenesis after long term arsenic exposure in a murine model. BALB/c mice were exposed to arsenic by daily gavages of 6 {mu}g/gm body weight for 1 year and were evaluated for markers of hepatic oxidative stress and fibrosis, as well as pro-inflammatory, pro-apoptotic and pro-fibrogenic factors at 9 and 12 months. Hepatic NADPH oxidase activity progressively increased in arsenic exposure with concomitant development of hepatic oxidative stress. Hepatic steatosis with occasional collection of mononuclear inflammatory cells and mild portal fibrosis were the predominant liver lesion observed after 9 months of arsenic exposure, while at 12 months, the changes included mild hepatic steatosis, inflammation, necrosis and significant fibrosis in periportal areas. The pathologic changes in the liver were associated with markers of hepatic stellate cells (HSCs) activation, matrix reorganization and fibrosis including {alpha}-smooth muscle actin, transforming growth factor-{beta}1, PDGF-R{beta}, pro-inflammatory cytokines and enhanced expression of tissue inhibitor of metalloproteinase-1 and pro({alpha}) collagen type I. Moreover, pro-apoptotic protein Bax was dominantly expressed and Bcl-2 was down-regulated along with increased number of TUNEL positive hepatocytes in liver of arsenic exposed mice. Furthermore, HSCs activation due to increased hepatic oxidative stress observed after in vivo arsenic exposure was recapitulated in co-culture model of isolated HSCs and hepatocytes exposed to arsenic. These findings have implications not only for the understanding of the pathology of arsenic related liver fibrosis but also for the design of preventive strategies in chronic arsenicosis.

  7. PTH-enhanced structural allograft healing is associated with decreased angiopoietin-2-mediated arteriogenesis, mast cell accumulation, and fibrosis.

    PubMed

    Dhillon, Robinder S; Xie, Chao; Tyler, Wakenda; Calvi, Laura M; Awad, Hani A; Zuscik, Michael J; O'Keefe, Regis J; Schwarz, Edward M

    2013-03-01

    Recombinant parathyroid hormone (rPTH) therapy has been evaluated for skeletal repair in animal studies and clinical trials based on its known anabolic effects, but its effects on angiogenesis and fibrosis remain poorly understood. We examined the effects of rPTH therapy on blood vessel formation and osseous integration in a murine femoral allograft model, which caused a significant increase in small vessel numbers, and decreased large vessel formation (p < 0.05). Histology showed that rPTH also reduced fibrosis around the allografts to similar levels observed in live autografts, and decreased mast cells at the graft-host junction. Similar effects on vasculogenesis and fibrosis were observed in femoral allografts from Col1caPTHR transgenic mice. Gene expression profiling revealed rPTH-induced angiopoietin-1 (8-fold), while decreasing angiopoietin-2 (70-fold) at day 7 of allograft healing. Finally, we show anti-angiopoietin-2 peptibody (L1-10) treatment mimics rPTH effects on angiogenesis and fibrosis. Collectively, these findings show that intermittent rPTH treatment enhances structural allograft healing by two processes: (1) anabolic effects on new bone formation via small vessel angiogenesis, and (2) inhibition of angiopoietin-2-mediated arteriogenesis. The latter effect may function as a vascular sieve to limit mast cell access to the site of tissue repair, which decreases fibrosis around and between the fractured ends of bone. Thus, rPTH therapy may be generalizable to all forms of tissue repair that suffer from limited biointegration and excessive fibrosis.

  8. Computational Modeling Predicts Simultaneous Targeting of Fibroblasts and Epithelial Cells Is Necessary for Treatment of Pulmonary Fibrosis

    PubMed Central

    Warsinske, Hayley C.; Wheaton, Amanda K.; Kim, Kevin K.; Linderman, Jennifer J.; Moore, Bethany B.; Kirschner, Denise E.

    2016-01-01

    Pulmonary fibrosis is pathologic remodeling of lung tissue that can result in difficulty breathing, reduced quality of life, and a poor prognosis for patients. Fibrosis occurs as a result of insult to lung tissue, though mechanisms of this response are not well-characterized. The disease is driven in part by dysregulation of fibroblast proliferation and differentiation into myofibroblast cells, as well as pro-fibrotic mediator-driven epithelial cell apoptosis. The most well-characterized pro-fibrotic mediator associated with pulmonary fibrosis is TGF-β1. Excessive synthesis of, and sensitivity to, pro-fibrotic mediators as well as insufficient production of and sensitivity to anti-fibrotic mediators has been credited with enabling fibroblast accumulation. Available treatments neither halt nor reverse lung damage. In this study we have two aims: to identify molecular and cellular scale mechanisms driving fibroblast proliferation and differentiation as well as epithelial cell survival in the context of fibrosis, and to predict therapeutic targets and strategies. We combine in vitro studies with a multi-scale hybrid agent-based computational model that describes fibroblasts and epithelial cells in co-culture. Within this model TGF-β1 represents a pro-fibrotic mediator and we include detailed dynamics of TGF-β1 receptor ligand signaling in fibroblasts. PGE2 represents an anti-fibrotic mediator. Using uncertainty and sensitivity analysis we identify TGF-β1 synthesis, TGF-β1 activation, and PGE2 synthesis among the key mechanisms contributing to fibrotic outcomes. We further demonstrate that intervention strategies combining potential therapeutics targeting both fibroblast regulation and epithelial cell survival can promote healthy tissue repair better than individual strategies. Combinations of existing drugs and compounds may provide significant improvements to the current standard of care for pulmonary fibrosis. Thus, a two-hit therapeutic intervention strategy

  9. Computational modeling predicts simultaneous targeting of fibroblasts and epithelial cells is necessary for treatment of pulmonary fibrosis

    SciTech Connect

    Warsinske, Hayley C.; Wheaton, Amanda K.; Kim, Kevin K.; Linderman, Jennifer J.; Moore, Bethany B.; Kirschner, Denise E.

    2016-06-23

    Pulmonary fibrosis is pathologic remodeling of lung tissue that can result in difficulty breathing, reduced quality of life, and a poor prognosis for patients. Fibrosis occurs as a result of insult to lung tissue, though mechanisms of this response are not well-characterized. The disease is driven in part by dysregulation of fibroblast proliferation and differentiation into myofibroblast cells, as well as pro-fibrotic mediator-driven epithelial cell apoptosis. The most well-characterized pro-fibrotic mediator associated with pulmonary fibrosis is TGF-β1. Excessive synthesis of, and sensitivity to, pro-fibrotic mediators as well as insufficient production of and sensitivity to anti-fibrotic mediators has been credited with enabling fibroblast accumulation. Available treatments neither halt nor reverse lung damage. In this study we have two aims: to identify molecular and cellular scale mechanisms driving fibroblast proliferation and differentiation as well as epithelial cell survival in the context of fibrosis, and to predict therapeutic targets and strategies. We combine in vitro studies with a multi-scale hybrid agent-based computational model that describes fibroblasts and epithelial cells in co-culture. Within this model TGF-β1 represents a pro-fibrotic mediator and we include detailed dynamics of TGFβ1 receptor ligand signaling in fibroblasts. PGE2 represents an anti-fibrotic mediator. Using uncertainty and sensitivity analysis we identify TGF-β1 synthesis, TGF-β1 activation, and PGE2 synthesis among the key mechanisms contributing to fibrotic outcomes. We further demonstrate that intervention strategies combining potential therapeutics targeting both fibroblast regulation and epithelial cell survival can promote healthy tissue repair better than individual strategies. Combinations of existing drugs and compounds may provide significant improvements to the current standard of care for pulmonary fibrosis. In conclusion, a two

  10. Computational modeling predicts simultaneous targeting of fibroblasts and epithelial cells is necessary for treatment of pulmonary fibrosis

    DOE PAGES

    Warsinske, Hayley C.; Wheaton, Amanda K.; Kim, Kevin K.; ...

    2016-06-23

    Pulmonary fibrosis is pathologic remodeling of lung tissue that can result in difficulty breathing, reduced quality of life, and a poor prognosis for patients. Fibrosis occurs as a result of insult to lung tissue, though mechanisms of this response are not well-characterized. The disease is driven in part by dysregulation of fibroblast proliferation and differentiation into myofibroblast cells, as well as pro-fibrotic mediator-driven epithelial cell apoptosis. The most well-characterized pro-fibrotic mediator associated with pulmonary fibrosis is TGF-β1. Excessive synthesis of, and sensitivity to, pro-fibrotic mediators as well as insufficient production of and sensitivity to anti-fibrotic mediators has been credited withmore » enabling fibroblast accumulation. Available treatments neither halt nor reverse lung damage. In this study we have two aims: to identify molecular and cellular scale mechanisms driving fibroblast proliferation and differentiation as well as epithelial cell survival in the context of fibrosis, and to predict therapeutic targets and strategies. We combine in vitro studies with a multi-scale hybrid agent-based computational model that describes fibroblasts and epithelial cells in co-culture. Within this model TGF-β1 represents a pro-fibrotic mediator and we include detailed dynamics of TGFβ1 receptor ligand signaling in fibroblasts. PGE2 represents an anti-fibrotic mediator. Using uncertainty and sensitivity analysis we identify TGF-β1 synthesis, TGF-β1 activation, and PGE2 synthesis among the key mechanisms contributing to fibrotic outcomes. We further demonstrate that intervention strategies combining potential therapeutics targeting both fibroblast regulation and epithelial cell survival can promote healthy tissue repair better than individual strategies. Combinations of existing drugs and compounds may provide significant improvements to the current standard of care for pulmonary fibrosis. In conclusion, a two-hit therapeutic

  11. Skin elasticity as a measure of radiation fibrosis: is it reproducible and does it correlate with patient and physician-reported measures?

    PubMed

    Nguyen, Nhu-Tram A; Roberge, David; Freeman, Carolyn R; Wong, Cindy; Hines, Jerod; Turcotte, Robert E

    2014-10-01

    Current means of measuring RT-induced fibrosis are subjective. We evaluated the DermaLab suction cup system to measure objectively skin deflection as a surrogate for fibrosis. Sixty-nine patients with E-STS were treated with limb-sparing surgery and 50-66 Grays (Gy) of RT. Using a "scleroderma" DermaLab Suction Cup, the skin stiffness was measured by two clinicians. The National Cancer Institute Common Terminology Criteria for Adverse Events (NCI-CTCAE) scale, the Musculoskeletal Tumor Rating Scale (MSTS) and Toronto Extremity Salvage Score (TESS) questionnaires were completed for each patient. Levels of agreement between measurers were estimated using the Kappa (k) coefficient and the concordance correlation coefficient (CCC). All sixty-nine patients were included. The level of agreement between measurers for NCI-CTCAE grading was moderate (range k = 0.41-0.59). The CCC for the elasticity measurements were higher, with CCC = 0.82 for fibrotic skin and CCC 5 0.84 for normal skin. The elasticity measurements were significantly higher when MSTS scores were <30 and or TESS scores were <90. Suction Cup measurement of skin elasticity is more reproducible than CTCAE grading and shows promise in generating reproducible measurements for radiation-induced skin fibrosis. Furthermore, it correlates well with the MSTS and TESS.

  12. CGRP induction in cystic fibrosis airways alters the submucosal gland progenitor cell niche in mice

    PubMed Central

    Xie, Weiliang; Fisher, John T.; Lynch, Thomas J.; Luo, Meihui; Evans, Turan I.A.; Neff, Traci L.; Zhou, Weihong; Zhang, Yulong; Ou, Yi; Bunnett, Nigel W.; Russo, Andrew F.; Goodheart, Michael J.; Parekh, Kalpaj R.; Liu, Xiaoming; Engelhardt, John F.

    2011-01-01

    In cystic fibrosis (CF), a lack of functional CF transmembrane conductance regulator (CFTR) chloride channels causes defective secretion by submucosal glands (SMGs), leading to persistent bacterial infection that damages airways and necessitates tissue repair. SMGs are also important niches for slow-cycling progenitor cells (SCPCs) in the proximal airways, which may be involved in disease-related airway repair. Here, we report that calcitonin gene–related peptide (CGRP) activates CFTR-dependent SMG secretions and that this signaling pathway is hyperactivated in CF human, pig, ferret, and mouse SMGs. Since CGRP-expressing neuroendocrine cells reside in bronchiolar SCPC niches, we hypothesized that the glandular SCPC niche may be dysfunctional in CF. Consistent with this hypothesis, CFTR-deficient mice failed to maintain glandular SCPCs following airway injury. In wild-type mice, CGRP levels increased following airway injury and functioned as an injury-induced mitogen that stimulated SMG progenitor cell proliferation in vivo and altered the proliferative potential of airway progenitors in vitro. Components of the receptor for CGRP (RAMP1 and CLR) were expressed in a very small subset of SCPCs, suggesting that CGRP indirectly stimulates SCPC proliferation in a non-cell-autonomous manner. These findings demonstrate that CGRP-dependent pathways for CFTR activation are abnormally upregulated in CF SMGs and that this sustained mitogenic signal alters properties of the SMG progenitor cell niche in CF airways. This discovery may have important implications for injury/repair mechanisms in the CF airway. PMID:21765217

  13. Characterization of mitochondrial function in cells with impaired cystic fibrosis transmembrane conductance regulator (CFTR) function.

    PubMed

    Atlante, Anna; Favia, Maria; Bobba, Antonella; Guerra, Lorenzo; Casavola, Valeria; Reshkin, Stephan Joel

    2016-06-01

    Evidence supporting the occurrence of oxidative stress in Cystic Fibrosis (CF) is well established and the literature suggests that oxidative stress is inseparably linked to mitochondrial dysfunction. Here, we have characterized mitochondrial function, in particular as it regards the steps of oxidative phosphorylation and ROS production, in airway cells either homozygous for the F508del-CFTR allele or stably expressing wt-CFTR. We find that oxygen consumption, ΔΨ generation, adenine nucleotide translocator-dependent ADP/ATP exchange and both mitochondrial Complex I and IV activities are impaired in CF cells, while both mitochondrial ROS production and membrane lipid peroxidation increase. Importantly, treatment of CF cells with the small molecules VX-809 and 4,6,4'-trimethylangelicin, which act as "correctors" for F508del CFTR by rescuing the F508del CFTR-dependent chloride secretion, while having no effect per sè on mitochondrial function in wt-CFTR cells, significantly improved all the above mitochondrial parameters towards values found in the airway cells expressing wt-CFTR. This novel study on mitochondrial bioenergetics provides a springboard for future research to further understand the molecular mechanisms responsible for the involvement of mitochondria in CF and identify the proteins primarily responsible for the F508del-CFTR-dependent mitochondrial impairment and thus reveal potential novel targets for CF therapy.

  14. A mast cell-ILC2-Th9 pathway promotes lung inflammation in cystic fibrosis

    PubMed Central

    Moretti, Silvia; Renga, Giorgia; Oikonomou, Vasilis; Galosi, Claudia; Pariano, Marilena; Iannitti, Rossana G.; Borghi, Monica; Puccetti, Matteo; De Zuani, Marco; Pucillo, Carlo E.; Paolicelli, Giuseppe; Zelante, Teresa; Renauld, Jean-Christophe; Bereshchenko, Oxana; Sportoletti, Paolo; Lucidi, Vincenzina; Russo, Maria Chiara; Colombo, Carla; Fiscarelli, Ersilia; Lass-Flörl, Cornelia; Majo, Fabio; Ricciotti, Gabriella; Ellemunter, Helmut; Ratclif, Luigi; Talesa, Vincenzo Nicola; Napolioni, Valerio; Romani, Luigina

    2017-01-01

    T helper 9 (Th9) cells contribute to lung inflammation and allergy as sources of interleukin-9 (IL-9). However, the mechanisms by which IL-9/Th9 mediate immunopathology in the lung are unknown. Here we report an IL-9-driven positive feedback loop that reinforces allergic inflammation. We show that IL-9 increases IL-2 production by mast cells, which leads to expansion of CD25+ type 2 innate lymphoid cells (ILC2) and subsequent activation of Th9 cells. Blocking IL-9 or inhibiting CD117 (c-Kit) signalling counteracts the pathogenic effect of the described IL-9-mast cell-IL-2 signalling axis. Overproduction of IL-9 is observed in expectorates from cystic fibrosis (CF) patients, and a sex-specific variant of IL-9 is predictive of allergic reactions in female patients. Our results suggest that blocking IL-9 may be a therapeutic strategy to ameliorate inflammation associated with microbial colonization in the lung, and offers a plausible explanation for gender differences in clinical outcomes of patients with CF. PMID:28090087

  15. Myeloperoxidase–Hepatocyte–Stellate Cell Cross Talk Promotes Hepatocyte Injury and Fibrosis in Experimental Nonalcoholic Steatohepatitis

    PubMed Central

    Pulli, Benjamin; Ali, Muhammad; Iwamoto, Yoshiko; Zeller, Matthias W.G.; Schob, Stefan; Linnoila, Jenny J.

    2015-01-01

    Abstract Aims: Myeloperoxidase (MPO), a highly oxidative enzyme secreted by leukocytes has been implicated in human and experimental nonalcoholic steatohepatitis (NASH), but the underlying mechanisms remain unknown. In this study, we investigated how MPO contributes to progression from steatosis to NASH. Results: In C57Bl/6J mice fed a diet deficient in methionine and choline to induce NASH, neutrophils and to a lesser extent inflammatory monocytes are markedly increased compared with sham mice and secrete abundant amounts of MPO. Through generation of HOCl, MPO directly causes hepatocyte death in vivo. In vitro experiments demonstrate mitochondrial permeability transition pore induction via activation of SAPK/JNK and PARP. MPO also contributes to activation of hepatic stellate cells (HSCs), the most important source of collagen in the liver. In vitro MPO-activated HSCs have an activation signature (MAPK and PI3K-AKT phosphorylation) and upregulate COL1A1, α-SMA, and CXCL1. MPO-derived oxidative stress also activates transforming growth factor β (TGF-β) in vitro, and TGF-β signaling inhibition with SB-431542 decreased steatosis and fibrosis in vivo. Conversely, congenital absence of MPO results in reduced hepatocyte injury, decreased levels of TGF-β, fewer activated HSCs, and less severe fibrosis in vivo. Innovation and Conclusion: Cumulatively, these findings demonstrate important cross talk between inflammatory myeloid cells, hepatocytes, and HSCs via MPO and establish MPO as part of a proapoptotic and profibrotic pathway of progression in NASH, as well as a potential therapeutic target to ameliorate this disease. Antioxid. Redox Signal. 23, 1255–1269. PMID:26058518

  16. The improving effects on hepatic fibrosis of interferon-γ liposomes targeted to hepatic stellate cells

    NASA Astrophysics Data System (ADS)

    Li, Qinghua; Yan, Zhiqiang; Li, Feng; Lu, Weiyue; Wang, Jiyao; Guo, Chuanyong

    2012-07-01

    No satisfactory anti-fibrotic therapies have yet been applied clinically. One of the main reasons is the inability to specifically target the responsible cells to produce an available drug concentration and the side-effects. Exploiting the key role of the activated hepatic stellate cells (HSCs) in both hepatic fibrogenesis and over-expression of platelet-derived growth factor receptor-β (PDGFR-β), we constructed targeted sterically stable liposomes (SSLs) modified by a cyclic peptide (pPB) with affinity for the PDGFR-β to deliver interferon (IFN)-γ to HSCs. The pPB-SSL-IFN-γ showed satisfactory size distribution. In vitro pPB-SSL could be taken up by activated HSCs. The study of tissue distribution via living-body animal imaging showed that the pPB-SSL-IFN-γ mostly accumulated in the liver until 24 h. Furthermore, the pPB-SSL-IFN-γ showed more significant remission of hepatic fibrosis. In vivo the histological Ishak stage, the semiquantitative score for collagen in fibrotic liver and the serum levels of collagen type IV-C in fibrotic rats treated with pPB-SSL-IFN-γ were less than those treated with SSL-IFN-γ, IFN-γ and the control group. In vitro pPB-SSL-IFN-γ was also more effective in suppressing activated HSC proliferation and inducing apoptosis of activated HSCs. Thus the data suggest that pPB-SSL-IFN-γ might be a more effective anti-fibrotic agent and a new opportunity for clinical therapy of hepatic fibrosis.

  17. Plasma B-Lymphocyte Stimulator (BLyS) and B-cell Differentiation in Idiopathic Pulmonary Fibrosis Patients*

    PubMed Central

    Xue, Jianmin; Kass, Daniel J.; Bon, Jessica; Vuga, Louis; Tan, Jiangning; Csizmadia, Eva; Otterbein, Leo; Soejima, Makoto; Levesque, Marc C.; Gibson, Kevin F.; Kaminski, Naftali; Pilewski, Joseph M.; Donahoe, Michael; Sciurba, Frank C.; Duncan, Steven R.

    2013-01-01

    We hypothesized B-cells are involved in the pathogenesis of idiopathic pulmonary fibrosis (IPF), a progressive, restrictive lung disease that is refractory to glucocorticoids and other nonspecific therapies, and almost invariably lethal. Accordingly, we sought to identify clinically-associated B-cell-related abnormalities in these patients. Phenotypes of circulating B-cells were characterized by flow cytometry. Intrapulmonary processes were evaluated by immunohistochemistry. Plasma B-lymphocyte stimulating factor (BLyS) was assayed by ELISA. Circulating B-cells of IPF subjects were more antigen-differentiated, with greater plasmablast proportions (3.1±0.8%) than in normal controls (1.3±0.3%) (p<0.03), and the extent of this differentiation correlated with IPF patient lung volumes (r=0.44, p<0.03). CD20+ B-cell aggregates, diffuse parenchymal and perivascular immune complexes, and complement depositions were all prevalent in IPF lungs, but much less prominent or absent in normal lungs. Plasma concentrations of BLyS, an obligate factor for B-cell survival and differentiation, were significantly greater (p<0.0001) in 110 IPF (2.05±0.05 ng/ml) than among 53 normal (1.40±0.04 ng/ml) and 90 chronic obstructive pulmonary disease (COPD) subjects (1.59±0.05 ng/ml). BLyS levels were uniquely correlated among IPF patients with pulmonary artery pressures (r=0.58, p<0.0001). The 25% of IPF subjects with the greatest BLyS values also had diminished one-year survival (46±11%), compared to those with lesser BLyS concentrations (81±5%) (HR=4.0, 95%CI=1.8-8.7, p=0.0002). Abnormalities of B-cells and BLyS are common in IPF patients, and highly associated with disease manifestations and patient outcomes. These findings have implications regarding IPF pathogenesis, and illuminate the potential for novel treatment regimens that specifically target B-cells in patients with this lung disease. PMID:23872052

  18. Rhinovirus Load Is High despite Preserved Interferon-β Response in Cystic Fibrosis Bronchial Epithelial Cells

    PubMed Central

    Cammisano, Maria; Chen, He; Singh, Sareen; Kooi, Cora; Leigh, Richard; Beaudoin, Trevor; Rousseau, Simon; Lands, Larry C.

    2015-01-01

    Lung disease in cystic fibrosis (CF) is often exacerbated following acute upper respiratory tract infections caused by the human rhinovirus (HRV). Pathophysiology of these exacerbations is presently unclear and may involve deficient innate antiviral or exaggerated inflammatory responses in CF airway epithelial cells. Furthermore, responses of CF cells to HRV may be adversely affected by pre-exposure to virulence factors of Pseudomonas (P.) aeruginosa, the microorganism that frequently colonizes CF airways. Here we examined production of antiviral cytokine interferon-β and inflammatory chemokine interleukin-8, expression of the interferon-responsive antiviral gene 2’-5’-oligoadenylate synthetase 1 (OAS1), and intracellular virus RNA load in primary CF (delF508 CFTR) and healthy airway epithelial cells following inoculation with HRV16. Parallel cells were exposed to virulence factors of P. aeruginosa prior to and during HRV16 inoculation. CF cells exhibited production of interferon-β and interleukin-8, and expression of OAS1 at levels comparable to those in healthy cells, yet significantly higher HRV16 RNA load during early hours post-inoculation with HRV16. In line with this, HRV16 RNA load was higher in the CFBE41o- dF cell line overexpessing delF508 CFTR, compared with the isogenic control CFBE41o- WT (wild-type CFTR). Pre-exposure to virulence factors of P. aeruginosa did not affect OAS1 expression or HRV16 RNA load, but potentiated interleukin-8 production. In conclusion, CF cells demonstrate elevated HRV RNA load despite preserved interferon-β and OAS1 responses. High HRV load in CF airway epithelial cells appears to be due to deficiencies manifesting early during HRV infection, and may not be related to interferon-β. PMID:26599098

  19. Burkholderia cenocepacia ET12 Strain Activates TNFR1 Signaling in Cystic Fibrosis Airway Epithelial Cells

    PubMed Central

    Umadevi Sajjan, S.; Hershenson, Marc B.; Forstner, Janet F.; LiPuma, John J.

    2011-01-01

    Burkholderia cenocepacia is an important pulmonary pathogen in individuals with cystic fibrosis. Infection is often associated with severe pulmonary inflammation and some patients develop a fatal necrotizing pneumonia and sepsis (‘cepacia syndrome’). The mechanisms by which this species causes severe pulmonary inflammation are poorly understood. Here, we demonstrate that B. cenocepacia BC7, a potentially virulent representative of the epidemic ET12 lineage binds to tumor necrosis factor receptor I (TNFR1) and activates TNFR1-related signaling pathway similar to TNF-α, a natural ligand for TNFR1. This interaction participates in stimulating a robust IL-8 production from CF airway epithelial cells. In contrast, BC45, a relatively less virulent ET12 representative, and ATCC 25416, an environmental B. cepacia strain do not bind to TNFR1 and stimulate only minimal IL-8 production from CF cells. Further, TNFR1 expression is increased in CF airway epithelial cells compared to non-CF cells. We also show that B. cenocepacia ET12 strain colocaizes with TNFR1 in vitro and in the lungs of CF patient who died due to infection with B. cenocepacia, ET12 strain. Together, these results suggest that interaction of B. cenocepacia, ET12 strain with TNFR1 may contribute to robust inflammatory responses elicited by this organism. PMID:17697131

  20. Relationships among injury, fibrosis, and time in human kidney transplants

    PubMed Central

    Venner, Jeffery M.; Famulski, Konrad S.; Reeve, Jeff; Chang, Jessica; Halloran, Philip F.

    2016-01-01

    BACKGROUND. Kidney transplant biopsies offer an opportunity to understand the pathogenesis of organ fibrosis. We studied the relationships between the time of biopsy after transplant (TxBx), histologic fibrosis, diseases, and transcript expression. METHODS. Expression microarrays from 681 kidney transplant indication biopsies taken either early (n = 282, <1 year) or late (n = 399, >1 year) after transplant were used to analyze the molecular landscape of fibrosis in relationship to histologic fibrosis and diseases. RESULTS. Fibrosis was absent at transplantation but was present in some early biopsies by 4 months after transplant, apparently as a self-limited response to donation implantation injury not associated with progression to failure. The molecular phenotype of early biopsies represented the time sequence of the response to wounding: immediate expression of acute kidney injury transcripts, followed by fibrillar collagen transcripts after several weeks, then by the appearance of immunoglobulin and mast cell transcripts after several months as fibrosis appeared. Fibrosis in late biopsies correlated with injury, fibrillar collagen, immunoglobulin, and mast cell transcripts, but these were independent of time. Pathway analysis revealed epithelial response-to-wounding pathways such as Wnt/β-catenin. CONCLUSION. Fibrosis in late biopsies had different associations because many kidneys had potentially progressive diseases and subsequently failed. Molecular correlations with fibrosis in late biopsies were independent of time, probably because ongoing injury obscured the response-to-wounding time sequence. The results indicate that fibrosis in kidney transplants is driven by nephron injury and that progression to failure reflects continuing injury, not autonomous fibrogenesis. TRIAL REGISTRATION. INTERCOM study (www.clinicalTrials.gov; NCT01299168). FUNDING. Canada Foundation for Innovation and Genome Canada. PMID:27699214

  1. Genotype-phenotype correlation in cystic fibrosis patients bearing [H939R;H949L] allele

    PubMed Central

    Polizzi, Angela; Tesse, Riccardina; Santostasi, Teresa; Diana, Anna; Manca, Antonio; Logrillo, Vito Paolo; Cazzato, Maria Domenica; Pantaleo, Maria Giuseppa; Armenio, Lucio

    2011-01-01

    Cystic fibrosis (CF) is caused by CFTR (cystic fibrosis transmembrane conductance regulator) gene mutations. We ascertained five patients with a novel complex CFTR allele, with two mutations, H939R and H949L, inherited in cis in the same exon of CFTR gene, and one different mutation per patient inherited in trans in a wide population of 289 Caucasian CF subjects from South Italy. The genotype-phenotype relationship in patients bearing this complex allele was investigated. The two associated mutations were related to classical severe CF phenotypes. PMID:21931512

  2. Cystic Fibrosis patients have inducible IL-17+IL-22+ memory cells in lung draining lymph nodes

    PubMed Central

    Chan, Yvonne R.; Chen, Kong; Duncan, Steven R.; Lathrop, Kira; Latoche, Joseph; Logar, Alison; Pociask, Derek A.; Wahlberg, Brendon; Ray, Prabir; Ray, Anuradha; Pilewski, Joseph M.; Kolls, Jay K.

    2012-01-01

    Background Interleukin (IL)-17 is an important cytokine signature of a T helper differentiation pathway, Th17. This T cell subset is crucial in mediating autoimmune disease or antimicrobial immunity in animal models, but its presence and role in human disease remains to be completely characterized. Objective We set out to determine the frequency of Th17 cells in cystic fibrosis (CF), a disease in which there is recurrent infection with known pathogens. Methods Explanted lungs from patients undergoing transplant or organ donors (CF = 18, non-CF, non-bronchiectatic = 10) were collected. Hilar nodes and parenchymal lung tissue were processed. We examined them for Th17 signature by immunofluorescence and quantitative real time PCR. T cells were isolated and stimulated with antigens from Pseudomonas aeruginosa and Aspergillus. Cytokine profiles and staining by flow cytometry were used to assess the reactivity of these cells to antigen stimulation. Results We found a strong IL-17 phenotype in CF compared to non-CF controls. Within this tissue, we found pathogen-antigen-responsive CD4+IL17+ cells. There were double positive IL-17+IL-22+ cells and the IL-22+ population had higher proportions of memory characteristics. Antigen-specific Th17 responses were stronger in the draining lymph nodes compared to matched parenchymal lung. Conclusion Inducible proliferation of Th17(22) with memory cell characteristics is seen in CF lung. The function of these individual subpopulations will require further study regarding their development. T-cells are likely not the exclusive producers of IL-17 and IL-22 and this will require further characterization. PMID:22795370

  3. Toward an animal model of cystic fibrosis: Targeted interruption of exon 10 of the cystic fibrosis transmembrane regulator gene in embryonic stem cells

    SciTech Connect

    Koller, B.H.; Hyungsuk Kim; Latour, A.M.; Brigman, K.; Boucher, R.C. Jr.; Smithies, O. ); Scambler, P.; Wainwright, B. )

    1991-12-01

    A gene-targeting construct was made containing 7.8 kilobases of DNA spanning exon 10 of the mouse cystic fibrosis transmembrane regulator (CFTR) gene in which part of the exon has been replaced by two neomycin-resistance (Neo) genes driven by different promoters. (This replacement introduces a chain-termination codon at amino acid position 489 in the CFTR sequence). A herpes simplex thymidine kinase gene was on each end of the construct, which was electroporated into embryonic stem (ES) cells. Colonies resistant to G418, or to G418 plus ganciclovir, were selected and screened by Southern blotting or by PCR amplification. Five pools of G418-resistant cells gave PCR products diagnostic of targeting. Four independent clones of ES cells with a disrupted CFTR gene have been isolated from these pools. The frequency of targeting was 1/2500 G418-resistant colonies. This low frequency is not the consequence of marginal expression of the Neo genes in the targeted cells. The CFTR targeting events were clustered among our experiments in a manner suggesting that some unidentified factor(s), possible passage number, influences the recovery of CFTR-targeted cells.

  4. miR-1273g-3p modulates activation and apoptosis of hepatic stellate cells by directly targeting PTEN in HCV-related liver fibrosis.

    PubMed

    Niu, Xuemin; Fu, Na; Du, Jinghua; Wang, Rongqi; Wang, Yang; Zhao, Suxian; Du, Huijuan; Wang, Baoyu; Zhang, Yuguo; Sun, Dianxing; Nan, Yuemin

    2016-08-01

    MicroRNA (miRNA) play a pivotal role in the development of liver fibrosis. However, the functions of miRNA in hepatitis C virus (HCV)-related liver fibrosis remain unclear. In this study, we systematically analyzed the microarray data of the serum miRNA in patients with HCV-induced hepatic fibrosis. Among 41 dysregulated miRNA, miR-1273g-3p was the most significantly upregulated miRNA and correlated with the stage of liver fibrosis. Overexpression of miR-1273g-3p could inhibit translation of PTEN, increase the expression of α-SMA, Col1A1, and reduce apoptosis in HSCs. Hence, we conclude that miR-1273g-3p might affect the activation and apoptosis of HSCs by directly targeting PTEN in HCV-related liver fibrosis.

  5. Pulmonary Fibrosis

    MedlinePlus

    Pulmonary fibrosis is a condition in which the tissue deep in your lungs becomes scarred over time. This tissue ... may not get enough oxygen. Causes of pulmonary fibrosis include environmental pollutants, some medicines, some connective tissue ...

  6. Alteration and localization of glycan-binding proteins in human hepatic stellate cells during liver fibrosis.

    PubMed

    Zhong, Yaogang; Qin, Yannan; Dang, Liuyi; Jia, Liyuan; Zhang, Zhiwei; Wu, Haoxiang; Cui, Jihong; Bian, Huijie; Li, Zheng

    2015-10-01

    Glycan-binding proteins (GBPs) play an important role in cell adhesion, bacterial/viral infection, and cellular signaling pathways. However, little is known about the precision alteration of GBPs referred to pathological changes in hepatic stellate cells (HSCs) during liver fibrosis. Here, the carbohydrate microarrays were used to probe the alteration of GBPs in the activated HSCs and quiescent HSCs. As a result, 12 carbohydrates (e.g. Gal, GalNAc, and Man-9Glycan) showed increased signal, while seven carbohydrates (e.g. NeuAc, Lac, and GlcNAc-O-Ser) showed decreased signal in activated HSCs. Three carbohydrates (Gal, GalNAc, and NeuAc) were selected and subsequently used to validate the results of the carbohydrate microarrays as well as assess the distribution and localization of their binding proteins in HSCs and liver tissues by cy/histochemistry; the results showed that GBPs mainly distributed in the cytoplasma membrane and perinuclear region of cytoplasm. The immunocytochemistry was further used to verify some GBPs really exist in Golgi apparatus of the cells. The precision alteration and localization of GBPs referred to pathological changes in HSCs may provide pivotal information to help understand the biological functions of glycans how to exert through their recognition by a wide variety of GBPs. This study could lead to the development of new anti-fibrotic strategies.

  7. EZH2-mediated repression of Dkk1 promotes hepatic stellate cell activation and hepatic fibrosis.

    PubMed

    Yang, Yang; Chen, Xiao-Xia; Li, Wan-Xia; Wu, Xiao-Qin; Huang, Cheng; Xie, Juan; Zhao, Yu-Xin; Meng, Xiao-Ming; Li, Jun

    2017-03-23

    EZH2, a histone H3 lysine-27-specific methyltransferase, is involved in diverse physiological and pathological processes including cell proliferation and differentiation. However, the role of EZH2 in liver fibrosis is largely unknown. In this study, it was identified that EZH2 promoted Wnt pathway-stimulated fibroblasts in vitro and in vivo by repressing Dkk-1, which is a Wnt pathway antagonist. The expression of EZH2 was increased in CCl4 -induced rat liver and primary HSCs as well as TGF-β1-treated HSC-T6, whereas the expression of Dkk1 was reduced. Silencing of EZH2 prevented TGF-β1-induced proliferation of HSC-T6 cells and the expression of α-SMA. In addition, knockdown of Dkk1 promoted TGF-β1-induced activation of HSCs. Moreover, silencing of EZH2 could restore the repression of Dkk-1 through trimethylation of H3K27me3 in TGF-β1-treated HSC-T6 cells. Interestingly, inhibition of EZH2 had almost no effect on the activation of HSC when Dkk1 was silenced. Collectively, EZH2-mediated repression of Dkk1 promotes the activation of Wnt/β-catenin pathway, which is an essential event for HSC activation.

  8. Abnormal apical cell membrane in cystic fibrosis respiratory epithelium. An in vitro electrophysiologic analysis.

    PubMed Central

    Cotton, C U; Stutts, M J; Knowles, M R; Gatzy, J T; Boucher, R C

    1987-01-01

    The transepithelial chloride permeability of airway and sweat ductal epithelium has been reported to be decreased in patients with cystic fibrosis (CF). In the present study, we investigated whether the airway epithelial defect was in the cell path by characterizing the relative ion permeabilities of the apical membrane of respiratory epithelial cells from CF and normal subjects. Membrane electric potential difference (PD) and the responses to luminal Cl- replacement, isoproterenol, and amiloride were measured with intracellular microelectrodes. The PD across the apical barrier was smaller for CF (-11 mV) than normal (-29 mV) epithelia whereas the PD across the basolateral barrier was similar, (-26 and -34 mV respectively). In contrast to normal nasal epithelium, the apical membrane in CF epithelia was not Cl- permselective and was not responsive to isoproterenol. Amiloride, a selective Na+ channel blocker, induced a larger apical membrane hyperpolarization and a greater increase in transepithelial resistance in CF epithelia. Both reduced apical cell membrane Cl- conductance and increased Na+ conductance appear to contribute to the abnormal function of respiratory epithelia of CF patients. PMID:3793933

  9. Functional repair of CFTR by CRISPR/Cas9 in intestinal stem cell organoids of cystic fibrosis patients.

    PubMed

    Schwank, Gerald; Koo, Bon-Kyoung; Sasselli, Valentina; Dekkers, Johanna F; Heo, Inha; Demircan, Turan; Sasaki, Nobuo; Boymans, Sander; Cuppen, Edwin; van der Ent, Cornelis K; Nieuwenhuis, Edward E S; Beekman, Jeffrey M; Clevers, Hans

    2013-12-05

    Single murine and human intestinal stem cells can be expanded in culture over long time periods as genetically and phenotypically stable epithelial organoids. Increased cAMP levels induce rapid swelling of such organoids by opening the cystic fibrosis transmembrane conductor receptor (CFTR). This response is lost in organoids derived from cystic fibrosis (CF) patients. Here we use the CRISPR/Cas9 genome editing system to correct the CFTR locus by homologous recombination in cultured intestinal stem cells of CF patients. The corrected allele is expressed and fully functional as measured in clonally expanded organoids. This study provides proof of concept for gene correction by homologous recombination in primary adult stem cells derived from patients with a single-gene hereditary defect.

  10. AST to Platelet Index Correlates with Hepatic Cirrhosis But Not with Fibrosis in Pediatric Patients with Intestinal Failure

    PubMed Central

    Díaz, Juan J; Gura, Kathleen M.; Roda, Juliamna; Perez-Atayde, Antonio R.; Duggan, Christopher; Jaksic, Tom; Lo, Clifford W.

    2013-01-01

    Patients with Intestinal failure (IF) require parenteral nutrition (PN) support to obtain enough nutrients to sustain growth. long-term PN use is associated with significant liver damage. Objective To analyze the utility of a non-invasive test, the aspartate aminotransferase (AST) to platelet ratio index (APRI), in the diagnosis of liver disease in pediatric patients with IF. Methods Medical records of all Boston Children’s Hospital patients who received PN and underwent a liver biopsy from January 2006 until November 2010 were reviewed. Patients with a clinical diagnosis with IF were selected. APRI was calculated as follows (AST (U/L)/ upper normal limit) × 100/ platelets (109/L). Presence of fibrosis and cirrhosis was estimated using the METAVIR score in liver biopsies. Results 62 liver biopsies from 48 patients (22 female) were studied. Mean APRI values in the different METAVIR categories (0-1; 2-3; 4) were: 1.80, 1.17, and 4.24 respectively (ANOVA; P=0.053; Bonferroni test for cirrhosis versus fibrosis P=0.048). APRI could significantly predict cirrhosis (OR 1.2.; 95% CI 1.001-1.43) but not significant fibrosis (METAVIR 2-3, OR 1.00; 95% CI =0.86-1.18). Area under the receiver operating characteristic curve for cirrhosis was 0.67 (95% CI= 0.45-0.89; p=0.13). Conclusion APRI, a non invasive, easy to obtain bedside test significantly predicts cirrhosis but not fibrosis in pediatric patients with IFALD. As the clinicians need a non invasive test to differentiate among different stages of liver fibrosis rather than differentiating cirrhosis from normal, we cannot recommend the use of this test in pediatric patients with IFALD for this purpose. PMID:23666459

  11. Mobilization of endogenous bone marrow-derived stem cells in a thioacetamide-induced mouse model of liver fibrosis.

    PubMed

    El-Akabawy, Gehan; El-Mehi, Abeer

    2015-06-01

    The clinical significance of enhancing endogenous circulating haematopoietic stem cells is becoming increasingly recognized, and the augmentation of circulating stem cells using granulocyte-colony stimulating factor (G-CSF) has led to promising preclinical and clinical results for several liver fibrotic conditions. However, this approach is largely limited by cost and the infeasibility of maintaining long-term administration. Preclinical studies have reported that StemEnhance, a mild haematopoietic stem cell mobilizer, promotes cardiac muscle regeneration and remedies the manifestation of diabetes. However, the effectiveness of StemEnhance in ameliorating liver cirrhosis has not been studied. This study is the first to evaluate the beneficial effect of StemEnhance administration in a thioacetamide-induced mouse model of liver fibrosis. StemEnhance augmented the number of peripheral CD34-positive cells, reduced hepatic fibrosis, improved histopathological changes, and induced endogenous liver proliferation. In addition, VEGF expression was up-regulated, while TNF-α expression was down-regulated in thioacetamide-induced fibrotic livers after StemEnhance intake. These data suggest that StemEnhance may be useful as a potential therapeutic candidate for liver fibrosis by inducing reparative effects via mobilization of haematopoietic stem cells.

  12. Molecular magnetic resonance imaging of activated hepatic stellate cells with ultrasmall superparamagnetic iron oxide targeting integrin αvβ3 for staging liver fibrosis in rat model

    PubMed Central

    Zhang, Caiyuan; Liu, Huanhuan; Cui, Yanfen; Li, Xiaoming; Zhang, Zhongyang; Zhang, Yong; Wang, Dengbin

    2016-01-01

    Purpose To evaluate the expression level of integrin αvβ3 on activated hepatic stellate cells (HSCs) at different stages of liver fibrosis induced by carbon tetrachloride (CCl4) in rat model and the feasibility to stage liver fibrosis by using molecular magnetic resonance imaging (MRI) with arginine-glycine-aspartic acid (RGD) peptide modified ultrasmall superparamagnetic iron oxide nanoparticle (USPIO) specifically targeting integrin αvβ3. Materials and methods All experiments received approval from our Institutional Animal Care and Use Committee. Thirty-six rats were randomly divided into three groups of 12 subjects each, and intraperitoneally injected with CCl4 for either 3, 6, or 9 weeks. Controls (n=10) received pure olive oil. The change in T2* relaxation rate (ΔR2*) pre- and postintravenous administration of RGD-USPIO or naked USPIO was measured by 3.0T clinical MRI and compared by one-way analysis of variance or the Student’s t-test. The relationship between expression level of integrin αvβ3 and liver fibrotic degree was evaluated by Spearman’s ranked correlation. Results Activated HSCs were confirmed to be the main cell types expressing integrin αvβ3 during liver fibrogenesis. The protein level of integrin αv and β3 subunit expressed on activated HSCs was upregulated and correlated well with the progression of liver fibrosis (r=0.954, P<0.001; r=0.931, P<0.001, respectively). After injection of RGD-USPIO, there is significant difference in ΔR2* among rats treated with 0, 3, 6, and 9 weeks of CCl4 (P<0.001). The accumulation of iron particles in fibrotic liver specimen is significantly greater for RGD-USPIO than naked USPIO after being injected with equal dose of iron. Conclusion Molecular MRI of integrin αvβ3 expressed on activated HSCs by using RGD-USPIO may distinguish different liver fibrotic stages in CCl4 rat model and shows promising to noninvasively monitor the progression of the liver fibrosis and therapeutic response to

  13. Endotoxin Induces Fibrosis in Vascular Endothelial Cells through a Mechanism Dependent on Transient Receptor Protein Melastatin 7 Activity

    PubMed Central

    Echeverría, Cesar; Montorfano, Ignacio; Hermosilla, Tamara; Armisén, Ricardo; Velásquez, Luis A.; Cabello-Verrugio, Claudio; Varela, Diego; Simon, Felipe

    2014-01-01

    The pathogenesis of systemic inflammatory diseases, including endotoxemia-derived sepsis syndrome, is characterized by endothelial dysfunction. It has been demonstrated that the endotoxin lipopolysaccharide (LPS) induces the conversion of endothelial cells (ECs) into activated fibroblasts through endothelial­to­mesenchymal transition mechanism. Fibrogenesis is highly dependent on intracellular Ca2+ concentration increases through the participation of calcium channels. However, the specific molecular identity of the calcium channel that mediates the Ca2+ influx during endotoxin-induced endothelial fibrosis is still unknown. Transient receptor potential melastatin 7 (TRPM7) is a calcium channel that is expressed in many cell types, including ECs. TRPM7 is involved in a number of crucial processes such as the conversion of fibroblasts into activated fibroblasts, or myofibroblasts, being responsible for the development of several characteristics of them. However, the role of the TRPM7 ion channel in endotoxin-induced endothelial fibrosis is unknown. Thus, our aim was to study whether the TRPM7 calcium channel participates in endotoxin-induced endothelial fibrosis. Using primary cultures of ECs, we demonstrated that TRPM7 is a crucial protein involved in endotoxin-induced endothelial fibrosis. Suppression of TRPM7 expression protected ECs from the fibrogenic process stimulated by endotoxin. Downregulation of TRPM7 prevented the endotoxin-induced endothelial markers decrease and fibrotic genes increase in ECs. In addition, TRPM7 downregulation abolished the endotoxin-induced increase in ECM proteins in ECs. Furthermore, we showed that intracellular Ca2+ levels were greatly increased upon LPS challenge in a mechanism dependent on TRPM7 expression. These results demonstrate that TRPM7 is a key protein involved in the mechanism underlying endotoxin-induced endothelial fibrosis. PMID:24710004

  14. Endotoxin induces fibrosis in vascular endothelial cells through a mechanism dependent on transient receptor protein melastatin 7 activity.

    PubMed

    Echeverría, Cesar; Montorfano, Ignacio; Hermosilla, Tamara; Armisén, Ricardo; Velásquez, Luis A; Cabello-Verrugio, Claudio; Varela, Diego; Simon, Felipe

    2014-01-01

    The pathogenesis of systemic inflammatory diseases, including endotoxemia-derived sepsis syndrome, is characterized by endothelial dysfunction. It has been demonstrated that the endotoxin lipopolysaccharide (LPS) induces the conversion of endothelial cells (ECs) into activated fibroblasts through endothelial-to-mesenchymal transition mechanism. Fibrogenesis is highly dependent on intracellular Ca2+ concentration increases through the participation of calcium channels. However, the specific molecular identity of the calcium channel that mediates the Ca2+ influx during endotoxin-induced endothelial fibrosis is still unknown. Transient receptor potential melastatin 7 (TRPM7) is a calcium channel that is expressed in many cell types, including ECs. TRPM7 is involved in a number of crucial processes such as the conversion of fibroblasts into activated fibroblasts, or myofibroblasts, being responsible for the development of several characteristics of them. However, the role of the TRPM7 ion channel in endotoxin-induced endothelial fibrosis is unknown. Thus, our aim was to study whether the TRPM7 calcium channel participates in endotoxin-induced endothelial fibrosis. Using primary cultures of ECs, we demonstrated that TRPM7 is a crucial protein involved in endotoxin-induced endothelial fibrosis. Suppression of TRPM7 expression protected ECs from the fibrogenic process stimulated by endotoxin. Downregulation of TRPM7 prevented the endotoxin-induced endothelial markers decrease and fibrotic genes increase in ECs. In addition, TRPM7 downregulation abolished the endotoxin-induced increase in ECM proteins in ECs. Furthermore, we showed that intracellular Ca2+ levels were greatly increased upon LPS challenge in a mechanism dependent on TRPM7 expression. These results demonstrate that TRPM7 is a key protein involved in the mechanism underlying endotoxin-induced endothelial fibrosis.

  15. Correlated FLIM and PLIM for cell metabolism

    NASA Astrophysics Data System (ADS)

    Rück, A.; Breymayer, J.; Kalinina, S.

    2016-03-01

    Correlated imaging of phosphorescence and fluorescence lifetime parameters of metabolic markers is a challenge for direct investigating mechanisms related to cell metabolism and oxygen tension. A large variety of clinical phenotypes is associated with mitochondrial defects accomplished with changes in cell metabolism. In many cases the hypoxic microenvironment of cancer cells shifts metabolism from oxidative phosphorylation (OXPHOS) to anaerobic or aerobic glycolysis, a process known as "Warburg" effect. Also during stem cell differentiation a switch in cell metabolism is observed. A defective mitochondrial function associated with hypoxia has been invoked in many complex disorders such as type 2 diabetes, Alzheimers disease, cardiac ischemia/reperfusion injury, tissue inflammation and cancer. Cellular responses to oxygen tension have been studied extensively, optical imaging techniques based on time correlated single photon counting (TCSPC) to detect the underlying metabolic mechanisms are therefore of prominent interest. They offer the possibility by inspecting fluorescence decay characteristics of intrinsic coenzymes to directly image metabolic pathways. Moreover oxygen tension can be determined by considering the phosphorescence lifetime of a phosphorescent probe. The combination of both fluorescence lifetime imaging (FLIM) of coenzymes like NADH and FAD and phosphorescence lifetime (PLIM) of phosphorescent dyes could provide valuable information about correlation of metabolic pathways and oxygen tension.

  16. Suppression of renal fibrosis by galectin-1 in high glucose-treated renal epithelial cells

    SciTech Connect

    Okano, Kazuhiro Tsuruta, Yuki; Yamashita, Tetsuri; Takano, Mari; Echida, Yoshihisa; Nitta, Kosaku

    2010-11-15

    Diabetic nephropathy is the most common cause of chronic kidney disease. We investigated the ability of intracellular galectin-1 (Gal-1), a prototype of endogenous lectin, to prevent renal fibrosis by regulating cell signaling under a high glucose (HG) condition. We demonstrated that overexpression of Gal-1 reduces type I collagen (COL1) expression and transcription in human renal epithelial cells under HG conditions and transforming growth factor-{beta}1 (TGF-{beta}1) stimulation. Matrix metalloproteinase 1 (MMP1) is stimulated by Gal-1. HG conditions and TGF-{beta}1 treatment augment expression and nuclear translocation of Gal-1. In contrast, targeted inhibition of Gal-1 expression reduces COL1 expression and increases MMP1 expression. The Smad3 signaling pathway is inhibited, whereas two mitogen-activated protein kinase (MAPK) pathways, p38 and extracellular signal-regulated kinase (ERK), are activated by Gal-1, indicating that Gal-1 regulates these signaling pathways in COL1 production. Using specific inhibitors of Smad3, ERK, and p38 MAPK, we showed that ERK MAPK activated by Gal-1 plays an inhibitory role in COL1 transcription and that activation of the p38 MAPK pathway by Gal-1 plays a negative role in MMP1 production. Taken together, two MAPK pathways are stimulated by increasing levels of Gal-1 in the HG condition, leading to suppression of COL1 expression and increase of MMP1 expression.

  17. Neuroinflammatory Mechanisms of Connective Tissue Fibrosis: Targeting Neurogenic and Mast Cell Contributions

    PubMed Central

    Monument, Michael J.; Hart, David A.; Salo, Paul T.; Befus, A. Dean; Hildebrand, Kevin A.

    2015-01-01

    Significance: The pathogenesis of fibrogenic wound and connective tissue healing is complex and incompletely understood. Common observations across a vast array of human and animal models of fibroproliferative conditions suggest neuroinflammatory mechanisms are important upstream fibrogenic events. Recent Advances: As detailed in this review, mast cell hyperplasia is a common observation in fibrotic tissue. Recent investigations in human and preclinical models of hypertrophic wound healing and post-traumatic joint fibrosis provides evidence that fibrogenesis is governed by a maladaptive neuropeptide-mast cell-myofibroblast signaling pathway. Critical Issues: The blockade and manipulation of these factors is providing promising evidence that if timed correctly, the fibrogenic process can be appropriately regulated. Clinically, abnormal fibrogenic healing responses are not ubiquitous to all patients and the identification of those at-risk remains an area of priority. Future Directions: Ultimately, an integrated appreciation of the common pathobiology shared by many fibrogenic connective tissue conditions may provide a scientific framework to facilitate the development of novel antifibrotic prevention and treatment strategies. PMID:25785237

  18. Cystic fibrosis transmembrane conductance regulator modulates neurosecretory function in pulmonary neuroendocrine cell-related tumor cell line models.

    PubMed

    Pan, Jie; Bear, Christine; Farragher, Susan; Cutz, Ernest; Yeger, Herman

    2002-11-01

    The pulmonary neuroendocrine cell (PNEC) system consists of solitary cells and distinctive cell clusters termed neuroepithelial bodies (NEB) localized in the airway epithelium. PNEC/NEB express a variety of bioactive substances, including amine (serotonin, 5HT) and neuropeptides. We have previously shown that NEB cells are O(2) sensors expressing nicotinamide adenine diphosphate oxidase complex and O(2) sensitive K(+) channel. Recently, we demonstrated expression of functional cystic fibrosis transmembrane conductance regulator (CFTR) and Cl(-) conductances in NEB cells of rabbit neonatal lung. Because PNEC/NEB are sparsely distributed and difficult to study in native lung, we investigated small-cell lung carcinoma (SCLC) and carcinoid tumor cell lines (tumor counterparts of normal PNEC/NEB) as models for PNEC/NEB. SCLC (H146, H345) and carcinoid (H727) cell lines express neuroendocrine cell markers, including chromogranin A, neural cell adhesion molecule (N-CAM), 5HT, and tryptophan hydroxylase. We report that H146, H345, and H727 express CFTR messenger RNA (reverse transcription polymerase chain reaction) and protein (immunoblotting) and possess functional CFTR Cl(-) conductance, demonstrated by an iodide efflux assay inhibitable by transfection with antisense CFTR. Using an immunoassay to quantitate 5HT secretion, we also show that downregulation of CFTR abolishes hypoxia-induced 5HT release, and reduces secretory response to high potassium. Our findings suggest that CFTR may modulate neurosecretory activity of PNEC/NEB possessing O(2) sensor function. We propose that these tumor cell lines may be useful models for investigating the role of CFTR in PNEC/NEB functions in health and disease.

  19. Effect of bosentan is correlated with MMP-9/TIMP-1 ratio in bleomycin-induced pulmonary fibrosis

    PubMed Central

    Zuo, Wan-Li; Zhao, Jie-Min; Huang, Ji-Xiong; Zhou, Wei; Lei, Ze-Hong; Huang, Yan-Ming; Huang, Yan-Fen; Li, Hai-Gang

    2017-01-01

    Pulmonary fibrosis (PF) is a life-threatening non-tumorous disease characterized by progressive fibrosis and worsening lung function. Various drugs, such as bleomycin, can contribute to lung injury and PF, with lung injury potentially occurring in 10% of bleomycin users. Bleomycin is the most commonly used drug in the establishment of an animal model of PF in rats. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) serve an important role in controlling tissue organization and fibrosis following injury. The present study examined the effect of bosentan on fibrotic lung tissue in rats administrated with bleomycin. In total, 48 Wistar rats were administrated with bleomycin, with or without bosentan, while the control rats received saline. The lung tissues were microscopically examined by staining with hematoxylin and eosin and Masson's trichome. ELISA was also used to detect the MMP-9 and TIMP-1 concentrations in the plasma. The results indicated that the bosentan-treated groups on the next day and the 15th day showed significant reversal of pathological findings. In addition, the concentrations of MMP-9 and TIMP-1 appeared to be altered following bosentan treatment, improving the bleomycin-induced PF. Masson's trichome staining showed high collagen deposition in the lung tissue sections, which may be a direct result of the activity of MMP-9 and TIMP-1. Furthermore, the deposition of collagen was significantly inhibited in bosentan-treated groups. In conclusion, these results demonstrated that bosentan inhibited lung fibrosis induced by bleomycin and it may be used as an inhibitor of PF. PMID:28357073

  20. Paracellular transport through healthy and cystic fibrosis bronchial epithelial cell lines--do we have a proper model?

    PubMed

    Molenda, Natalia; Urbanova, Katarina; Weiser, Nelly; Kusche-Vihrog, Kristina; Günzel, Dorothee; Schillers, Hermann

    2014-01-01

    It has been reported recently that the cystic fibrosis transmembrane conductance regulator (CFTR) besides transcellular chloride transport, also controls the paracellular permeability of bronchial epithelium. The aim of this study was to test whether overexpressing wtCFTR solely regulates paracellular permeability of cell monolayers. To answer this question we used a CFBE41o- cell line transfected with wtCFTR or mutant F508del-CFTR and compered them with parental line and healthy 16HBE14o- cells. Transepithelial electrical resistance (TER) and paracellular fluorescein flux were measured under control and CFTR-stimulating conditions. CFTR stimulation significant decreased TER in 16HBE14o- and also in CFBE41o- cells transfected with wtCFTR. In contrast, TER increased upon stimulation in CFBE41o- cells and CFBE41o- cells transfected with F508del-CFTR. Under non-stimulated conditions, all four cell lines had similar paracellular fluorescein flux. Stimulation increased only the paracellular permeability of the 16HBE14o- cell monolayers. We observed that 16HBE14o- cells were significantly smaller and showed a different structure of cell-cell contacts than CFBE41o- and its overexpressing clones. Consequently, 16HBE14o- cells have about 80% more cell-cell contacts through which electrical current and solutes can leak. Also tight junction protein composition is different in 'healthy' 16HBE14o- cells compared to 'cystic fibrosis' CFBE41o- cells. We found that claudin-3 expression was considerably stronger in 16HBE14o- cells than in the three CFBE41o- cell clones and thus independent of the presence of functional CFTR. Together, CFBE41o- cell line transfection with wtCFTR modifies transcellular conductance, but not the paracellular permeability. We conclude that CFTR overexpression is not sufficient to fully reconstitute transport in CF bronchial epithelium. Hence, it is not recommended to use those cell lines to study CFTR-dependent epithelial transport.

  1. Angiogenesis and liver fibrosis

    PubMed Central

    Elpek, Gülsüm Özlem

    2015-01-01

    Recent data indicate that hepatic angiogenesis, regardless of the etiology, takes place in chronic liver diseases (CLDs) that are characterized by inflammation and progressive fibrosis. Because anti-angiogenic therapy has been found to be efficient in the prevention of fibrosis in experimental models of CLDs, it is suggested that blocking angiogenesis could be a promising therapeutic option in patients with advanced fibrosis. Consequently, efforts are being directed to revealing the mechanisms involved in angiogenesis during the progression of liver fibrosis. Literature evidences indicate that hepatic angiogenesis and fibrosis are closely related in both clinical and experimental conditions. Hypoxia is a major inducer of angiogenesis together with inflammation and hepatic stellate cells. These profibrogenic cells stand at the intersection between inflammation, angiogenesis and fibrosis and play also a pivotal role in angiogenesis. This review mainly focuses to give a clear view on the relevant features that communicate angiogenesis with progression of fibrosis in CLDs towards the-end point of cirrhosis that may be translated into future therapies. The pathogenesis of hepatic angiogenesis associated with portal hypertension, viral hepatitis, non-alcoholic fatty liver disease and alcoholic liver disease are also discussed to emphasize the various mechanisms involved in angiogenesis during liver fibrogenesis. PMID:25848465

  2. Andrographolide attenuates skeletal muscle dystrophy in mdx mice and increases efficiency of cell therapy by reducing fibrosis

    PubMed Central

    2014-01-01

    Background Duchenne muscular dystrophy (DMD) is characterized by the absence of the cytoskeletal protein dystrophin, muscle wasting, increased transforming growth factor type beta (TGF-β) signaling, and fibrosis. At the present time, the only clinically validated treatments for DMD are glucocorticoids. These drugs prolong muscle strength and ambulation of patients for a short term only and have severe adverse effects. Andrographolide, a bicyclic diterpenoid lactone, has traditionally been used for the treatment of colds, fever, laryngitis, and other infections with no or minimal side effects. We determined whether andrographolide treatment of mdx mice, an animal model for DMD, affects muscle damage, physiology, fibrosis, and efficiency of cell therapy. Methods mdx mice were treated with andrographolide for three months and skeletal muscle histology, creatine kinase activity, and permeability of muscle fibers were evaluated. Fibrosis and TGF-β signaling were evaluated by indirect immunofluorescence and Western blot analyses. Muscle strength was determined in isolated skeletal muscles and by a running test. Efficiency of cell therapy was determined by grafting isolated skeletal muscle satellite cells onto the tibialis anterior of mdx mice. Results mdx mice treated with andrographolide exhibited less severe muscular dystrophy than untreated dystrophic mice. They performed better in an exercise endurance test and had improved muscle strength in isolated muscles, reduced skeletal muscle impairment, diminished fibrosis and a significant reduction in TGF-β signaling. Moreover, andrographolide treatment of mdx mice improved grafting efficiency upon intramuscular injection of dystrophin-positive satellite cells. Conclusions These results suggest that andrographolide could be used to improve quality of life in individuals with DMD. PMID:24655808

  3. The anti-hepatic fibrosis activity of ergosterol depended on upregulation of PPARgamma in HSC-T6 cells.

    PubMed

    Tai, Chen-Jei; Choong, Chen-Yen; Lin, Yu-Chun; Shi, Yeu-Ching; Tai, Cheng-Jeng

    2016-04-01

    Advanced glycation endproducts (AGEs) were shown to play an important role in metabolic syndrome and were suggested to contribute to the development of hepatic fibrosis. Evidence indicates that AGEs resulted in hepatic fibrosis coupled to the activation of the receptor for AGEs (RAGE) in hepatic stellate cells (HSCs). NADPH oxidase is downstream of the RAGE signaling pathway, resulting in an increase in reactive oxygen species (ROS), alpha-smooth muscle actin (alpha-SMA), RAGE, and matrix metalloproteinase-9 (MMP-9). This study was designed to evaluate the effects of ergosterol on RAGE signaling in HSC-T6 cells. Ergosterol suppressed the activation of HSC-T6 cells induced by AGEs, and attenuated overexpressions of alpha-SMA, MMP-9, and epithelial-mesenchymal transition (EMT) markers, including N-cadherin and vimentin. We also found that these inhibitory effects of ergosterol on the activation of HSCs were dependent on peroxisome proliferator-activated receptor-gamma (PPARgamma) confirmed by PPARgamma reporter assay and PPARgamma knockdown. In addition, ergosterol also showed an inhibitory effect on the generation of AGEs, fructosamine, and α-dicarbonyl compounds in this study. Our results show that ergosterol can be used as a protective agent against hepatic fibrosis caused by induction of AGEs.

  4. Tomatidine inhibits replication of Staphylococcus aureus small-colony variants in cystic fibrosis airway epithelial cells.

    PubMed

    Mitchell, Gabriel; Gattuso, Mariza; Grondin, Gilles; Marsault, Éric; Bouarab, Kamal; Malouin, François

    2011-05-01

    Small-colony variants (SCVs) often are associated with chronic Staphylococcus aureus infections, such as those encountered by cystic fibrosis (CF) patients. We report here that tomatidine, the aglycon form of the plant secondary metabolite tomatine, has a potent growth inhibitory activity against SCVs (MIC of 0.12 μg/ml), whereas the growth of normal S. aureus strains was not significantly altered by tomatidine (MIC, >16 μg/ml). The specific action of tomatidine was bacteriostatic for SCVs and was clearly associated with their dysfunctional electron transport system, as the presence of the electron transport inhibitor 4-hydroxy-2-heptylquinoline-N-oxide (HQNO) caused normal S. aureus strains to become susceptible to tomatidine. Inversely, the complementation of SCVs' respiratory deficiency conferred resistance to tomatidine. Tomatidine provoked a general reduction of macromolecular biosynthesis but more specifically affected the incorporation of radiolabeled leucine in proteins of HQNO-treated S. aureus at a concentration corresponding to the MIC against SCVs. Furthermore, tomatidine inhibited the intracellular replication of a clinical SCV in polarized CF-like epithelial cells. Our results suggest that tomatidine eventually will find some use in combination therapy with other traditional antibiotics to eliminate persistent forms of S. aureus.

  5. Chlorzoxazone or 1-EBIO increases Na(+) absorption across cystic fibrosis airway epithelial cells.

    PubMed

    Gao, L; Yankaskas, J R; Fuller, C M; Sorscher, E J; Matalon, S; Forman, H J; Venglarik, C J

    2001-11-01

    Previous studies demonstrated that chlorzoxazone or 1-ethyl-2-benzimidazolinone (1-EBIO) enhances transepithelial Cl(-) secretion by increasing basolateral K(+) conductance (G(K)) (Singh AK, Devor DC, Gerlach AC, Gondor M, Pilewski JM, and Bridges RJ. J Pharmacol Exp Ther 292: 778-787, 2000). Hence these compounds may be useful to treat cystic fibrosis (CF) airway disease. The goal of the present study was to determine whether chlorzoxazone or 1-EBIO altered ion transport across Delta F508-CF transmembrane conductance regulator homozygous CFT1 airway cells. CFT1 monolayers exhibited a basal short-circuit current that was abolished by apical amiloride (inhibition constant 320 nM) as expected for Na(+) absorption. The addition of chlorzoxazone (400 microM) or 1-EBIO (2 mM) increased the amiloride-sensitive I(sc) approximately 2.5-fold. This overlapping specificity may preclude use of these compounds as CF therapeutics. Assaying for changes in the basolateral G(K) with a K(+) gradient plus the pore-forming antibiotic amphotericin B revealed that chlorzoxazone or 1-EBIO evoked an approximately 10-fold increase in clotrimazole-sensitive G(K). In contrast, chlorzoxazone did not alter epithelial Na(+) channel-mediated currents across basolateral-permeabilized monolayers or in Xenopus oocytes. These data further suggest that alterations in basolateral G(K) alone can modulate epithelial Na(+) transport.

  6. AMPK agonists ameliorate sodium and fluid transport and inflammation in cystic fibrosis airway epithelial cells.

    PubMed

    Myerburg, Michael M; King, J Darwin; Oyster, Nicholas M; Fitch, Adam C; Magill, Amy; Baty, Catherine J; Watkins, Simon C; Kolls, Jay K; Pilewski, Joseph M; Hallows, Kenneth R

    2010-06-01

    The metabolic sensor AMP-activated kinase (AMPK) inhibits both the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) Cl(-) channel and epithelial Na(+) channel (ENaC), and may inhibit secretion of proinflammatory cytokines in epithelia. Here we have tested in primary polarized CF and non-CF human bronchial epithelial (HBE) cells the effects of AMPK activators, metformin and 5-aminoimidazole-4-carboxamide-1-beta-D-riboside (AICAR), on various parameters that contribute to CF lung disease: ENaC-dependent short-circuit currents (I(sc)), airway surface liquid (ASL) height, and proinflammatory cytokine secretion. AMPK activation after overnight treatment with either metformin (2-5 mM) or AICAR (1 mM) substantially inhibited ENaC-dependent I(sc) in both CF and non-CF airway cultures. Live-cell confocal images acquired 60 minutes after apical addition of Texas Red-dextran-containing fluid revealed significantly greater ASL heights after AICAR and metformin treatment relative to controls, suggesting that AMPK-dependent ENaC inhibition slows apical fluid reabsorption. Both metformin and AICAR decreased secretion of various proinflammatory cytokines, both with and without prior LPS stimulation. Finally, prolonged exposure to more physiologically relevant concentrations of metformin (0.03-1 mM) inhibited ENaC currents and decreased proinflammatory cytokine levels in CF HBE cells in a dose-dependent manner. These findings suggest that novel therapies to activate AMPK in the CF airway may be beneficial by blunting excessive sodium and ASL absorption and by reducing excessive airway inflammation, which are major contributors to CF lung disease.

  7. AMPK Agonists Ameliorate Sodium and Fluid Transport and Inflammation in Cystic Fibrosis Airway Epithelial Cells

    PubMed Central

    Myerburg, Michael M.; King, J Darwin; Oyster, Nicholas M.; Fitch, Adam C.; Magill, Amy; Baty, Catherine J.; Watkins, Simon C.; Kolls, Jay K.; Pilewski, Joseph M.; Hallows, Kenneth R.

    2010-01-01

    The metabolic sensor AMP-activated kinase (AMPK) inhibits both the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) Cl− channel and epithelial Na+ channel (ENaC), and may inhibit secretion of proinflammatory cytokines in epithelia. Here we have tested in primary polarized CF and non-CF human bronchial epithelial (HBE) cells the effects of AMPK activators, metformin and 5-aminoimidazole-4-carboxamide-1-β-D-riboside (AICAR), on various parameters that contribute to CF lung disease: ENaC-dependent short-circuit currents (Isc), airway surface liquid (ASL) height, and proinflammatory cytokine secretion. AMPK activation after overnight treatment with either metformin (2–5 mM) or AICAR (1 mM) substantially inhibited ENaC-dependent Isc in both CF and non-CF airway cultures. Live-cell confocal images acquired 60 minutes after apical addition of Texas Red–dextran-containing fluid revealed significantly greater ASL heights after AICAR and metformin treatment relative to controls, suggesting that AMPK-dependent ENaC inhibition slows apical fluid reabsorption. Both metformin and AICAR decreased secretion of various proinflammatory cytokines, both with and without prior LPS stimulation. Finally, prolonged exposure to more physiologically relevant concentrations of metformin (0.03–1 mM) inhibited ENaC currents and decreased proinflammatory cytokine levels in CF HBE cells in a dose-dependent manner. These findings suggest that novel therapies to activate AMPK in the CF airway may be beneficial by blunting excessive sodium and ASL absorption and by reducing excessive airway inflammation, which are major contributors to CF lung disease. PMID:19617399

  8. Phase correlation imaging of unlabeled cell dynamics

    PubMed Central

    Ma, Lihong; Rajshekhar, Gannavarpu; Wang, Ru; Bhaduri, Basanta; Sridharan, Shamira; Mir, Mustafa; Chakraborty, Arindam; Iyer, Rajashekar; Prasanth, Supriya; Millet, Larry; Gillette, Martha U.; Popescu, Gabriel

    2016-01-01

    We present phase correlation imaging (PCI) as a novel approach to study cell dynamics in a spatially-resolved manner. PCI relies on quantitative phase imaging time-lapse data and, as such, functions in label-free mode, without the limitations associated with exogenous markers. The correlation time map outputted in PCI informs on the dynamics of the intracellular mass transport. Specifically, we show that PCI can extract quantitatively the diffusion coefficient map associated with live cells, as well as standard Brownian particles. Due to its high sensitivity to mass transport, PCI can be applied to studying the integrity of actin polymerization dynamics. Our results indicate that the cyto-D treatment blocking the actin polymerization has a dominant effect at the large spatial scales, in the region surrounding the cell. We found that PCI can distinguish between senescent and quiescent cells, which is extremely difficult without using specific markers currently. We anticipate that PCI will be used alongside established, fluorescence-based techniques to enable valuable new studies of cell function. PMID:27615512

  9. Respiratory epithelial cell expression of human transforming growth factor-alpha induces lung fibrosis in transgenic mice.

    PubMed Central

    Korfhagen, T R; Swantz, R J; Wert, S E; McCarty, J M; Kerlakian, C B; Glasser, S W; Whitsett, J A

    1994-01-01

    Increased production of EGF or TGF-alpha by the respiratory epithelial cells has been associated with the pathogenesis of various forms of lung injury. Growth factors and cytokines are thought to act locally, via paracrine and autocrine mechanisms, to stimulate cell proliferation and matrix deposition by interstitial lung cells resulting in pulmonary fibrosis. To test whether TGF-alpha mediates pulmonary fibrotic responses, we have generated transgenic mice expressing human TGF-alpha under control of regulatory regions of the human surfactant protein C (SP-C) gene. Human TGF-alpha mRNA was expressed in pulmonary epithelial cells in the lungs of the transgenic mice. Adult mice bearing the SP-C-TGF-alpha transgene developed severe pulmonary fibrosis. Fibrotic lesions were observed in peribronchial, peribronchiolar, and perivascular regions, as well as subjacent to pleural surfaces. Lesions consisted of fibrous tissue that included groups of epithelial cells expressing endogenous SP-C mRNA, consistent with their identification as distal respiratory epithelial cells. Peripheral fibrotic regions consisted of thickened pleura associated with extensive collagen deposition. Alveolar architecture was disrupted in the transgenic mice with loss of alveoli in the lung parenchyma. Pulmonary epithelial cell expression of TGF-alpha in transgenic mice disrupts alveolar morphogenesis and produces fibrotic lesions mediated by paracrine signaling between respiratory epithelial and interstitial cells of the lung. Images PMID:8163670

  10. Squamous cell carcinoma of the oral mucosa after release of submucous fibrosis and bilateral small radial forearm flap reconstruction.

    PubMed

    Celik, Naci; Wei, Fu-chan; Chang, Yang-ming; Yang, Wen-guei; Chen, Da-jeng; Tsai, Chi-ying

    2002-07-01

    Oral submucous fibrosis is a collagen disorder that affects the submucosal layer of the upper digestive tract. The major cause is the habit of betel quid chewing, which is common in central, southern, and southeast Asia. The progressive and irreversible course of disease results with trismus, dysphagia, xerostomia, and rhinolalia. The most serious complication of this disorder is the development of oral carcinoma, and the incidence in different series varies from 1.9 to 10 percent. A sufficient mouth opening can be achieved by complete release of fibrotic tissue, and coronoidectomy and temporal muscle myotomy when needed, and reconstruction of the resultant defect can be best achieved by microsurgical free-tissue transfer because of the discouraging results with skin grafting or local flaps. From April of 1997 to May of 2001, a total of 26 patients received reconstructive surgery with small radial forearm flaps after release of submucous fibrosis with or without temporalis muscle myotomy and coronoidectomy. All patients were men, with a mean age of 40.1 years (range, 18 to 62 years) and all had a history of betel nut chewing ranging from 8 to 40 years. The interincisal distance ranged from 5 to 29 mm, with a mean of 15 mm, before operation. After the release procedure, the interincisal distance increased to 40 mm (range, 35 to 50 mm). At a follow-up period of 3 to 48 months, the interincisal distance was a mean of 35 mm (range, 18 to 57 mm), with an average increase of 20 mm compared with the preoperative distance. During follow-up, three patients developed squamous cell carcinoma of the oral cavity 24 to 36 months after submucous fibrosis release. Two of them occurred in the release site and the other one occurred at the soft palate. Oral cancer occurred in three of 13 patients who had received release of submucous fibrosis and who were followed for longer than 2 years (range, 24 to 48 months), which means that 23 percent of these patients developed squamous cell

  11. Deficiency of NOX1 or NOX4 Prevents Liver Inflammation and Fibrosis in Mice through Inhibition of Hepatic Stellate Cell Activation

    PubMed Central

    Lan, Tian; Kisseleva, Tatiana; Brenner, David A.

    2015-01-01

    Reactive oxygen species (ROS) produced by nicotinamide adenine dinucleotide phosphate oxidase (NOX) play a key role in liver injury and fibrosis. Previous studies demonstrated that GKT137831, a dual NOX1/4 inhibitor, attenuated liver fibrosis in mice as well as pro-fibrotic genes in hepatic stellate cells (HSCs) as well as hepatocyte apoptosis. The effect of NOX1 and NOX4 deficiency in liver fibrosis is unclear, and has never been directly compared. HSCs are the primary myofibroblasts in the pathogenesis of liver fibrosis. Therefore, we aimed to determine the role of NOX1 and NOX4 in liver fibrosis, and investigated whether NOX1 and NOX4 signaling mediates liver fibrosis by regulating HSC activation. Mice were treated with carbon tetrachloride (CCl4) to induce liver fibrosis. Deficiency of either NOX1 or NOX4 attenuates liver injury, inflammation, and fibrosis after CCl4 compared to wild-type mice. NOX1 or NOX4 deficiency reduced lipid peroxidation and ROS production in mice with liver fibrosis. NOX1 and NOX4 deficiency are approximately equally effective in preventing liver injury in the mice. The NOX1/4 dual inhibitor GKT137831 suppressed ROS production as well as inflammatory and proliferative genes induced by lipopolysaccharide (LPS), platelet-derived growth factor (PDGF), or sonic hedgehog (Shh) in primary mouse HSCs. Furthermore, the mRNAs of proliferative and pro-fibrotic genes were downregulated in NOX1 and NOX4 knock-out activated HSCs (cultured on plastic for 5 days). Finally, NOX1 and NOX4 protein levels were increased in human livers with cirrhosis compared with normal controls. Thus, NOX1 and NOX4 signaling mediates the pathogenesis of liver fibrosis, including the direct activation of HSC. PMID:26222337

  12. Genomic Analysis of Kidney Allograft Injury Identifies Hematopoietic Cell Kinase as a Key Driver of Renal Fibrosis.

    PubMed

    Wei, Chengguo; Li, Li; Menon, Madhav C; Zhang, Weijia; Fu, Jia; Kidd, Brian; Keung, Karen L; Woytovich, Christopher; Greene, Ilana; Xiao, Wenzhen; Salem, Fadi; Yi, Zhengzi; He, John Cijiang; Dudley, Joel T; Murphy, Barbara

    2016-12-07

    Renal fibrosis is the common pathway of progression for patients with CKD and chronic renal allograft injury (CAI), but the underlying mechanisms remain obscure. We performed a meta-analysis in human kidney biopsy specimens with CAI, incorporating data available publicly and from our Genomics of Chronic Renal Allograft Rejection study. We identified an Src family tyrosine kinase, hematopoietic cell kinase (Hck), as upregulated in allografts in CAI. Querying the Kinase Inhibitor Resource database revealed that dasatinib, a Food and Drug Administration-approved drug, potently binds Hck with high selectivity. In vitro, Hck overexpression activated the TGF-β/Smad3 pathway, whereas HCK knockdown inhibited it. Treatment of tubular cells with dasatinib reduced the expression of Col1a1 Dasatinib also reduced proliferation and α-SMA expression in fibroblasts. In a murine model with unilateral ureteric obstruction, pretreatment with dasatinib significantly reduced the upregulation of profibrotic markers, phosphorylation of Smad3, and renal fibrosis observed in kidneys pretreated with vehicle alone. Dasatinib treatment also improved renal function, reduced albuminuria, and inhibited expression of profibrotic markers in animal models with lupus nephritis and folic acid nephropathy. These data suggest that Hck is a key mediator of renal fibrosis and dasatinib could be developed as an antifibrotic drug.

  13. Activated hepatic stellate cells impair NK cell anti-fibrosis capacity through a TGF-β-dependent emperipolesis in HBV cirrhotic patients.

    PubMed

    Shi, Jijing; Zhao, Juanjuan; Zhang, Xin; Cheng, Yongqian; Hu, Jinhua; Li, Yuanyuan; Zhao, Xin; Shang, Qinghua; Sun, Yanling; Tu, Bo; Shi, Lei; Gao, Bin; Wang, Fu-Sheng; Zhang, Zheng

    2017-03-14

    Natural killer (NK) cells can induce liver fibrosis remission by killing hepatic stellate cells (HSCs) and producing interferon (IFN)-γ in a mouse model; however, their anti-fibrotic immune-characteristics and regulatory mechanisms by HSCs remain to be determined, especially in livers from HBV-infected liver cirrhosis (LC) patients. We analyzed frequency, phenotype and anti-fibrotic function of hepatic and peripheral NK subsets in 43 HBV-LC patients. We found that hepatic NK subsets from LC patients displayed a decreased frequency, activation status and anti-fibrotic activity compared with those from chronic hepatitis B patients, which were mainly mediated by increased intrahepatic tumour-growth factor (TGF)-β because blockade of TGF-β significantly reversed NK anti-fibrotic function in vitro. In vivo, hepatic NK cells were enriched in proximity to the α-smooth muscle actin (α-SMA+) area within mild fibrosis regions; while in severe fibrotic areas, they were either directly attached to or separated from the α-SMA+ region. NK cells from LC patients could enter HSCs to form emperipolesis (a cell-in-cell structure) and become apoptotic; anti-TGF-β treatment ameliorated this emperipolesis. This finding suggested a novel mechanism by which activated HSCs impair NK cells' anti-fibrosis capacity through a TGF-β-dependent emperipolesis in LC patients, providing an anti-fibrotic rational by enhancing NK cell activity.

  14. Orphan nuclear receptor NR4A2 inhibits hepatic stellate cell proliferation through MAPK pathway in liver fibrosis.

    PubMed

    Chen, Pengguo; Li, Jie; Huo, Yan; Lu, Jin; Wan, Lili; Li, Bin; Gan, Run; Guo, Cheng

    2015-01-01

    Hepatic stellate cells (HSCs) play a crucial role in liver fibrosis, which is a pathological process characterized by extracellular matrix accumulation. NR4A2 is a nuclear receptor belonging to the NR4A subfamily and vital in regulating cell growth, metabolism, inflammation and other biological functions. However, its role in HSCs is unclear. We analyzed NR4A2 expression in fibrotic liver and stimulated HSCs compared with control group and studied the influence on cell proliferation, cell cycle, cell apoptosis and MAPK pathway after NR4A2 knockdown. NR4A2 expression was examined by real-time polymerase chain reaction, Western blotting, immunohistochemistry and immunofluorescence analyses. NR4A2 expression was significantly lower in fibrotic liver tissues and PDGF BB or TGF-β stimulated HSCs compared with control group. After NR4A2 knockdown α-smooth muscle actin and Col1 expression increased. In addition, NR4A2 silencing led to the promotion of cell proliferation, increase of cell percentage in S phase and reduced phosphorylation of ERK1/2, P38 and JNK in HSCs. These results indicate that NR4A2 can inhibit HSC proliferation through MAPK pathway and decrease extracellular matrix in liver fibrogenesis. NR4A2 may be a promising therapeutic target for liver fibrosis.

  15. Elevated furin levels in human cystic fibrosis cells result in hypersusceptibility to exotoxin A–induced cytotoxicity

    PubMed Central

    Ornatowski, Wojciech; Poschet, Jens F.; Perkett, Elizabeth; Taylor-Cousar, Jennifer L.; Deretic, Vojo

    2007-01-01

    Progressive pulmonary disease and infections with Pseudomonas aeruginosa remain an intractable problem in cystic fibrosis (CF). At the cellular level, CF is characterized by organellar hyperacidification, which results in altered protein and lipid glycosylation. Altered pH of the trans-Golgi network (TGN) may further disrupt the protein processing and packaging that occurs in this organelle. Here we measured activity of the major TGN endoprotease furin and demonstrated a marked upregulation in human CF cells. Increased furin activity was linked to elevated production in CF of the immunosuppressive and tissue remodeling cytokine TGF-β and its downstream effects, including macrophage deactivation and augmented collagen secretion by epithelial cells. As furin is responsible for the proteolytic processing of a range of endogenous and exogenous substrates including growth factors and bacterial toxins, we determined that elevated furin-dependent activation of exotoxin A caused increased cell death in CF respiratory epithelial cells compared with genetically matched CF transmembrane conductance regulator–corrected cells. Thus elevated furin levels in CF respiratory epithelial cells contributes to bacterial toxin–induced cell death, fibrosis, and local immunosuppression. These data suggest that the use of furin inhibitors may represent a strategy for pharmacotherapy in CF. PMID:17948127

  16. Human Adipose-derived Mesenchymal Stem Cells Attenuate Early Stage of Bleomycin Induced Pulmonary Fibrosis: Comparison with Pirfenidone

    PubMed Central

    Reddy, Manoj; Fonseca, Lyle; Gowda, Shashank; Chougule, Basavraj; Hari, Aarya; Totey, Satish

    2016-01-01

    Background and Objectives Idiopathic pulmonary fibrosis (IPF) is a progressive, irreversible, invariably fatal fibrotic lung disease with no lasting option for therapy. Mesenchymal stem cells (MSCs) could be a promising modality for the treatment of IPF. Aim of the study was to investigate improvement in survivability and anti-fibrotic efficacy of human adipose-derived mesenchymal stem cells (AD-MSCs) in comparison with pirfenidone in the bleomycin-induced pulmonary fibrosis model. Methods Human AD-MSCs were administered intravenously on day 3, 6 and 9 after an intra-tracheal challenge with bleomycin, whereas, pirfenidone was given orally in drinking water at the rate of 100 mg/kg body weight three times a day daily from day 3 onward. AD-MSCs were labelled with PKH-67 before administration to detect engraftment. Disease severity and improvement was assessed and compared between sham control and vehicle control groups using Kaplan-Meier survival analysis, biochemical and molecular analysis, histopathology and high resolution computed tomography (HRCT) parameters at the end of study. Results Results demonstrated that AD-MSCs significantly increase survivability; reduce organ weight and collagen deposition better than pirfenidone group. Histological analyses and HRCT of the lung revealed that AD-MSCs afforded protection against bleomycin induced fibrosis and protect architecture of the lung. Gene expression analysis revealed that AD-MSCs potently suppressed pro-fibrotic genes induced by bleomycin. More importantly, AD-MSCs were found to inhibit pro-inflammatory related transcripts. Conclusions Our results provided direct evidence that AD-MSC-mediated immunomodulation and anti-fibrotic effect in the lungs resulted in marked protection in pulmonary fibrosis, but at an early stage of disease. PMID:27871152

  17. Activated hepatic stellate cells impair NK cell anti-fibrosis capacity through a TGF-β-dependent emperipolesis in HBV cirrhotic patients

    PubMed Central

    Shi, Jijing; Zhao, Juanjuan; Zhang, Xin; Cheng, Yongqian; Hu, Jinhua; Li, Yuanyuan; Zhao, Xin; Shang, Qinghua; Sun, Yanling; Tu, Bo; Shi, Lei; Gao, Bin; Wang, Fu-Sheng; Zhang, Zheng

    2017-01-01

    Natural killer (NK) cells can induce liver fibrosis remission by killing hepatic stellate cells (HSCs) and producing interferon (IFN)-γ in a mouse model; however, their anti-fibrotic immune-characteristics and regulatory mechanisms by HSCs remain to be determined, especially in livers from HBV-infected liver cirrhosis (LC) patients. We analyzed frequency, phenotype and anti-fibrotic function of hepatic and peripheral NK subsets in 43 HBV-LC patients. We found that hepatic NK subsets from LC patients displayed a decreased frequency, activation status and anti-fibrotic activity compared with those from chronic hepatitis B patients, which were mainly mediated by increased intrahepatic tumour-growth factor (TGF)-β because blockade of TGF-β significantly reversed NK anti-fibrotic function in vitro. In vivo, hepatic NK cells were enriched in proximity to the α-smooth muscle actin (α-SMA+) area within mild fibrosis regions; while in severe fibrotic areas, they were either directly attached to or separated from the α-SMA+ region. NK cells from LC patients could enter HSCs to form emperipolesis (a cell-in-cell structure) and become apoptotic; anti-TGF-β treatment ameliorated this emperipolesis. This finding suggested a novel mechanism by which activated HSCs impair NK cells’ anti-fibrosis capacity through a TGF-β-dependent emperipolesis in LC patients, providing an anti-fibrotic rational by enhancing NK cell activity. PMID:28291251

  18. PPARγ activation following apoptotic cell instillation promotes resolution of lung inflammation and fibrosis via regulation of efferocytosis and proresolving cytokines

    PubMed Central

    Yoon, Y-S; Kim, S-Y; Kim, M-J; Lim, J-H; Cho, M-S; Kang, J L

    2015-01-01

    Changes in macrophage phenotype have been implicated in apoptotic cell-mediated immune modulation via induction of peroxisome proliferator-activated receptor-γ (PPARγ). In this study, we characterized PPARγ induction by apoptotic cell instillation over the course of bleomycin-induced lung injury in C57BL/6 mice. Next, the role of PPARγ activation in resolving lung inflammation and fibrosis was investigated. Our data demonstrate that apoptotic cell instillation after bleomycin results in immediate and prolonged enhancement of PPARγ mRNA and protein in alveolar macrophages and lung. Moreover, PPARγ activity and expression of its target molecules, including CD36, macrophage mannose receptor, and arginase 1, were persistently enhanced following apoptotic cell instillation. Coadministration of the PPARγ antagonist, GW9662, reversed the enhanced efferocytosis, and the reduced proinflammatory cytokine expression, neutrophil recruitment, myeloperoxidase activity, hydroxyproline contents, and fibrosis markers, including type 1 collagen α2, fibronectin and α-smooth muscle actin (α-SMA), in the lung by apoptotic cell instillation. In addition, inhibition of PPARγ activity reversed the expression of transforming growth factor-β (TGF-β), interleukin (IL)-10, and hepatocyte growth factor (HGF). These findings indicate that one-time apoptotic cell instillation contributes to anti-inflammatory and antifibrotic responses via upregulation of PPARγ expression and subsequent activation, leading to regulation of efferocytosis and production of proresolving cytokines. PMID:25586556

  19. Abnormal ion content, hydration and granule expansion of the secretory granules from cystic fibrosis airway glandular cells

    SciTech Connect

    Baconnais, S.; Delavoie, F. |; Zahm, J.M.; Milliot, M.; Castillon, N.; Terryn, C.; Banchet, V.; Michel, J.; Danos, O.; Merten, M.; Chinet, T.; Zierold, K.; Bonnet, N.; Puchelle, E. , E-Mail: edith.puchelle@univ-reims.fr; Balossier, G.

    2005-10-01

    The absence or decreased expression of cystic fibrosis transmembrane conductance regulator (CFTR) induces increased Na{sup +} absorption and hyperabsorption of the airway surface liquid (ASL) resulting in a dehydrated and hyperviscous ASL. Although the implication of abnormal airway submucosal gland function has been suggested, the ion and water content in the Cystic Fibrosis (CF) glandular secretory granules, before exocytosis, is unknown. We analyzed, in non-CF and CF human airway glandular cell lines (MM-39 and KM4, respectively), the ion content in the secretory granules by electron probe X-ray microanalysis and the water content by quantitative dark field imaging on freeze-dried cryosections. We demonstrated that the ion content (Na{sup +}, Mg{sup 2+}, P, S and Cl{sup -}) is significantly higher and the water content significantly lower in secretory granules from the CF cell line compared to the non-CF cell line. Using videomicroscopy, we observed that the secretory granule expansion was deficient in CF glandular cells. Transfection of CF cells with CFTR cDNA or inhibition of non-CF cells with CFTR{sub inh}-172, respectively restored or decreased the water content and granule expansion, in parallel with changes in ion content. We hypothesize that the decreased water and increased ion content in glandular secretory granules may contribute to the dehydration and increased viscosity of the ASL in CF.

  20. Tonsil-derived mesenchymal stem cells ameliorate CCl4-induced liver fibrosis in mice via autophagy activation.

    PubMed

    Park, Minhwa; Kim, Yu-Hee; Woo, So-Youn; Lee, Hye Jin; Yu, Yeonsil; Kim, Han Su; Park, Yoon Shin; Jo, Inho; Park, Joo-Won; Jung, Sung-Chul; Lee, Hyukjin; Jeong, Byeongmoon; Ryu, Kyung-Ha

    2015-02-27

    Liver transplantation is the treatment of choice for chronic liver failure, although it is complicated by donor shortage, surgery-related complications, and immunological rejection. Cell transplantation is an alternative, minimally invasive treatment option with potentially fewer complications. We used human palatine tonsil as a novel source of mesenchymal stem cells (T-MSCs) and examined their ability to differentiate into hepatocyte-like cells in vivo and in vitro. Carbon tetrachloride (CCl4) mouse model was used to investigate the ability of T-MSCs to home to the site of liver injury. T-MSCs were only detected in the damaged liver, suggesting that they are disease-responsive. Differentiation of T-MSCs into hepatocyte-like cells was confirmed in vitro as determined by expression of hepatocyte markers. Next, we showed resolution of liver fibrosis by T-MSCs via reduction of TGF-β expression and collagen deposition in the liver. We hypothesized that autophagy activation was a possible mechanism for T-MSC-mediated liver recovery. In this report, we demonstrate for the first time that T-MSCs can differentiate into hepatocyte-like cells and ameliorate liver fibrosis via autophagy activation and down-regulation of TGF-β. These findings suggest that T-MSCs could be used as a novel source for stem cell therapy targeting liver diseases.

  1. Analysis of correlation between the process of thyroid fibrosis and TGFB1 gene expression level in fine-needle aspiration biopsy (FNAB) thyroid specimens collected from patients with Hashimoto's thyroiditis and non-toxic goitre.

    PubMed

    Cyniak-Magierska, A; Januszkiewicz-Caulier, J; Brzeziańska, E; Lewiński, A

    2010-07-01

    Transforming growth factor beta 1 (TGFB1) stimulates the production of various extracellular matrix components; at the same time, it inhibits matrix degradation. These actions of TGFB1 contribute to tissue repair, however, an altered expression of TGFB1 can be a causative factor of fibrosis processes, including thyroid fibrosis which follows chronic thyroiditis. The aim of our study was to examine a potential correlation between TGFB1 gene expression level in fine-needle aspiration biopsy (FNAB) thyroid specimens and fibrosis of the thyroid gland in two types of thyroid lesions. Fibrosis of the thyroid tissue was assessed, based on the expression levels of fibrosis-associated genes (COL1A1 and COL3A1) in thyroid FNAB samples, on the FNAB specimen cellularity and other features of the tissue fibrosis assessed during cytological examination, as well as on the size of thyroid gland and its function. Following routine cytological examination, 63 thyroid FNAB specimens, received from patients with Hashimoto's thyroiditis (HT, n=30) and non-toxic goitre (NTG, n=33), were quantitatively evaluated regarding TGFB1, COL1A1 and COL3A1 expression level by real-time PCR in the ABI PRISM 7500 Sequence Detection System. The obtained results showed statistically significant differences regarding the expression level (RQ) of TGFB1 and of COL1A1 genes between the groups with HT and with NTG (higher expression in HT group). No significant differences, concerning the expression level of COL3A1 gene, were observed for the studied groups (HT vs. NTG). In HT group statistically significant correlation was found between TGFB1 gene and COL3A1 gene expression levels (p<0.05). The correlation in question might suggest excessive extracellular matrix deposition and could--possibly--contribute to thyroid fibrosis mechanism in the course of chronic thyroiditis.

  2. IL-33 promotes ST2-dependent lung fibrosis by the induction of alternatively activated macrophages and innate lymphoid cells in mice

    PubMed Central

    Li, Dong; Guabiraba, Rodrigo; Besnard, Anne-Gaëlle; Komai-Koma, Mousa; Jabir, Majid S.; Zhang, Li; Graham, Gerard J.; Kurowska-Stolarska, Mariola; Liew, Foo Y.; McSharry, Charles; Xu, Damo

    2014-01-01

    Background The initiation and regulation of pulmonary fibrosis are not well understood. IL-33, an important cytokine for respiratory diseases, is overexpressed in the lungs of patients with idiopathic pulmonary fibrosis. Objectives We aimed to determine the effects and mechanism of IL-33 on the development and severity of pulmonary fibrosis in murine bleomycin-induced fibrosis. Methods Lung fibrosis was induced by bleomycin in wild-type or Il33r (St2)−/− C57BL/6 mice treated with the recombinant mature form of IL-33 or anti–IL-33 antibody or transferred with type 2 innate lymphoid cells (ILC2s). The development and severity of fibrosis was evaluated based on lung histology, collagen levels, and lavage cytology. Cytokine and chemokine levels were quantified by using quantitative PCR, ELISA, and cytometry. Results IL-33 is constitutively expressed in lung epithelial cells but is induced in macrophages by bleomycin. Bleomycin enhanced the production of the mature but reduced full-length form of IL-33 in lung tissue. ST2 deficiency, anti–IL-33 antibody treatment, or alveolar macrophage depletion attenuated and exogenous IL-33 or adoptive transfer of ILC2s enhanced bleomycin-induced lung inflammation and fibrosis. These pathologic changes were accompanied, respectively, by reduced or increased IL-33, IL-13, TGF-β1, and inflammatory chemokine production in the lung. Furthermore, IL-33 polarized M2 macrophages to produce IL-13 and TGF-β1 and induced the expansion of ILC2s to produce IL-13 in vitro and in vivo. Conclusions IL-33 is a novel profibrogenic cytokine that signals through ST2 to promote the initiation and progression of pulmonary fibrosis by recruiting and directing inflammatory cell function and enhancing profibrogenic cytokine production in an ST2- and macrophage-dependent manner. PMID:24985397

  3. The herbal compound Songyou Yin (SYY) inhibits hepatocellular carcinoma growth and improves survival in models of chronic fibrosis via paracrine inhibition of activated hepatic stellate cells

    PubMed Central

    Xue, Tong-Chun; Zhang, Quan-Bao; Zhang, Ke-Zhi; Zhang, Qiang-Bo; You, Yang; Tian, Hui; Qin, Lun-Xiu; Tang, Zhao-You

    2015-01-01

    Chronic fibrosis is a major risk factor for the development of hepatocellular carcinoma (HCC). The pathological progression of hepatic fibrosis has been linked to cellular processes that promote tumor growth and metastasis. Several recent studies have highlighted the cross-talk between tumor cells and activated hepatic stellate cells (aHSCs) in HCC. The herbal compound Songyou Yin (SYY) is known to attenuate hepatoma cell invasion and metastasis via down-regulation of cytokine secretion by aHSCs. However the underlying mechanism of SYY treatment in reversal of hepatic fibrosis and metastasis of liver cancers is not known. In the current study, a nude mouse model with liver fibrosis bearing orthotopic xenograft was established and we found that SYY could reduce associated fibrosis, inhibit tumor growth and improve survival. In the subcutaneous tumor model with fibrosis, we found that SYY could inhibit liver cancer. In vitro, hepatoma cells incubated with conditioned media (CM) from SYY treated aHSCs showed reduced proliferation, decrease in colony formation and invasive potential. SYY treated group showed altered gene expression, with 1205 genes up-regulated and 1323 genes down-regulated. Gene cluster analysis indicated that phosphatidylinositol-3-kinase (PI3K) was one of the key genes altered in the expression profiles. PI3K related markers were all significantly down-regulated. ELISA also indicated decreased secretion of cytokines which were regulated by PI3K/AKT signaling after SYY treatment in the hepatic stellate cell line, LX2. These data clearly demonstrate that SYY therapy inhibits HCC invasive and metastatic potential and improves survival in nude mice models with chronic fibrosis background via inhibition of cytokine secretion by activated hepatic stellate cells. PMID:26517671

  4. Implication of the satellite cell in dystrophic muscle fibrosis: a self-perpetuating mechanism of collagen overproduction.

    PubMed

    Alexakis, Catherine; Partridge, Terence; Bou-Gharios, George

    2007-08-01

    Because of its mechanical function, skeletal muscle is heavily influenced by the composition of its extracellular matrix (ECM). Fibrosis generated by chronic damage, such as occurs in muscular dystrophies, is thus particularly disastrous in this tissue. Here, we examined the interrelationship between the muscle satellite cell and the production of collagen type I, a major component of fibrotic ECM, by using both C2C12, a satellite cell-derived cell line, and primary muscle satellite cells. In C2C12 cells, we found that expression of collagen type I mRNA decreases substantially during skeletal muscle differentiation. On a single-cell level, collagen type I and myogenin became mutually exclusive after 3 days in differentiation medium, whereas addition of collagen markedly suppressed differentiation of C2C12 cells. Primary cultures of satellite cells associated with isolated single fibers of the young (4 wk old) mdx dystrophic mouse and of C57BL/10ScSn wild-type controls expressed collagen type I and type III mRNA and protein. This pattern persisted in wild-type mice at all ages. But, curiously, in older (18-mo-old) mdx mice, although the myogenic cells continued to express type III collagen, type I expression became restricted to nonmyogenic cells. These cells typically constituted part of a cellular sheet surrounding the old mdx fibers. This combination of features strongly suggests that the progression to fibrosis in dystrophic muscle involves changes in the mechanisms controlling matrix production, which generates positive feedback that results in a reprogramming of myoblasts to a profibrotic function.

  5. Correlation of Cell and Substrate Mechanical Properties

    NASA Astrophysics Data System (ADS)

    Setton, Tedhar; Levine, Joshua; Levine, Joseph; Guan, E.; Collazo, Lourdes; Ge, Shouren; Entcheva, Emilia; Rafailovich, Miriam

    2003-03-01

    The mechanical properties of neonatal rat ventricular fibroblasts plated onto elastomer surfaces were studied in vitro and correlated to the mechanical response of the substrate. In order to differentiate the response of the cells to mechanical as opposed to mechanical modifications in their environment, only the rheological properties of the substrates were modified. In the case of entangled polymers this can be accomplished either by varying the molecular weight or the thickness of polymer films spun cast onto rigid supports. Scanning lateral force microscopy, which has been shown to be an effective technique for measuring relative modulii of surfaces(1) was used to track the mechanical response of the substrates as a function of processing procedures, molecular weight, both in liquid, air, and following fibronectin incubation. The response of the living cells was then compared to that of the underlying substrate. The samples were then stained and the distribution of actin correlated to the mechanical response. 1. S. Ge et al. Phys. Rev. Lett. 11, (2000)2340

  6. Role of IL-10-producing regulatory B cells in modulating T-helper cell immune responses during silica-induced lung inflammation and fibrosis

    PubMed Central

    Liu, Fangwei; Dai, Wujing; Li, Chao; Lu, Xiaowei; Chen, Ying; Weng, Dong; Chen, Jie

    2016-01-01

    Silicosis is characterized by chronic lung inflammation and fibrosis, which are seriously harmful to human health. Previous research demonstrated that uncontrolled T-helper (Th) cell immune responses were involved in the pathogenesis of silicosis. Lymphocytes also are reported to have important roles. Existing studies on lymphocyte regulation of Th immune responses were limited to T cells, such as the regulatory T (Treg) cell, which could negatively regulate inflammation and promote the process of silicosis. However, other regulatory subsets in silicosis have not been investigated in detail, and the mechanism of immune homeostasis modulation needs further exploration. Another regulatory lymphocyte, the regulatory B cell, has recently drawn increasing attention. In this study, we comprehensively showed the role of IL-10-producing regulatory B cell (B10) in a silicosis model of mice. B10 was inducible by silica instillation. Insufficient B10 amplified inflammation and attenuated lung fibrosis by promoting the Th1 immune response. Insufficient B10 clearly inhibited Treg and decreased the level of IL-10. Our study indicated that B10 could control lung inflammation and exacerbate lung fibrosis by inhibiting Th1 response and modulating the Th balance. The regulatory function of B10 could be associated with Treg induction and IL-10 secretion. PMID:27354007

  7. Experimental models of liver fibrosis.

    PubMed

    Crespo Yanguas, Sara; Cogliati, Bruno; Willebrords, Joost; Maes, Michaël; Colle, Isabelle; van den Bossche, Bert; de Oliveira, Claudia Pinto Marques Souza; Andraus, Wellington; Alves, Venâncio Avancini; Leclercq, Isabelle; Vinken, Mathieu

    2016-05-01

    Hepatic fibrosis is a wound healing response to insults and as such affects the entire world population. In industrialized countries, the main causes of liver fibrosis include alcohol abuse, chronic hepatitis virus infection and non-alcoholic steatohepatitis. A central event in liver fibrosis is the activation of hepatic stellate cells, which is triggered by a plethora of signaling pathways. Liver fibrosis can progress into more severe stages, known as cirrhosis, when liver acini are substituted by nodules, and further to hepatocellular carcinoma. Considerable efforts are currently devoted to liver fibrosis research, not only with the goal of further elucidating the molecular mechanisms that drive this disease, but equally in view of establishing effective diagnostic and therapeutic strategies. The present paper provides a state-of-the-art overview of in vivo and in vitro models used in the field of experimental liver fibrosis research.

  8. Management strategies for liver fibrosis.

    PubMed

    Altamirano-Barrera, Alejandra; Barranco-Fragoso, Beatriz; Méndez-Sánchez, Nahum

    2017-01-01

    Liver fibrosis resulting from chronic liver injury are major causes of morbidity and mortality worldwide. Among causes of hepatic fibrosis, viral infection is most common (hepatitis B and C). In addition, obesity rates worldwide have accelerated the risk of liver injury due to nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH). Also liver fibrosis is associated with the consumption of alcohol, or autoimmune hepatitis and chronic cholangiophaties. The response of hepatocytes to inflammation plays a decisive role in the physiopathology of hepatic fibrosis, which involves the recruitment of both pro- and anti-inflammatory cells such as monocytes and macrophages. As well as the production of other cytokines and chemokines, which increase the stimulus of hepatic stellate cells by activating proinflammatory cells. The aim of this review is to identify the therapeutic options available for the treatment of the liver fibrosis, enabling the prevention of progression when is detected in time.

  9. Gardenia jasminoides attenuates hepatocellular injury and fibrosis in bile duct-ligated rats and human hepatic stellate cells

    PubMed Central

    Chen, Ying-Hua; Lan, Tian; Li, Jing; Qiu, Chun-Hui; Wu, Teng; Gou, Hong-Ju; Lu, Min-Qiang

    2012-01-01

    AIM: To investigate the anti-hepatofibrotic effects of Gardenia jasminoides in liver fibrosis. METHODS: Male Sprague-Dawley rats underwent common bile duct ligation (BDL) for 14 d and were treated with Gardenia jasminoides by gavage. The effects of Gardenia jasminoides on liver fibrosis and the detailed molecular mechanisms were also assessed in human hepatic stellate cells (LX-2) in vitro. RESULTS: Treatment with Gardenia jasminoides decreased serum alanine aminotransferase (BDL vs BDL + 100 mg/kg Gardenia jasminoides, 146.6 ± 15 U/L vs 77 ± 6.5 U/L, P = 0.0007) and aspartate aminotransferase (BDL vs BDL + 100 mg/kg Gardenia jasminoides, 188 ± 35.2 U/L vs 128 ± 19 U/L, P = 0.005) as well as hydroxyproline (BDL vs BDL + 100 mg/kg Gardenia jasminoides, 438 ± 40.2 μg/g vs 228 ± 10.3 μg/g liver tissue, P = 0.004) after BDL. Furthermore, Gardenia jasminoides significantly reduced liver mRNA and/or protein expression of transforming growth factor β1 (TGF-β1), collagen type I (Col I) and α-smooth muscle actin (α-SMA). Gardenia jasminoides significantly suppressed the upregulation of TGF-β1, Col I and α-SMA in LX-2 exposed to recombinant TGF-β1. Moreover, Gardenia jasminoides inhibited TGF-β1-induced Smad2 phosphorylation in LX-2 cells. CONCLUSION: Gardenia jasminoides exerts antifibrotic effects in the liver fibrosis and may represent a novel antifibrotic agent. PMID:23326120

  10. Effects of decoy molecules targeting NF-kappaB transcription factors in Cystic fibrosis IB3–1 cells

    PubMed Central

    Finotti, Alessia; Borgatti, Monica; Bezzerri, Valentino; Nicolis, Elena; Lampronti, Ilaria; Dechecchi, Maria; Mancini, Irene; Cabrini, Giulio; Saviano, Michele; Avitabile, Concetta; Romanelli, Alessandra; Gambari, Roberto

    2012-01-01

    One of the clinical features of cystic fibrosis (CF) is a deep inflammatory process, which is characterized by production and release of cytokines and chemokines, among which interleukin 8 (IL-8) represents one of the most important. Accordingly, there is a growing interest in developing therapies against CF to reduce the excessive inflammatory response in the airways of CF patients. Since transcription factor NF-kappaB plays a critical role in IL-8 expression, the transcription factor decoy (TFD) strategy might be of interest. In order to demonstrate that TFD against NF-kappaB interferes with the NF-kappaB pathway we proved, by chromatin immunoprecipitation (ChIP) that treatment with TFD oligodeoxyribonucleotides of cystic fibrosis IB3–1 cells infected with Pseudomonas aeruginosa leads to a decrease occupancy of the Il-8 gene promoter by NF-kappaB factors. In order to develop more stable therapeutic molecules, peptide nucleic acids (PNAs) based agents were considered. In this respect PNA-DNA-PNA (PDP) chimeras are molecules of great interest from several points of view: (1) they can be complexed with liposomes and microspheres; (2) they are resistant to DNases, serum and cytoplasmic extracts; (3) they are potent decoy molecules. By using electrophoretic mobility shift assay and RT-PCR analysis we have demonstrated that (1) the effects of PDP/PDP NF-kappaB decoy chimera on accumulation of pro-inflammatory mRNAs in P.aeruginosa infected IB3–1 cells reproduce that of decoy oligonucleotides; in particular (2) the PDP/PDP chimera is a strong inhibitor of IL-8 gene expression; (3) the effect of PDP/PDP chimeras, unlike those of ODN-based decoys, are observed even in the absence of protection with lipofectamine. These informations are of great impact, in our opinion, for the development of stable molecules to be used in non-viral gene therapy of cystic fibrosis. PMID:22772035

  11. Adipose-derived stem cells ameliorate renal interstitial fibrosis through inhibition of EMT and inflammatory response via TGF-β1 signaling pathway.

    PubMed

    Song, Yan; Peng, Changliang; Lv, Shasha; Cheng, Jing; Liu, Shanshan; Wen, Qing; Guan, Guangju; Liu, Gang

    2017-03-01

    Adipose-derived stem cells (ADSCs) have been successfully used to treat acute kidney injury or acute renal failure. However, the effect of ADSCs on treating renal interstitial fibrosis remains unknown. Here, we assessed the therapeutic efficacy of ADSCs on renal interstitial fibrosis induced by unilateral ureter obstruction (UUO) and explored the potential mechanisms. After 7days of UUO, rats were injected with ADSCs (5×10(6)) or vehicle via tail vein. We found that ADSCs administration significantly ameliorated renal interstitial fibrosis, the occurrence of epithelial-mesenchymal transition (EMT) and inflammatory response. Furthermore, ADSCs administration could inhibit the activation of transforming growth factor-β1 (TGF-β1) signaling pathway, which might play a crucial role in renal interstitial fibrosis of the UUO model rats. These results suggested that ADSCs treatment attenuates renal interstitial fibrosis possibly through inhibition of EMT and inflammatory response via TGF-β1 signaling pathway. Therefore, ADSCs may be an effective therapeutic strategy for the treatment of renal interstitial fibrosis.

  12. Cystic Fibrosis

    MedlinePlus

    Cystic fibrosis (CF) is an inherited disease of the mucus and sweat glands. It affects mostly your lungs, pancreas, liver, intestines, sinuses, and sex organs. CF causes your mucus to be thick and sticky. ...

  13. Immunophenotypic and Ultrastructural Analysis of Mast Cells in Hermansky-Pudlak Syndrome Type-1: A Possible Connection to Pulmonary Fibrosis

    PubMed Central

    Kirshenbaum, Arnold S.; Cruse, Glenn; Desai, Avanti; Bandara, Geethani; Leerkes, Maarten; Lee, Chyi-Chia R.; Fischer, Elizabeth R.; O’Brien, Kevin J.; Gochuico, Bernadette R.; Stone, Kelly; Gahl, William A.; Metcalfe, Dean D.

    2016-01-01

    Hermansky-Pudlak Syndrome type-1 (HPS-1) is an autosomal recessive disorder caused by mutations in HPS1 which result in reduced expression of the HPS-1 protein, defective lysosome-related organelle (LRO) transport and absence of platelet delta granules. Patients with HPS-1 exhibit oculocutaneous albinism, colitis, bleeding and pulmonary fibrosis postulated to result from a dysregulated immune response. The effect of the HPS1 mutation on human mast cells (HuMCs) is unknown. Since HuMC granules classify as LROs along with platelet granules and melanosomes, we set out to determine if HPS-1 cutaneous and CD34+ culture-derived HuMCs have distinct granular and cellular characteristics. Cutaneous and cultured CD34+-derived HuMCs from HPS-1 patients were compared with normal cutaneous and control HuMCs, respectively, for any morphological and functional differences. One cytokine-independent HPS-1 culture was expanded, cloned, designated the HP proMastocyte (HPM) cell line and characterized. HPS-1 and idiopathic pulmonary fibrosis (IPF) alveolar interstitium showed numerous HuMCs; HPS-1 dermal mast cells exhibited abnormal granules when compared to healthy controls. HPS-1 HuMCs showed increased CD63, CD203c and reduced mediator release following FcɛRI aggregation when compared with normal HuMCs. HPM cells also had the duplication defect, expressed FcɛRI and intracytoplasmic proteases and exhibited less mediator release following FcɛRI aggregation. HPM cells constitutively released IL-6, which was elevated in patients’ serum, in addition to IL-8, fibronectin-1 (FN-1) and galectin-3 (LGALS3). Transduction with HPS1 rescued the abnormal HPM morphology, cytokine and matrix secretion. Microarray analysis of HPS-1 HuMCs and non-transduced HPM cells confirmed upregulation of differentially expressed genes involved in fibrogenesis and degranulation. Cultured HPS-1 HuMCs appear activated as evidenced by surface activation marker expression, a decrease in mediator content and

  14. Cystic Fibrosis Transmembrane Conductance Regulator is an Epithelial Cell Receptor for Clearance of Pseudomonas aeruginosa from the Lung

    NASA Astrophysics Data System (ADS)

    Pier, Gerald B.; Grout, Martha; Zaidi, Tanweer S.

    1997-10-01

    The cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride ion channel, but its relationship to the primary clinical manifestation of CF, chronic Pseudomonas aeruginosa pulmonary infection, is unclear. We report that CFTR is a cellular receptor for binding, endocytosing, and clearing P. aeruginosa from the normal lung. Murine cells expressing recombinant human wild-type CFTR ingested 30-100 times as many P. aeruginosa as cells lacking CFTR or expressing mutant Δ F508 CFTR protein. Purified CFTR inhibited ingestion of P. aeruginosa by human airway epithelial cells. The first extracellular domain of CFTR specifically bound to P. aeruginosa and a synthetic peptide of this region inhibited P. aeruginosa internalization in vivo, leading to increased bacterial lung burdens. CFTR clears P. aeruginosa from the lung, indicating a direct connection between mutations in CFTR and the clinical consequences of CF.

  15. Client views and attitudes to non-invasive prenatal diagnosis for sickle cell disease, thalassaemia and cystic fibrosis.

    PubMed

    Hill, Melissa; Compton, Cecilia; Karunaratna, Madhavi; Lewis, Celine; Chitty, Lyn

    2014-12-01

    In the near future the availability of non-invasive prenatal diagnosis (NIPD) for single gene disorders will change the prenatal diagnosis options available to couples who are carriers of conditions such as cystic fibrosis, sickle cell disorder and thalassaemia. Client opinions about NIPD are needed to inform the implementation of NIPD for single gene disorders. This qualitative study used two focus groups (n = 12) and one-to-one interviews (n = 16) with carriers and support group representatives of sickle cell disease, thalassaemia and cystic fibrosis. Discussions were digitally recorded, transcribed verbatim and analysed using thematic analysis. Opinions about NIPD were very positive and participants valued the opportunity to have safe and early testing. Uptake of prenatal testing is likely to increase as women who had previously declined invasive testing expressed interest in having NIPD. Participant concerns about NIPD centred on the need for accuracy to be high to be used for subsequent decision making about termination of pregnancy. Participants also raised concerns that less thought may be given to having a blood test compared to an invasive test and that the perceived ease of a blood test may bring increased pressure to have testing. Participants thought NIPD should be offered through existing specialist services to ensure appropriate genetic counseling and support. Maintaining all testing options is important as some people may prefer invasive testing over NIPD if invasive testing was more accurate or if invasive testing could give information about other conditions such as Down syndrome.

  16. Dysfunction of the non-neuronal cholinergic system in the airways and blood cells of patients with cystic fibrosis.

    PubMed

    Wessler, Ignaz; Bittinger, Fernando; Kamin, Wolfgang; Zepp, Fred; Meyer, Eckhard; Schad, Arno; Kirkpatrick, Charles James

    2007-05-30

    The non-neuronal cholinergic system is widely expressed in human airways, skin and immune cells. Choline acetyltransferase (ChAT), acetylcholine and nicotine/muscarine receptors are demonstrated in epithelial surface cells, submucosal glands, airway smooth muscle fibres and immune cells. Moreover, acetylcholine is involved in the regulation of cell functions like proliferation, differentiation, migration, organization of the cytoskeleton, cell-cell contact, secretion and transport of ions and water. Cystic fibrosis (CF), the most frequent genetic disorder, is known to be caused by a mutation of the CF-gene coding for the cystic fibrosis transmembrane regulator protein (CFTR). CFTR represents a regulating transport protein for ion channels and processes involving endo- and exocytosis. Despite the identification of the genetic mutation knowledge of the underlying cellular pathways is limited. In the present experiments the cholinergic system was investigated in the peripheral blood and in the lung of CF patients undergoing lung transplantation (n=7). Acetylcholine content in bronchi and lung parenchyma of CF was reduced by 70% compared to controls (tumor-free tissue obtained from patients with lung tumor; n=13). In contrast, ChAT activity was elevated to some extent (p>0.05) in CF, and esterase activity did not differ from control. Acetylcholine content extracted from peripheral leucocytes (30 ml) was also reduced by 70% in CF (n=13) compared to healthy volunteers (n=9). Double labelling experiments with anti-CF antibodies and anti-ChAT antibodies showed a co-localization in peripheral lymphocytes, giving first evidence that CFTR may be linked with the intracellular storage/transport of non-neuronal acetylcholine. It is concluded that the non-neuronal cholinergic system is involved in the pathogenesis of CF. A reduced content of non-neuronal acetylcholine could contribute to the deleterious changes of epithelial ion and water movements in CF, because acetylcholine

  17. The low PLC-δ1 expression in cystic fibrosis bronchial epithelial cells induces upregulation of TRPV6 channel activity.

    PubMed

    Vachel, Laura; Norez, Caroline; Jayle, Christophe; Becq, Frédéric; Vandebrouck, Clarisse

    2015-01-01

    Increase of Ca(2+) influx in Cystic Fibrosis (CF) cells has been reported to be related to Transient Receptor Potential Canonical (TRPC6) channel, which is implicated in a functional coupling with Cystic Fibrosis Transmembrane conductance Regulator (CFTR). Several members of the Transient Receptor Potential Vanilloid (TRPV) channels family have already been described as emerging target for respiratory diseases. Two specific isoforms, TRPV5 and TRPV6 are of particular interest in the context of CF Ca(2+) homeostasis as they are highly selective toward Ca(2+) and constitutively activated. Thus, we investigated the involvement of these channels in Ca(2+) influx in CF and non-CF human bronchial epithelial cell lines. 16HBE14o-, CFBE41o- cell lines, primary human airway epithelial cells (hAEC) and freshly isolated human airway epithelial cells from CF and non-CF individuals were used. We showed that both channels are expressed in CF and non-CF cells and constitutive Ca(2+) influx was significantly higher (85%) in cells from CF individuals compared to cells from non-CF ones. Using the selective inhibitor of TRPV6 channel SOR-C27 and a siRNA strategy, our results revealed that TRPV6 was mostly involved in the increase of Ca(2+) influx. TRPV6 channel is negatively regulated by the PLC-PIP2 pathway. We measured the Ca(2+) influx in the presence of the non-specific PLC inhibitor, U73122, in non-CF human bronchial epithelial cells. Ca(2+) influx was increased by 33% with U73122 and this increase was largely reduced in the presence of SOR-C27. PLC inhibition in CF cells by U73122 had no effect on Ca(2+) influx. These results showed that PLC-PIP2 pathway is dysregulated in CF cells and leads to the increase of TRPV6 activity. The regulation of TRPV6 by PLC-PIP2 pathway implicates the specific PLC isoform, PLC-δ1. Immunoblot experiments revealed that expression of PLC-δ1 was decreased by 70% in CF cells. TRPV6 activity was normalized but not the level of expression of PLC-δ1

  18. The Mitochondrial Complex I Activity Is Reduced in Cells with Impaired Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Function

    PubMed Central

    Valdivieso, Angel G.; Clauzure, Mariángeles; Marín, María C.; Taminelli, Guillermo L.; Massip Copiz, María M.; Sánchez, Francisco; Schulman, Gustavo; Teiber, María L.; Santa-Coloma, Tomás A.

    2012-01-01

    Cystic fibrosis (CF) is a frequent and lethal autosomal recessive disease. It results from different possible mutations in the CFTR gene, which encodes the CFTR chloride channel. We have previously studied the differential expression of genes in CF and CF corrected cell lines, and found a reduced expression of MTND4 in CF cells. MTND4 is a mitochondrial gene encoding the MTND4 subunit of the mitochondrial Complex I (mCx-I). Since this subunit is essential for the assembly and activity of mCx-I, we have now studied whether the activity of this complex was also affected in CF cells. By using Blue Native-PAGE, the in-gel activity (IGA) of the mCx-I was found reduced in CFDE and IB3-1 cells (CF cell lines) compared with CFDE/6RepCFTR and S9 cells, respectively (CFDE and IB3-1 cells ectopically expressing wild-type CFTR). Moreover, colon carcinoma T84 and Caco-2 cells, which express wt-CFTR, either treated with CFTR inhibitors (glibenclamide, CFTR(inh)-172 or GlyH101) or transfected with a CFTR-specific shRNAi, showed a significant reduction on the IGA of mCx-I. The reduction of the mCx-I activity caused by CFTR inhibition under physiological or pathological conditions may have a profound impact on mitochondrial functions of CF and non-CF cells. PMID:23185247

  19. Infusion of Bone Marrow Mononuclear Cells Reduces Lung Fibrosis but Not Inflammation in the Late Stages of Murine Silicosis

    PubMed Central

    Lopes-Pacheco, Miquéias; Ventura, Túlio G.; de Oliveira, Helena D'Anunciação; Monção-Ribeiro, Leonardo C.; Gutfilen, Bianca; de Souza, Sergio A. L.; Rocco, Patrícia R. M.; Borojevic, Radovan; Morales, Marcelo M.; Takiya, Christina M.

    2014-01-01

    We hypothesized that infusion of bone marrow mononuclear cells (BMMCs) in the late stages of silica-induced damage would reduce the remodelling process in a murine model of silicosis. C57BL/6 mice were assigned to 2 groups. In the SIL group, mice were instilled with a silica particle suspension intratracheally. Control (C) mice received saline under the same protocol. On the 40th day, some of the animals from both groups were killed. The others were treated with either saline or BMMCs (1×106cells) intravenously (C+BMMC and SIL+BMMC), and the mice were killed 70 days after the start of the protocol. In the mice in the SIL+BMMC group, collagen deposition, the presence of silica particles inside nodules, the presence of macrophages and cells reactive for inducible nitric oxide synthase were reduced. Lung parameters also improved. Beyond that, the total and differential cellularity of bronchoalveolar lavage fluid, immunoexpression of transforming growth factor-β, the number of T regulatory cells and apoptosis were increased. However, the presence of male donor cells in lung tissue was not observed using GFP+ cells (40d) or Y chromosome DNA (70d). Therefore, BMMC therapy in the late stages of experimental silicosis improved lung function by diminishing fibrosis but inflammatory cells persisted, which could be related to expansion of T regulatory cells, responsible for the beneficial effects of cell therapy. PMID:25299237

  20. Relationships among CFTR expression, HCO3- secretion, and host defense may inform gene- and cell-based cystic fibrosis therapies.

    PubMed

    Shah, Viral S; Ernst, Sarah; Tang, Xiao Xiao; Karp, Philip H; Parker, Connor P; Ostedgaard, Lynda S; Welsh, Michael J

    2016-05-10

    Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel. Airway disease is the major source of morbidity and mortality. Successful implementation of gene- and cell-based therapies for CF airway disease requires knowledge of relationships among percentages of targeted cells, levels of CFTR expression, correction of electrolyte transport, and rescue of host defense defects. Previous studies suggested that, when ∼10-50% of airway epithelial cells expressed CFTR, they generated nearly wild-type levels of Cl(-) secretion; overexpressing CFTR offered no advantage compared with endogenous expression levels. However, recent discoveries focused attention on CFTR-mediated HCO3 (-) secretion and airway surface liquid (ASL) pH as critical for host defense and CF pathogenesis. Therefore, we generated porcine airway epithelia with varying ratios of CF and wild-type cells. Epithelia with a 50:50 mix secreted HCO3 (-) at half the rate of wild-type epithelia. Likewise, heterozygous epithelia (CFTR(+/-) or CFTR(+/∆F508)) expressed CFTR and secreted HCO3 (-) at ∼50% of wild-type values. ASL pH, antimicrobial activity, and viscosity showed similar relationships to the amount of CFTR. Overexpressing CFTR increased HCO3 (-) secretion to rates greater than wild type, but ASL pH did not exceed wild-type values. Thus, in contrast to Cl(-) secretion, the amount of CFTR is rate-limiting for HCO3 (-) secretion and for correcting host defense abnormalities. In addition, overexpressing CFTR might produce a greater benefit than expressing CFTR at wild-type levels when targeting small fractions of cells. These findings may also explain the risk of airway disease in CF carriers.

  1. The effect of ambroxol on chloride transport, CFTR and ENaC in cystic fibrosis airway epithelial cells.

    PubMed

    Varelogianni, Georgia; Hussain, Rashida; Strid, Hilja; Oliynyk, Igor; Roomans, Godfried M; Johannesson, Marie

    2013-11-01

    Ambroxol, a mucokinetic anti-inflammatory drug, has been used for treatment of cystic fibrosis (CF). The respiratory epithelium is covered by the airway surface liquid (ASL), the thickness and composition of which is determined by Cl(-) efflux via the cystic fibrosis transmembrane conductance regulator (CFTR) and Na(+) influx via the epithelial Na(+) channel (ENaC). In cells expressing wt-CFTR, ambroxol increased the Cl(-) conductance, but not the bicarbonate conductance of the CFTR channels. We investigated whether treatment with ambroxol enhances chloride transport and/or CFTR and ENaC expression in CF airway epithelial cells (CFBE) cells. CFBE cells were treated with 100 µM ambroxol for 2, 4 or 8 h. mRNA expression for CFTR and ENaC subunits was analysed by real-time polymerase chain reaction (RT-PCR); protein expression was measured by Western blot. The effect of ambroxol on Cl(-) transport was measured by Cl(-) efflux measurements with a fluorescent chloride probe. Ambroxol significantly stimulated Cl(-) efflux from CFBE cells (a sixfold increase after 8 h treatment), and enhanced the expression of the mRNA of CFTR and α-ENaC, and of the CFTR protein. No significant difference was observed in β-ENaC after exposure to ambroxol, whereas mRNA expression of γ-ENaC was reduced. No significant effects of ambroxol on the ENaC subunits were observed by Western blot. Ambroxol did not significantly affect the intracellular Ca(2+) concentration. Upregulation of CFTR and enhanced Cl(-) efflux after ambroxol treatment should promote transepithelial ion and water transport, which may improve hydration of the mucus, and therefore be beneficial to CF-patients.

  2. Adenovirus-mediated expression of orphan nuclear receptor NR4A2 targeting hepatic stellate cell attenuates liver fibrosis in rats

    PubMed Central

    Chen, Pengguo; Li, Jie; Huo, Yan; Lu, Jin; Wan, Lili; Yang, Quanjun; Huang, Jinlu; Gan, Run; Guo, Cheng

    2016-01-01

    Liver fibrosis is a wound-healing response characterized with the accumulation of extracellular matrix (ECM). And hepatic stellate cells (HSCs) are the principal cell source of ECM. NR4A2 (Nurr1) is a member of orphan nuclear receptor NR4A family and acts as transcription factor. It participates in regulating cell differentiation, proliferation and apoptosis. We previously demonstrated that NR4A2 expression in fibrotic liver reduced significantly compared with normal liver and NR4A2 knockout in HSCs promoted ECM production. In the present study we explored the role of NR4A2 on liver fibrosis. Studies in cultured HSCs demonstrated that NR4A2 over-expression suppressed the activation of HSCs, such as ECM production and invasion ability. Moreover cell cycle was arrested, cell apoptosis was promoted and cell signaling pathway was influenced. Adenovirus-mediated delivery of NR4A2 in rats ameliorated significantly dimethylnitrosamine (DMN) induced liver fibrosis. The In vivo experiments produced results consistent with in vitro experiments. Taken together these results demonstrate NR4A2 enhancement attenuates liver fibrosis via suppressing the activation of HSCs and NR4A2 may be an ideal target for anti-fibrotic therapy. PMID:27646469

  3. Murine junctional adhesion molecules JAM-B and JAM-C mediate endothelial and stellate cell interactions during hepatic fibrosis.

    PubMed

    Hintermann, Edith; Bayer, Monika; Ehser, Janine; Aurrand-Lions, Michel; Pfeilschifter, Josef M; Imhof, Beat A; Christen, Urs

    2016-07-03

    Classical junctional adhesion molecules JAM-A, JAM-B and JAM-C influence vascular permeability, cell polarity as well as leukocyte recruitment and immigration into inflamed tissue. As the vasculature becomes remodelled in chronically injured, fibrotic livers we aimed to determine distribution and role of junctional adhesion molecules during this pathological process. Therefore, livers of naïve or carbon tetrachloride-treated mice were analyzed by immunohistochemistry to localize all 3 classical junctional adhesion molecules. Hepatic stellate cells and endothelial cells were isolated and subjected to immunocytochemistry and flow cytometry to determine localization and functionality of JAM-B and JAM-C. Cells were further used to perform contractility and migration assays and to study endothelial tubulogenesis and pericytic coverage by hepatic stellate cells. We found that in healthy tissue, JAM-A was ubiquitously expressed whereas JAM-B and JAM-C were restricted to the vasculature. During fibrosis, JAM-B and JAM-C levels increased in endothelial cells and JAM-C was de novo generated in myofibroblastic hepatic stellate cells. Soluble JAM-C blocked contractility but increased motility in hepatic stellate cells. Furthermore, soluble JAM-C reduced endothelial tubulogenesis and endothelial cell/stellate cell interaction. Thus, during liver fibrogenesis, JAM-B and JAM-C expression increase on the vascular endothelium. More importantly, JAM-C appears on myofibroblastic hepatic stellate cells linking them as pericytes to JAM-B positive endothelial cells. This JAM-B/JAM-C mediated interaction between endothelial cells and stellate cells stabilizes vessel walls and may control the sinusoidal diameter. Increased hepatic stellate cell contraction mediated by JAM-C/JAM-C interaction may cause intrahepatic vasoconstriction, which is a major complication in liver cirrhosis.

  4. Mature cystic fibrosis airway neutrophils suppress T cell function: evidence for a role of arginase 1 but not programmed death-ligand 1.

    PubMed

    Ingersoll, Sarah A; Laval, Julie; Forrest, Osric A; Preininger, Marcela; Brown, Milton R; Arafat, Dalia; Gibson, Greg; Tangpricha, Vin; Tirouvanziam, Rabindra

    2015-06-01

    Bacteria colonize cystic fibrosis (CF) airways, and although T cells with appropriate Ag specificity are present in draining lymph nodes, they are conspicuously absent from the lumen. To account for this absence, we hypothesized that polymorphonuclear neutrophils (PMNs), recruited massively into the CF airway lumen and actively exocytosing primary granules, also suppress T cell function therein. Programmed death-ligand 1 (PD-L1), which exerts T cell suppression at a late step, was expressed bimodally on CF airway PMNs, delineating PD-L1(hi) and PD-L1(lo) subsets, whereas healthy control (HC) airway PMNs were uniformly PD-L1(hi). Blood PMNs incubated in CF airway fluid lost PD-L1 over time; in coculture, Ab blockade of PD-L1 failed to inhibit the suppression of T cell proliferation by CF airway PMNs. In contrast with PD-L1, arginase 1 (Arg1), which exerts T cell suppression at an early step, was uniformly high on CF and HC airway PMNs. However, arginase activity was high in CF airway fluid and minimal in HC airway fluid, consistent with the fact that Arg1 activation requires primary granule exocytosis, which occurs in CF, but not HC, airway PMNs. In addition, Arg1 expression on CF airway PMNs correlated negatively with lung function and positively with arginase activity in CF airway fluid. Finally, combined treatment with arginase inhibitor and arginine rescued the suppression of T cell proliferation by CF airway fluid. Thus, Arg1 and PD-L1 are dynamically modulated upon PMN migration into human airways, and, Arg1, but not PD-L1, contributes to early PMN-driven T cell suppression in CF, likely hampering resolution of infection and inflammation.

  5. MATURE CYSTIC FIBROSIS AIRWAY NEUTROPHILS SUPPRESS T-CELL FUNCTION: EVIDENCE FOR A ROLE OF ARGINASE 1, BUT NOT PROGRAMMED DEATH-LIGAND 1

    PubMed Central

    Ingersoll, Sarah A.; Laval, Julie; Forrest, Osric A.; Preininger, Marcela; Brown, Milton R.; Arafat, Dalia; Gibson, Greg; Tangpricha, Vin; Tirouvanziam, Rabindra

    2015-01-01

    Bacteria colonize cystic fibrosis (CF) airways, and while T cells with appropriate antigen specificity are present in draining lymph nodes, they are conspicuously absent from the lumen. To account for this absence, we hypothesized that polymorphonuclear neutrophils (PMNs), recruited massively into the CF airway lumen and actively exocytosing primary granules, also suppress T-cell function therein. Programmed Death-Ligand 1 (PD-L1), which exerts T-cell suppression at a late step, was expressed bimodally on CF airway PMNs, delineating PD-L1hi and PD-L1lo subsets, while healthy control (HC) airway PMNs were uniformly PD-L1hi. Blood PMNs incubated in CF airway fluid lost PD-L1 over time, and in coculture, antibody blockade of PD-L1 failed to inhibit the suppression of T-cell proliferation by CF airway PMNs. In contrast with PD-L1, arginase 1 (Arg1), which exerts T-cell suppression at an early step, was uniformly high on CF and HC airway PMNs. However, arginase activity was high in CF airway fluid and minimal in HC airway fluid, consistent with the fact that Arg1 activation requires primary granule exocytosis, which occurs in CF, but not HC, airway PMNs. In addition, Arg1 expression on CF airway PMNs correlated negatively with lung function and positively with arginase activity in CF airway fluid. Finally, combined treatment with arginase inhibitor and arginine rescued the suppression of T-cell proliferation by CF airway fluid. Thus, Arg1 and PD-L1 are dynamically modulated upon PMN migration into human airways, and, Arg1, but not PD-L1, contributes to early PMN-driven T-cell suppression in CF, likely hampering resolution of infection and inflammation. PMID:25926674

  6. Idiopathic Pulmonary Fibrosis

    MedlinePlus

    ... the NHLBI on Twitter. What Is Idiopathic Pulmonary Fibrosis? Pulmonary fibrosis (PULL-mun-ary fi-BRO-sis) is a ... time. The formation of scar tissue is called fibrosis. As the lung tissue thickens, your lungs can' ...

  7. Cystic fibrosis - resources

    MedlinePlus

    Resources - cystic fibrosis ... The following organizations are good resources for information on cystic fibrosis : Cystic Fibrosis Foundation -- www.cff.org March of Dimes -- www.marchofdimes.org/baby/cystic-fibrosis-and- ...

  8. GTP-Binding Proteins Inhibit cAMP Activation of Chloride Channels in Cystic Fibrosis Airway Epithelial Cells

    NASA Astrophysics Data System (ADS)

    Schwiebert, Erik M.; Kizer, Neil; Gruenert, Dieter C.; Stanton, Bruce A.

    1992-11-01

    Cystic fibrosis (CF) is a genetic disease characterized, in part, by defective regulation of Cl^- secretion by airway epithelial cells. In CF, cAMP does not activate Cl^- channels in the apical membrane of airway epithelial cells. We report here whole-cell patch-clamp studies demonstrating that pertussis toxin, which uncouples heterotrimeric GTP-binding proteins (G proteins) from their receptors, and guanosine 5'-[β-thio]diphosphate, which prevents G proteins from interacting with their effectors, increase Cl^- currents and restore cAMP-activated Cl^- currents in airway epithelial cells isolated from CF patients. In contrast, the G protein activators guanosine 5'-[γ-thio]triphosphate and AlF^-_4 reduce Cl^- currents and inhibit cAMP from activating Cl^- currents in normal airway epithelial cells. In CF cells treated with pertussis toxin or guanosine 5'-[β-thio]diphosphate and in normal cells, cAMP activates a Cl^- conductance that has properties similar to CF transmembrane-conductance regulator Cl^- channels. We conclude that heterotrimeric G proteins inhibit cAMP-activated Cl^- currents in airway epithelial cells and that modulation of the inhibitory G protein signaling pathway may have the therapeutic potential for improving cAMP-activated Cl^- secretion in CF.

  9. Restoring homeostasis of CD4⁺ T cells in hepatitis-B-virus-related liver fibrosis.

    PubMed

    Cheng, Li-Sha; Liu, Yun; Jiang, Wei

    2015-10-14

    Immune-mediated liver injury is widely seen during hepatitis B virus (HBV) infection. Unsuccessful immune clearance of HBV results in chronic hepatitis and increases the risk of liver cirrhosis and hepatocellular carcinoma. HBV-related liver fibrosis (HBVLF), occurring as a result of HBV-induced chronic hepatitis, is a reversible, intermediate stage of chronic hepatitis B (CHB) and liver cirrhosis. Therefore, defining the pathogenesis of HBVLF is of practical significance for achieving better clinical outcomes. Recently, the homeostasis of CD4(+) T cells was considered to be pivotal in the process of HBVLF. To better uncover the underlying mechanisms, in this review, we systematically retrospect the impacts of different CD4(+) T-cell subsets on CHB and HBVLF. We emphasize CD4(+) T-cell homeostasis and the important balance between regulatory T (Treg) and T helper 17 (Th17) cells. We discuss some cytokines associated with Treg and Th17 cells such as interleukin (IL)-17, IL-22, IL-21, IL-23, IL-10, IL-35 and IL-33, as well as surface molecules such as programmed cell death protein 1, cytotoxic T lymphocyte-associated antigen 4, T cell immunoglobulin domain and mucin domain-containing molecule 3 and cannabinoid receptor 2 that have potential therapeutic implications for the homeostasis of CD4(+) T cells in CHB and HBVLF.

  10. Glycaemic regulation and insulin secretion are abnormal in cystic fibrosis pigs despite sparing of islet cell mass.

    PubMed

    Uc, Aliye; Olivier, Alicia K; Griffin, Michelle A; Meyerholz, David K; Yao, Jianrong; Abu-El-Haija, Maisam; Buchanan, Katherine M; Vanegas Calderón, Oriana G; Abu-El-Haija, Marwa; Pezzulo, Alejandro A; Reznikov, Leah R; Hoegger, Mark J; Rector, Michael V; Ostedgaard, Lynda S; Taft, Peter J; Gansemer, Nick D; Ludwig, Paula S; Hornick, Emma E; Stoltz, David A; Ode, Katie L; Welsh, Michael J; Engelhardt, John F; Norris, Andrew W

    2015-01-01

    Diabetes is a common and significant co-morbidity in cystic fibrosis (CF). The pathogenesis of cystic fibrosis related diabetes (CFRD) is incompletely understood. Because exocrine pancreatic disease is similar between humans and pigs with CF, the CF pig model has the potential to contribute significantly to the understanding of CFRD pathogenesis. We determined the structure of the endocrine pancreas in fetal, newborn and older CF and non-CF pigs and assessed endocrine pancreas function by intravenous glucose tolerance test (IV-GTT). In fetal pigs, pancreatic insulin and glucagon density was similar between CF and non-CF. In newborn and older pigs, the insulin and glucagon density was unchanged between CF and non-CF per total pancreatic area, but increased per remnant lobular tissue in CF reflecting exocrine pancreatic loss. Although fasting glucose levels were not different between CF and non-CF newborns, CF newborns demonstrated impaired glucose tolerance and increased glucose area under the curve during IV-GTT. Second phase insulin secretion responsiveness was impaired in CF newborn pigs and significantly lower than that observed in non-CF newborns. Older CF pigs had elevated random blood glucose levels compared with non-CF. In summary, glycaemic abnormalities and insulin secretion defects were present in newborn CF pigs and spontaneous hyperglycaemia developed over time. Functional changes in CF pig pancreas were not associated with a decline in islet cell mass. Our results suggest that functional islet abnormalities, independent of structural islet loss, contribute to the early pathogenesis of CFRD.

  11. Transplantation of mesenchymal stem cells expressing TIMP-1-shRNA improves hepatic fibrosis in CCl4-treated rats

    PubMed Central

    Zhu, Yingwei; Miao, Zongning; Gong, Lei; Chen, Weichang

    2015-01-01

    This study was to investigate the therapeutic effect of intravenous transplantation of TIMP-1-silencing mesenchymal stem cells (MSCs) in a rat model of liver fibrosis. MSCs were transduced with a lentiviral vector expressing tissue inhibitor of metalloproteinase 1 (TIMP-1)-shRNA, and the liver cirrhosis model was established by injection of CCl4 (1 ml/kg body weight twice a week for 4 weeks) in Sprague Dawley rats. The survived 36 rats were randomly divided into 3 groups: control group, MSCs group, and TIMP-1-shRNA group. At 4 weeks after establishment of animal model, 3×106 MSCs were intravenously injected. In TIMP-1-shRNA group, MSCs expressing TIMP-1-shRNA were transplanted. Animals were sacrificed 4 weeks later. Blood was collected for the detection of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). The livers were harvested for histological examination. At 5 days after transfection, strong fluorescence was detectable in each group. TIMP-1-shRNA group had the lowest TIMP-1 expression. Following MSCs transplantation, serum ALT and AST reduced in rats with hepatic cirrhosis, and histology showed less fibrotic areas and collagens, as compared to control group. These improvements were more obvious in the TIMP-1-shRNA group. Our study indicates that transplantation of MSCs expressing TIMP-1-shRNA is able to inhibit the progression of liver fibrosis and possibly restore the liver function in a rat model. PMID:26464632

  12. A muscle resident cell population promotes fibrosis in hindlimb skeletal muscles of mdx mice through the Wnt canonical pathway.

    PubMed

    Trensz, Frédéric; Haroun, Sonia; Cloutier, Alexandre; Richter, Martin V; Grenier, Guillaume

    2010-11-01

    Previous work has pointed to a role for the Wnt canonical pathway in fibrosis formation in aged skeletal muscles. In the present study, we studied the dystrophic mdx mouse, which displays skeletal muscle fibrosis. Our results indicated that the muscle resident stromal cell (mrSC) population in the muscles of dystrophic mice is higher than in the muscles of age-matched wild-type mice. Wnt3a promoted the proliferation of and collagen expression by cultured mrSCs but arrested the growth of and collagen expression by cultured myoblasts. Injections of Wnt3A in the tibialis anterior muscles of adult wild-type mice significantly enhanced the mrSC population and collagen deposition compared with the contralateral muscles. Conversely, an injection of the Wnt antagonist Dickkof protein (DKK1) into the skeletal muscles of mdx mice significantly reduced collagen deposition. These results suggested that the Wnt canonical pathway expands the population of mrSCs and stimulates their production of collagen as observed during aging and in various myopathies.

  13. Adipocytokines and Hepatic Fibrosis

    PubMed Central

    Saxena, Neeraj K.; Anania, Frank A.

    2015-01-01

    Obesity and metabolic syndrome pose significant risk for progression of many types of chronic illnesses, including liver disease. Hormones released from adipocytes, adipocytokines, associated with obesity and metabolic syndrome, have been shown to control hepatic inflammation and fibrosis. Hepatic fibrosis is the final common pathway that can result in cirrhosis, and can ultimately require liver transplantation. Initially, two key adipocytokines, leptin and adiponectin, appeared to control many fundamental aspects of the cell and molecular biology related to hepatic fibrosis and its resolution. Leptin appears to act as a profibrogenic molecule while adiponectin possesses strong-anti-fibrotic properties. In this review, we emphasize pertinent data associated with these, and recently discovered, adipocytokines that may drive or halt the fibrogenic response in the liver. PMID:25656826

  14. CD147 promotes liver fibrosis progression via VEGF-A/VEGFR2 signalling-mediated cross-talk between hepatocytes and sinusoidal endothelial cells.

    PubMed

    Yan, Zhaoyong; Qu, Kai; Zhang, Jing; Huang, Qichao; Qu, Ping; Xu, Xinsen; Yuan, Peng; Huang, Xiaojun; Shao, Yongping; Liu, Chang; Zhang, Hongxin; Xing, Jinliang

    2015-10-01

    Although previous evidence indicates close involvement of CD147 in the pathogenesis of liver fibrosis, the underlying molecular mechanisms and its therapeutic value remain largely unknown. In the present study, we investigated the biological roles of CD147 in liver fibrosis and assessed its therapeutic value as a target molecule in the CCl4-induced liver fibrosis mouse model. We found that CD147 was highly expressed in both hepatocytes and SECs (sinusoidal endothelial cells) in fibrotic liver tissues. Additionally, it was significantly associated with the fibrosis stage. TGF-β1 (transforming growth factor β1) was found to be mainly responsible for the up-regulation of CD147. Bioinformatic and experimental data suggest a functional link between CD147 expression and VEGF-A (vascular endothelial growth factor A)/VEGR-2 (VEGF receptor 2) signalling-mediated angiogenesis in fibrotic liver tissues. Furthermore, we observed that the CD147-induced activation of the PI3K (phosphoinositide 3-kinase)/Akt signalling pathway promotes the production of VEGF-A in hepatocytes and expression of VEGFR-2 in SECs, which was found to enhance the angiogenic capability of SECs. Finally, our data indicate that blocking of CD147 using an mAb (monoclonal antibody) attenuated liver fibrosis progression via inhibition of VEGF-A/VEGFR-2 signalling and subsequent amelioration of microvascular abnormality in the CCl4-induced mouse model. Our findings suggest a novel functional mechanism that CD147 may promote liver fibrosis progression via inducing the VEGF-A/VEGFR-2 signalling pathway-mediated cross-talk between hepatocytes and SECs. New strategies based on the intervention of CD147 can be expected for prevention of liver fibrosis.

  15. Blocking TGF-β1 by P17 peptides attenuates gastric cancer cell induced peritoneal fibrosis and prevents peritoneal dissemination in vitro and in vivo.

    PubMed

    Lv, Zhi-Dong; Zhao, Wei-Jun; Jin, Li-Ying; Wang, Wen-Juan; Dong, Qian; Li, Na; Xu, Hui-Mian; Wang, Hai-Bo

    2017-04-01

    Our previous study demonstrated that the peritoneal stroma environment favors proliferation of tumor cells by serving as a rich source of growth factors and chemokines known to be involved in tumor metastasis. In this study, we investigated the interaction between gastric cancer cells and peritoneal mesothelial cells, and determined the effects of TGF-β1 in this processing. Human peritoneal tissues and peritoneal wash fluid were obtained, which examined by hematoxylin and eosin staining or ELISA for measurements of TGF-β1 levels. The peritoneal mesothelial cells were co-incubated with the supernatants of gastric cancer, the expression of TGF-β1, collagen and fibronectin was observed by ELISA and western blot. We then investigated the effects of serum-free conditioned media from HSC-39 gastric cancer cells on the peritoneum of nude mice, and the effects of peritoneal fibrosis on the development of peritoneal metastasis in vivo. The peritoneum from gastric patients were thickened and contained extensive fibrosis. After co-culture both gastric tumor cells and mesothelial cells, we found that TGF-β1 expression was greatly increased in the co-culture system compared to individual culture condition. Serum-free Conditioned Media from HSC-39 was able to induce extracellular matrix expression in vitro and in vivo, and tumorigenicity in mice with peritoneal fibrosis was greater than in mice with normal peritoneum, while blocking TGF-β1 by peptide P17 can partially inhibit these effects. In conclusion, these results indicated that the interaction of gastric cancer with peritoneal fibrosis and determined that TGF-β1 plays a key role in induction of peritoneal fibrosis, which in turn affected dissemination of gastric cancer.

  16. MicroRNA-29b inhibits TGF-β1-induced fibrosis via regulation of the TGF-β1/Smad pathway in primary human endometrial stromal cells

    PubMed Central

    LI, JINGXIONG; CEN, BOHONG; CHEN, SIPING; HE, YUANLI

    2016-01-01

    Transforming growth factor (TGF)-β1 has a key role in the regulation of fibrosis and organ dysfunction. During the pathogenesis and progression of vital organ fibrosis, the microRNA (miR)-29 family is irregularly downregulated and exogenous supplementation of miR-29b has a strong anti-fibrotic capacity. However, whether TGF-β1 is able to provoke endometrial fibrosis, and the role of miR-29 in endometrial fibrosis remain unclear. In the present study, RT-qPCR, immunocytochemistry, western blot analysis, scanning electron microscopy, immunofluorescence staining, cell proliferation assay and flow cytometric analysis were employed. The results demonstrated that the expression levels of collagen, type 1, alpha 1 (COL1A1), α-smooth muscle actin (α-SMA) and phosphorylated (p)-Smad2/3 were increased, whereas miR-29b and maternally expressed gene 3 (MEG3) were decreased in primary endometrial stromal cells (ESCs) in response to TGF-β1 stimulation, in a time and dose-dependent manner. Furthermore, overexpression of miR-29b markedly reduced the expression levels of COL1A1 and α-SMA, and decreased the expression and nuclear accumulation of p-Smad2/3. In addition, ectopic overexpression of miR-29b increased the expression levels of MEG3, inhibited myofibroblast-like cell proliferation and induced apoptosis. These findings indicated that miR-29b may have a significant anti-fibrotic role, and may attenuate TGF-β1-induced fibrosis in ESCs. Therefore, exogenous miR-29b may serve as a potential therapeutic agent for the treatment of endometrial fibrosis. PMID:27035110

  17. Cardiac Fibrosis: The Fibroblast Awakens

    PubMed Central

    Travers, Joshua G.; Kamal, Fadia A.; Robbins, Jeffrey; Yutzey, Katherine E.; Blaxall, Burns C.

    2016-01-01

    Myocardial fibrosis is a significant global health problem associated with nearly all forms of heart disease. Cardiac fibroblasts comprise an essential cell type in the heart that is responsible for the homeostasis of the extracellular matrix; however upon injury, these cells transform to a myofibroblast phenotype and contribute to cardiac fibrosis. This remodeling involves pathological changes that include chamber dilation, cardiomyocyte hypertrophy and apoptosis, and ultimately leads to the progression to heart failure. Despite the critical importance of fibrosis in cardiovascular disease, our limited understanding of this cell population impedes the development of potential therapies that effectively target this cell type and its pathological contribution to disease progression. This review summarizes current knowledge regarding the origins and roles of fibroblasts, mediators and signaling pathways known to influence fibroblast function after myocardial injury, as well as novel therapeutic strategies under investigation to attenuate cardiac fibrosis. PMID:26987915

  18. Complement Component 5 Mediates Development of Fibrosis, via Activation of Stellate Cells, in 2 Mouse Models of Chronic Pancreatitis

    PubMed Central

    Sendler, Matthias; Beyer, Georg; Mahajan, Ujjwal M.; Kauschke, Vivien; Maertin, Sandrina; Schurmann, Claudia; Homuth, Georg; Völker, Uwe; Völzke, Henry; Halangk, Walter; Wartmann, Thomas; Weiss, Frank-Ulrich; Hegyi, Peter; Lerch, Markus M.; Mayerle, Julia

    2015-01-01

    Background & Aims Little is known about the pathogenic mechanisms of chronic pancreatitis. We investigated the roles of complement component 5 (C5) in pancreatic fibrogenesis in mice and patients. Methods Chronic pancreatitis was induced by ligation of the midpancreatic duct, followed by a single supramaximal intraperitoneal injection of cerulein, in C57Bl6 (control) and C5-deficient mice. Some mice were given injections of 2 different antagonists of the receptor for C5a over 21 days. In a separate model, mice were given injections of cerulein for 10 weeks to induce chronic pancreatitis. Direct effects of C5 were studied in cultured primary cells. We performed genotype analysis for the single-nucleotide polymorphisms rs 17611 and rs 2300929 in C5 in patients with pancreatitis and healthy individuals (controls). Blood cells from 976 subjects were analyzed by transcriptional profiling. Results During the initial phase of pancreatitis, levels of pancreatic damage were similar between C5-deficient and control mice. During later stages of pancreatitis, C5-deficient mice and mice given injections of C5a-receptor antagonists developed significantly less pancreatic fibrosis than control mice. Primary pancreatic stellate cells were activated in vitro by C5a. There were no differences in the rs 2300929 SNP between subjects with or without pancreatitis, but the minor allele rs17611 was associated with a significant increase in levels of C5 in whole blood. Conclusions In mice, loss of C5 or injection of a C5a-receptor antagonist significantly reduced the level of fibrosis of chronic pancreatitis, but this was not a consequence of milder disease in early stages of pancreatitis. C5 might be a therapeutic target for chronic pancreatitis. PMID:26001927

  19. Wilms' tumor 1 (Wt1) regulates pleural mesothelial cell plasticity and transition into myofibroblasts in idiopathic pulmonary fibrosis

    PubMed Central

    Karki, Suman; Surolia, Ranu; Hock, Thomas David; Guroji, Purusotham; Zolak, Jason S.; Duggal, Ryan; Ye, Tong; Thannickal, Victor J.; Antony, Veena B.

    2014-01-01

    Pleural mesothelial cells (PMCs), which are derived from the mesoderm, exhibit an extraordinary capacity to undergo phenotypic changes during development and disease. PMC transformation and trafficking has a newly defined role in idiopathic pulmonary fibrosis (IPF); however, the contribution of Wilms' tumor 1 (Wt1)-positive PMCs to the generation of pathognomonic myofibroblasts remains unclear. PMCs were obtained from IPF lung explants and healthy donor lungs that were not used for transplantation. Short hairpin Wt1-knockdown PMCs (sh Wt1) were generated with Wt1 shRNA, and morphologic and functional assays were performed in vitro. Loss of Wt1 abrogated the PMC phenotype and showed evidence of mesothelial-to-mesenchymal transition (MMT), with a reduced expression of E-cadherin and an increase in the profibrotic markers α-smooth muscle actin (α-SMA) and fibronectin, along with increased migration and contractility, compared with that of the control. Migration of PMCs in response to active transforming growth factor (TGF)-β1 was assessed by live-cell imaging with 2-photon microscopy and 3D imaging, of Wt1-EGFP transgenic mice. Lineage-tracing experiments to map the fate of Wt1+ PMCs in mouse lung in response to TGF-β1 were also performed by using a Cre-loxP system. Our results, for the first time, demonstrate that Wt1 is necessary for the morphologic integrity of pleural membrane and that loss of Wt1 contributes to IPF via MMT of PMCs into a myofibroblast phenotype.—Karki, S., Surolia, R., Hock, T. D., Guroji, P., Zolak, J. S., Duggal, R., Ye, T., Thannickal, V., J., Antony, V. B. Wilms' tumor 1 (Wt1) regulates pleural mesothelial cell plasticity and transition into myofibroblasts in idiopathic pulmonary fibrosis. PMID:24265486

  20. Metformin against TGFβ-induced epithelial-to-mesenchymal transition (EMT): from cancer stem cells to aging-associated fibrosis.

    PubMed

    Cufí, Silvia; Vazquez-Martin, Alejandro; Oliveras-Ferraros, Cristina; Martin-Castillo, Begoña; Joven, Jorge; Menendez, Javier A

    2010-11-15

    Transforming Growth Factor-b (TGFb) is a major driving force of the Epithelial-to-Mesenchymal (EMT) genetic program, which becomes overactive in the pathophysiology of many age-related human diseases.  TGFb-driven EMT is sufficient to generate migrating cancer stem cells by directly linking the acquisition of cellular motility with the maintenance of tumor-initiating (stemness) capacity.  Chronic diseases exhibiting excessive fibrosis can be caused by repeated and sustained infliction of TGFb-driven EMT, which increases collagen and extracellular matrix synthesis.  Pharmacological prevention and/or reversal of TGFb-induced EMT may therefore have important clinical applications in the management of cancer metastasis as well as in the prevention and/or treatment of end-state organ failures.  Earlier studies from our group have revealed that clinically-relevant concentrations of the biguanide derivative metformin, the most widely used oral agent to lower blood glucose concentration in patients with type 2 diabetes and metabolic syndrome, notably decreased both the self-renewal and the proliferation of trastuzumab-refractory breast cancer stem cell populations.  Given that: a.) tumor-initiating cancer stem cells display a significant enrichment in the expression of basal/mesenchymal or myoepithelial markers, including an increased secretion of TGFb; b.) metformin treatment impedes the ontogeny of generating the stem cell phenotype by transcriptionally repressing key drivers of the EMT genetic program (e.g. ZEB1, TWIST1, SNAIL2 [Slug], TGFbs), we recently hypothesized that prevention of TGFb-induced EMT might represent a common molecular mechanism underlying the anti-cancer stem cells and anti-fibrotic actions of metformin.  Remarkably, metformin exposure not only impedes TGFb-promoted loss of the epithelial marker E-cadherin in MCF-7 breast cancer cells but it prevents further TGF-induced cell scattering and accumulation of the mesenchymal marker vimentin in

  1. Induced Pluripotent Stem Cells Inhibit Bleomycin-Induced Pulmonary Fibrosis in Mice through Suppressing TGF-β1/Smad-Mediated Epithelial to Mesenchymal Transition

    PubMed Central

    Zhou, Yan; He, Zhong; Gao, Yuan; Zheng, Rui; Zhang, Xiaoye; Zhao, Li; Tan, Mingqi

    2016-01-01

    Pulmonary fibrosis is a progressive and irreversible fibrotic lung disorder with high mortality and few treatment options. Recently, induced pluripotent stem (iPS) cells have been considered as an ideal resource for stem cell-based therapy. Although, an earlier study demonstrated the therapeutic effect of iPS cells on pulmonary fibrosis, the exact mechanisms remain obscure. The present study investigated the effects of iPS cells on inflammatory responses, transforming growth factor (TGF)-β1 signaling pathway, and epithelial to mesenchymal transition (EMT) during bleomycin (BLM)-induced lung fibrosis. A single intratracheal instillation of BLM (5 mg/kg) was performed to induce pulmonary fibrosis in C57BL/6 mice. Then, iPS cells (c-Myc-free) were administrated intravenously at 24 h following BLM instillation. Three weeks after BLM administration, pulmonary fibrosis was evaluated. As expected, treatment with iPS cells significantly limited the pathological changes, edema, and collagen deposition in lung tissues of BLM-induced mice. Mechanically, treatment with iPS cells obviously repressed the expression ratios of matrix metalloproteinase-2 (MMP-2) to its tissue inhibitor -2 (TIMP-2) and MMP-9/TIMP-1 in BLM-induced pulmonary tissues. In addition, iPS cell administration remarkably suppressed BLM-induced up-regulation of pulmonary inflammatory mediators, including tumor necrosis factor-α, interleukin (IL)-1β, IL-6, inducible nitric oxide synthase, nitric oxide, cyclooxygenase-2 and prostaglandin E2. We further demonstrated that transplantation of iPS cells markedly inhibited BLM-mediated activation of TGF-β1/Mothers against decapentaplegic homolog 2/3 (Smad2/3) and EMT in lung tissues through up-regulating epithelial marker E-cadherin and down-regulating mesenchymal markers including fibronectin, vimentin and α-smooth muscle actin. Moreover, in vitro, iPS cell-conditioned medium (iPSC-CM) profoundly inhibited TGF-β1-induced EMT signaling pathway in mouse alveolar

  2. JNK1 and JNK2 regulate α-SMA in hepatic stellate cells during CCl4 -induced fibrosis in the rat liver.

    PubMed

    Hong, Il-Hwa; Park, Sang-Joon; Goo, Moon-Jung; Lee, Hye-Rim; Park, Jin-Kyu; Ki, Mi-Ran; Kim, Sang-Hyeob; Lee, Eun-Mi; Kim, Ah-Young; Jeong, Kyu-Shik

    2013-10-01

    Following liver injuries, hepatic stellate cells (HSCs) express α-SMA. Mitogen activated protein kinase (MAPK) signaling pathways mediate α-SMA expression in distinct cell types. However, the regulation of α-SMA expression by MAPKs in HSCs has been rarely studied. We aimed to study the role of MAPKs in the activation of HSCs during liver fibrosis. Liver fibrosis of rats was induced by carbon tetrachloride. HSC-T6 cells, murine embryonic fibroblasts, JNK1(-/-) and JNK2(-/-) cells were used for in vitro studies. Immunohistochemistry and immunoblot analysis were used. We have found that the expression of JNK and α-SMA co-localized in HSCs during liver fibrosis, but ERK and p38 expressed in macrophages. The expression of α-SMA was up-regulated by JNK1 and JNK2 in non-stress condition. Under TGF-β stimulation, however, the level α-SMA expression was increased by only JNK1, but not significantly changed by JNK2. We suggest that JNKs are responsible for α-SMA regulation, and especially JNK1 has a major role in up-regulation of α-SMA expression in HSCs under stress condition induced by TGF-β during liver fibrosis.

  3. Loss of discoidin domain receptor 2 promotes hepatic fibrosis after chronic carbon tetrachloride through altered paracrine interactions between hepatic stellate cells and liver-associated macrophages.

    PubMed

    Olaso, Elvira; Arteta, Beatriz; Benedicto, Aitor; Crende, Olatz; Friedman, Scott L

    2011-12-01

    Hepatic stellate cells (HSCs) interact with fibrillar collagen through the discoidin domain receptor 2 (DDR2) in acute hepatic injury, generating increased fibrosis. However, the contribution of DDR2 signaling to chronic liver fibrosis in vivo is unclear, despite its relevance to chronic human liver disease. We administered carbon tetrachloride (CCl(4)) to DDR2(+/+) and DDR2(-/-) mice twice weekly, and liver tissues and isolated HSCs were analyzed. In contrast to changes seen in acute injury, after chronic CCl(4) administration, DDR2(-/-) livers had increased collagen deposition, gelatinolytic activity, and HSC density. Increased basal gene expression of osteopontin, transforming growth factor-β1, monocyte chemoattractant protein-1, and IL-10 and reduced basal gene expression of matrix metalloproteinase-2, matrix metalloproteinase-13, and collagen type I in quiescent DDR2(-/-) HSCs were amplified further after chronic CCl(4). In concordance, DDR2(-/-) HSCs isolated from chronically injured livers had enhanced in vitro migration and proliferation, but less extracellular matrix degradative activity. Macrophages from chronic CCl(4)-treated DDR2(-/-) livers showed stronger chemoattractive activity toward DDR2(-/-) HSCs than DDR2(+/+) macrophages, increased extracellular matrix degradation, and higher cytokine mRNA expression. In conclusion, loss of DDR2 promotes chronic liver fibrosis after CCl(4) injury. The fibrogenic sinusoidal milieu generated in chronic DDR2(-/-) livers recruits more HSCs to injured regions, which enhances fibrosis. Together, these findings suggest that DDR2 normally orchestrates gene programs and paracrine interactions between HSCs and macrophages that together attenuate chronic hepatic fibrosis.

  4. Evaluation of TGF-β1 and MCP-1 expression and tubulointerstitial fibrosis in children with Henoch-Schönlein purpura nephritis and IgA nephropathy: A clinical correlation

    PubMed Central

    Shuiai, Zhao; Huijun, Shen; Weizhong, Gu; Aimin, Liu; Jianhua, Mao

    2017-01-01

    OBJECTIVES: Henoch-Schönlein purpura nephritis and immunoglobulin A nephropathy are two diseases with similar clinical presentations but very different prognoses. Transforming growth factor β1 and monocyte chemoattractant protein-1 have been associated with the development of tissue fibrosis. We examined the development of tubulointerstitial fibrosis and its relationship with Transforming growth factor β1 and monocyte chemoattractant protein-1 expression in these patients. METHODS: Renal tissue samples were collected by renal biopsy from 50 children with Henoch-Schönlein purpura nephritis and 50 children with immunoglobulin A nephropathy. Hematoxylin and eosin and Masson's trichrome-stained tissues were examined using light microscopy. Tubulointerstitial fibrosis was graded using the method described by Bohle et al. 1. The immunohistochemical detection of Transforming growth factor β1 and monocyte chemoattractant protein-1 expression was correlated with the tubulointerstitial fibrosis grade. Clinical Trial registration number: ZJCH-2012-0105. RESULTS: Transforming growth factor β1 and monocyte chemoattractant protein-1 expression in the renal tissues was significantly greater in the patients with immunoglobulin A nephropathy than in the patients with Henoch-Schönlein purpura nephritis (both p<0.001). The immunoglobulin A nephropathy patients had a higher tubulointerstitial fibrosis grade than the Henoch-Schönlein purpura nephritis patients (p<0.001). The tubulointerstitial fibrosis grade was in accordance with the Transforming growth factor β1 and monocyte chemoattractant protein-1 expression levels in both diseases (both p<0.001). CONCLUSION: Transforming growth factor β1 and monocyte chemoattractant protein-1 expression was associated with the development of immunoglobulin A nephropathy and Henoch-Schönlein purpura nephritis. Further studies are needed to better evaluate this association. PMID:28273242

  5. Morin ameliorates chemically induced liver fibrosis in vivo and inhibits stellate cell proliferation in vitro by suppressing Wnt/β-catenin signaling

    SciTech Connect

    MadanKumar, Perumal; NaveenKumar, Perumal; Manikandan, Samidurai; Devaraj, Halagowder; NiranjaliDevaraj, Sivasithamparam

    2014-06-01

    The anti-fibrotic effect of morin was examined in LX-2 cells (culture-activated human hepatic stellate cells) and in diethylnitrosamine induced rat model of liver fibrosis. The in vitro study was designed to determine whether morin affects the survival of cultured LX-2 cells, while the in vivo study was designed to evaluate the antioxidant and anti-fibrotic efficacy of morin on diethylnitrosamine induced liver fibrosis in male albino Wistar rat. The activities of liver function enzymes in serum, liver lipid peroxide levels, activities of serum antioxidant enzymes and liver architecture were monitored to cast light on the antioxidant and hepatoprotective nature of morin. To establish the anti-fibrotic effects of morin, the levels of key Wnt signaling molecules which are strongly associated with the signal transduction pathway of HSC activation were measured. Overall, from the in vitro results, it was observed that morin at 50 μM concentration inhibited the proliferation of cultured LX-2 cells, inhibited Wnt signaling and induced G1 cell cycle arrest. The in vivo results further confirmed that morin by downregulating the expressions of GSK-3β, β-catenin and cyclin D1 ameliorated DEN-induced liver fibrosis. Hence morin could be employed as a promising chemopreventive natural supplement for liver fibrosis. - Highlights: • In vivo and in vitro results revealed the active participation of Wnt signaling. • Morin at 50 μM inhibited LX-2 cell proliferation by suppressing Wnt signaling. • Morin exhibited hepatoprotective effects against DEN induced liver fibrosis. • Morin inhibited HSC activation in vivo by downregulating Wnt/β-catenin signaling.

  6. Achromobacter xylosoxidans genomic characterization and correlation of randomly amplified polymorphic DNA profiles with relevant clinical features [corrected] of cystic fibrosis patients.

    PubMed

    Magni, Annarita; Trancassini, Maria; Varesi, Paola; Iebba, Valerio; Curci, Anna; Pecoraro, Claudia; Cimino, Giuseppe; Schippa, Serena; Quattrucci, Serena

    2010-04-01

    Achromobacter xylosoxidans is an emerging pathogen increasingly being isolated from respiratory samples of cystic fibrosis (CF) patients. Its role and clinical significance in lung pathogenesis have not yet been clarified. The aim of the present study was to genetically characterize A. xylosoxidans strains isolated from CF patients by use of randomly amplified polymorphic DNA (RAPD) profiles and to look for a possible correlation between RAPD profiles and the patients' clinical features, such as their spirometry values, the presence of concomitant chronic bacterial flora at the time of isolation, and the persistent or intermittent presence of A. xylosoxidans strains. A set of 106 strains of A. xylosoxidans were typed by RAPD analysis, and their profiles were analyzed by agglomerative hierarchical classification (AHC) and associated with the patient characteristics mentioned above by factorial discriminant analysis (FDA). The overall results obtained in this study showed that (i) there is a marked genetic relationship between strains isolated from the same patients at different times, (ii) characteristic RAPD profiles are associated with different predicted classes for forced expiratory volume in 1 s (FEV1%), (iii) some characteristic RAPD profiles are associated with different concomitant chronic flora (CCF) profiles, and (iv) there is a significant division of RAPD profiles into "persistent strains" and "intermittent strains" of A. xylosoxidans. These findings seem to imply that the lung habitats found in CF patients are capable of shaping and selecting the colonizing bacterial flora, as seems to be the case for the A. xylosoxidans strains studied.

  7. Oxidative Stress and Pulmonary Fibrosis

    PubMed Central

    Cheresh, Paul; Kim, Seok-Jo; Tulasiram, Sandhya; Kamp, David W.

    2012-01-01

    Oxidative stress is implicated as an important molecular mechanism underlying fibrosis in a variety of organs, including the lungs. However, the causal role of reactive oxygen species (ROS) released from environmental exposures and inflammatory / interstitial cells in mediating fibrosis as well as how best to target an imbalance in ROS production in patients with fibrosis are not firmly established. We focus on the role of ROS in pulmonary fibrosis and, where possible, highlight overlapping molecular pathways in other organs. The key origins of oxidative stress in pulmonary fibrosis (e.g. environmental toxins, mitochondria / NADPH oxidase of inflammatory and lung target cells, and depletion of antioxidant defenses) are reviewed. The role of alveolar epithelial cell (AEC) apoptosis by mitochondria- and p53-regulated death pathways are examined. We emphasize an emerging role for the endoplasmic reticulum (ER) in pulmonary fibrosis. After briefly summarizing how ROS trigger a DNA damage response, we concentrate on recent studies implicating a role for mitochondrial DNA (mtDNA) damage and repair mechanisms focusing on 8-oxoguanine DNA glycosylase (Ogg1) as well as crosstalk between ROS production, mtDNA damage, p53, Ogg1, and mitochondrial aconitase (ACO2). Finally, the association between ROS and TGF-β1-induced fibrosis is discussed. Novel insights into the molecular basis of ROS-induced pulmonary diseases and, in particular, lung epithelial cell death may promote the development of unique therapeutic targets for managing pulmonary fibrosis as well as fibrosis in other organs and tumors, and in aging; diseases for which effective management is lacking. PMID:23219955

  8. Paracellular Transport through Healthy and Cystic Fibrosis Bronchial Epithelial Cell Lines – Do We Have a Proper Model?

    PubMed Central

    Weiser, Nelly; Kusche-Vihrog, Kristina; Günzel, Dorothee; Schillers, Hermann

    2014-01-01

    It has been reported recently that the cystic fibrosis transmembrane conductance regulator (CFTR) besides transcellular chloride transport, also controls the paracellular permeability of bronchial epithelium. The aim of this study was to test whether overexpressing wtCFTR solely regulates paracellular permeability of cell monolayers. To answer this question we used a CFBE41o– cell line transfected with wtCFTR or mutant F508del-CFTR and compered them with parental line and healthy 16HBE14o– cells. Transepithelial electrical resistance (TER) and paracellular fluorescein flux were measured under control and CFTR-stimulating conditions. CFTR stimulation significant decreased TER in 16HBE14o– and also in CFBE41o– cells transfected with wtCFTR. In contrast, TER increased upon stimulation in CFBE41o– cells and CFBE41o– cells transfected with F508del-CFTR. Under non-stimulated conditions, all four cell lines had similar paracellular fluorescein flux. Stimulation increased only the paracellular permeability of the 16HBE14o– cell monolayers. We observed that 16HBE14o– cells were significantly smaller and showed a different structure of cell-cell contacts than CFBE41o– and its overexpressing clones. Consequently, 16HBE14o– cells have about 80% more cell-cell contacts through which electrical current and solutes can leak. Also tight junction protein composition is different in ‘healthy’ 16HBE14o– cells compared to ‘cystic fibrosis’ CFBE41o– cells. We found that claudin-3 expression was considerably stronger in 16HBE14o– cells than in the three CFBE41o– cell clones and thus independent of the presence of functional CFTR. Together, CFBE41o– cell line transfection with wtCFTR modifies transcellular conductance, but not the paracellular permeability. We conclude that CFTR overexpression is not sufficient to fully reconstitute transport in CF bronchial epithelium. Hence, it is not recommended to use those cell lines to study CFTR

  9. Retinoic Acid Ameliorates Pancreatic Fibrosis and Inhibits the Activation of Pancreatic Stellate Cells in Mice with Experimental Chronic Pancreatitis via Suppressing the Wnt/β-Catenin Signaling Pathway

    PubMed Central

    Yin, Guojian; Fan, Yuting; Wu, Deqing; Qiu, Lei; Yu, Ge; Xing, Miao; Hu, Guoyong; Wang, Xingpeng; Wan, Rong

    2015-01-01

    Pancreatic fibrosis, a prominent feature of chronic pancreatitis (CP), induces persistent and permanent damage in the pancreas. Pancreatic stellate cells (PSCs) provide a major source of extracellular matrix (ECM) deposition during pancreatic injury, and persistent activation of PSCs plays a vital role in the progression of pancreatic fibrosis. Retinoic acid (RA), a retinoid, has a broad range of biological functions, including regulation of cell differentiation and proliferation, attenuating progressive fibrosis of multiple organs. In the present study, we investigated the effects of RA on fibrosis in experimental CP and cultured PSCs. CP was induced in mice by repetitive cerulein injection in vivo, and mouse PSCs were isolated and activated in vitro. Suppression of pancreatic fibrosis upon administration of RA was confirmed based on reduction of histological damage, α-smooth muscle actin (α-SMA) expression and mRNA levels of β-catenin, platelet-derived growth factor (PDGF)-Rβ transforming growth factor (TGF)-βRII and collagen 1α1 in vivo. Wnt 2 and β-catenin protein levels were markedly down-regulated, while Axin 2 expression level was up-regulated in the presence of RA, both in vivo and in vitro. Nuclear translation of β-catenin was significantly decreased following RA treatment, compared with cerulein-induced CP in mice and activated PSCs. Furthermore, RA induced significant PSC apoptosis, inhibited proliferation, suppressed TCF/LEF-dependent transcriptional activity and ECM production of PSC via down-regulation of TGFβRII, PDGFRβ and collagen 1α1 in vitro. These results indicate a critical role of the Wnt/β-catenin signaling pathway in RA-induced effects on CP and PSC regulation and support the potential of RA as a suppressor of pancreatic fibrosis in mice. PMID:26556479

  10. Melatonin Pretreatment Enhances the Homing of Bone Marrow-derived Mesenchymal Stem Cells Following Transplantation in a Rat Model of Liver Fibrosis

    PubMed Central

    Mortezaee, Keywan; Pasbakhsh, Parichehr; Kashani, Iraj Ragerdi; Sabbaghziarani, Fatemeh; Omidi, Ameneh; Zendedel, Adib; Ghasemi, Soudabeh; Dehpour, Ahmad Reza

    2016-01-01

    Background: Bone marrow-derived mesenchymal stem cells (BMMSCs) transplantation has been considered as a promising milestone in liver fibrosis treatment. However, low amounts of homing are a major obstacle. We aimed to investigate the role of melatonin pretreatment in BMMSC homing into experimental liver fibrosis. Methods: BMMSCs were obtained, grown, propagated and preconditioned with 5 µM melatonin and analyzed for multipotency and immunophenotypic features at passage three. The cells were labelled with CM-Dil and infused into the rats received the i.p. injection of carbon tetrachloride (CCl4) for five weeks to induce liver fibrosis. Animals were divided into two groups: One group received BMMSCs, whereas the other group received melatonin-pretreated BMMSCs (MT-BMMSCs). After cell injection at 72 h, animals were sacrificed, and the liver tissues were assessed for further evaluations: fibrosis using Masson’s trichrome and hematoxylin and eosin staining and homing using fluorescent microscopy and flow cytometry. Results: BMMSCs and MT-BMMSCs expressed a high level of CD44 but low levels of CD11b, CD45 and CD34 (for all P≤0.05) and were able to differentiate into adipocytes and Schwann cells. CCl4 induction resulted in extensive collagen deposition, tissue disruption and fatty accumulation with no obvious difference between the two groups. There was a significant increase in homing of MT-BMMSCs in both florescent microscopy (P≤0.001) and flow cytometry (P≤0.01) assays, as compared with non-treated BMMSCs. Conclusion: This study indicates the improved homing potential of BMMSCs in pretreatment with melatonin. Therefore, this strategy may represent an applied approach for improving the stem cell therapy of liver fibrosis. PMID:27130910

  11. Challenges in pulmonary fibrosis · 2 : Bronchiolocentric fibrosis

    PubMed Central

    Cordier, Jean‐François

    2007-01-01

    Bronchiolocentric fibrosis is essentially represented by the pathological pattern of constrictive fibrotic bronchiolitis obliterans. The corresponding clinical condition (obliterative bronchiolitis) is characterised by dyspnoea, airflow obstruction at lung function testing and air trapping with characteristic mosaic features on expiratory high resolution CT scans. Bronchiolitis obliterans may result from many causes including acute diffuse bronchiolar damage after inhalation of toxic gases or fumes, alloimmune chronic processes after lung or haematopoietic stem cell transplantation, or connective tissue disease (especially rheumatoid arthritis). Airway‐centred interstitial fibrosis and bronchiolar metaplasia are other features of bronchiolocentric fibrosis. PMID:17600295

  12. Cytokines in human lung fibrosis.

    PubMed

    Martinet, Y; Menard, O; Vaillant, P; Vignaud, J M; Martinet, N

    1996-01-01

    Fibrosis is a pathological process characterized by the replacement of normal tissue by mesenchymal cells and the extracellular matrix produced by these cells. The sequence of events leading to fibrosis of an organ involves the subsequent processes of injury with inflammation and disruption of the normal tissue architecture, followed by tissue repair with accumulation of mesenchymal cells in the area of derangement. The same sequence of events occurs in wound healing with normal granulation tissue and scar formation, but, while normal scar formation is very localized and transient, in contrast, in fibrosis, the repair process is exaggerated and usually widespread and can be chronic. Inflammatory cells (mainly mononuclear phagocytes), platelets, endothelial cells, and type II pneumocytes play a direct and indirect role in tissue injury and repair. The evaluation of three human fibrotic lung diseases, two diffuse [idiopathic pulmonary fibrosis (IPF), and the adult respiratory distress syndrome (ARDS)], and one focal (tumor stroma in lung cancer), has shown that several cytokines participate to the local injury and inflammatory reaction [interleukin-1 (IL-1), interleukin-8 (IL-8), monocyte chemotactic protein-1 (MCP-1), tumor necrosis factor-alpha (TNF-alpha)], while other cytokines are involved in tissue repair and fibrosis [platelet-derived growth factor (PDGF), insulin-like growth factor-1 (IGF-1), transforming growth factor-beta (TGF-beta), and basic-fibroblast growth factor (b-FGF)]. A better understanding of the cytokines and cytokine networks involved in lung fibrosis leads to the possibility of new therapeutic approaches.

  13. [Dynamic expression profile of HBsAg according to hepatic parenchyma cells' volume at different liver fibrosis stages in the immune clearance phase].

    PubMed

    Wu, Zhe-bin; Cao, Hong; Liu, Ting; Wu, Ze-qian; Ke, Wei-min; Gao, Zhi-liang

    2012-10-01

    The aim of this study was to determine the dynamic expression profile of hepatitis B surface antigen (HBsAg) according to hepatic parenchyma cells' volume at different stages of liver fibrosis during the immune clearance phase. Eighty-nine patients with HBeAg-positive chronic hepatitis B (CHB) in the immune clearance stage were recruited for study. Each patient's serum HBsAg levels were detected by electrochemiluminescence. The serum HBsAg levels were apportioned according to hepatic parenchyma cells' volume at liver fibrosis stages 1, 2, 3, and 4 and compared by ANOVA. The unapportioned serum HBsAg levels (IU/mL) at liver fibrosis stages 1 (227.2+/-237.7), 2 (211.0+/-131.4), 3(300.1+/-144.6), and 4 (278.7+/-148.8) were not significantly different (all comparisons, P range: 0.061 to 0.759). However, when the serum HBsAg levels were apportioned by the same hepatic parenchyma cells' volume at liver fibrosis stages 1 (343.9+/-359.8), 2 (336.4+/-209.5), 3 (508.7+/-245.1), and 4 (525.2+/-274.8), the levels were significantly different (all comparisons, F = 3.045 and P = 0.033; stage 1 vs. 3, P = 0.041; stage 1 vs. 4, P = 0.046; stage 2 vs. 3, P = 0.028; stage 2 vs. 4, P = 0.034). During the immune clearance phase of chronic hepatitis B, increased HBsAg expression is associated with increased hepatic parenchyma cells' volume and progressive liver fibrosis stage.

  14. Antihepatic Fibrosis Effect of Active Components Isolated from Green Asparagus (Asparagus officinalis L.) Involves the Inactivation of Hepatic Stellate Cells.

    PubMed

    Zhong, Chunge; Jiang, Chunyu; Xia, Xichun; Mu, Teng; Wei, Lige; Lou, Yuntian; Zhang, Xiaoshu; Zhao, Yuqing; Bi, Xiuli

    2015-07-08

    Green asparagus (Asparagus officinalis L.) is a vegetable with numerous nutritional properties. In the current study, a total of 23 compounds were isolated from green asparagus, and 9 of these compounds were obtained from this genus for the first time. Preliminary data showed that the ethyl acetate (EtOAc)-extracted fraction of green asparagus exerted a stronger inhibitory effect on the growth of t-HSC/Cl-6 cells, giving an IC50 value of 45.52 μg/mL. The biological activities of the different compounds isolated from the EtOAc-extracted fraction with respect to antihepatic fibrosis were investigated further. Four compounds, C3, C4, C10, and C12, exhibited profound inhibitory effect on the activation of t-HSC/Cl-6 cells induced by TNF-α. The activation t-HSC/Cl-6 cells, which led to the production of fibrotic matrix (TGF-β1, activin C) and accumulation of TNF-α, was dramatically decreased by these compounds. The mechanisms by which these compounds inhibited the activation of hepatic stellate cells appeared to be associated with the inactivation of TGF-β1/Smad signaling and c-Jun N-terminal kinases, as well as the ERK phosphorylation cascade.

  15. Trimethylangelicin promotes the functional rescue of mutant F508del CFTR protein in cystic fibrosis airway cells.

    PubMed

    Favia, Maria; Mancini, Maria T; Bezzerri, Valentino; Guerra, Lorenzo; Laselva, Onofrio; Abbattiscianni, Anna C; Debellis, Lucantonio; Reshkin, Stephan J; Gambari, Roberto; Cabrini, Giulio; Casavola, Valeria

    2014-07-01

    Cystic fibrosis transmembrane conductance regulator (CFTR) carrying the F508del mutation is retained in endoplasmic reticulum and fails to traffic to the cell surface where it functions as a protein kinase A (PKA)-activated chloride channel. Pharmacological correctors that rescue the trafficking of F508del CFTR may overcome this defect; however, the rescued F508del CFTR still displays reduced chloride permeability. Therefore, a combined administration of correctors and potentiators of the gating defect is ideal. We recently found that 4,6,4'-trimethylangelicin (TMA), besides inhibiting the expression of the IL-8 gene in airway cells in which the inflammatory response was challenged with Pseudomonas aeruginosa, also potentiates the cAMP/PKA-dependent activation of wild-type CFTR or F508del CFTR that has been restored to the plasma membrane. Here, we demonstrate that long preincubation with nanomolar concentrations of TMA is able to effectively rescue both F508del CFTR-dependent chloride secretion and F508del CFTR cell surface expression in both primary or secondary airway cell monolayers homozygous for F508del mutation. The correction effect of TMA seems to be selective for CFTR and persisted for 24 h after washout. Altogether, the results suggest that TMA, besides its anti-inflammatory and potentiator activities, also displays corrector properties.

  16. Cultured Mycelium Cordyceps sinensis allevi¬ates CCl4-induced liver inflammation and fibrosis in mice by activating hepatic natural killer cells

    PubMed Central

    Peng, Yuan; Huang, Kai; Shen, Li; Tao, Yan-yan; Liu, Cheng-hai

    2016-01-01

    Aim: Recent evidence shows that cultured mycelium Cordyceps sinensis (CMCS) effectively protects against liver fibrosis in mice. Here, we investigated whether the anti-fibrotic action of CMCS was related to its regulation of the activity of hepatic natural killer (NK) cells in CCl4-treated mice. Methods: C57BL/6 mice were injected with 10% CCl4 (2 mL/kg, ip) 3 times per week for 4 weeks, and received CMCS (120 mg·kg−1·d−1, ig) during this period. In another part of experiments, the mice were also injected with an NK cell-deleting antibody ASGM-1 (20 μg, ip) 5 times in the first 3 weeks. After the mice were sacrificed, serum liver function, and liver inflammation, hydroxyproline content and collagen deposition were assessed. The numbers of hepatic NK cells and expression of NKG2D (activation receptor of NK cells) on isolated liver lymphocytes were analyzed using flow cytometry. Desmin expression and cell apoptosis in liver tissues were studied using desmin staining and TUNEL assay, respectively. The levels of α-SMA, TGF-β, RAE-1δ and RAE-1ε in liver tissues were determined by RT-qPCR. Results: In CCl4-treated mice, CMCS administration significantly improved liver function, attenuated liver inflammation and fibrosis, and increased the numbers of hepatic NK cells and expression level of NKG2D on hepatic NK cells. Furthermore, CMCS administration significantly decreased desmin expression in liver tissues, and increased TUNEL staining adjacent to hepatic stellate cells. Injection with NK cell-deleting ASGM-1 not only diminished the numbers of hepatic NK cells, but also greatly accelerated liver inflammation and fibrosis in CCl4-treated mice. In CCl4-treated mice with NK cell depletion, CMCS administration decelerated the rate of liver fibrosis development, and mildly upregulated the numbers of hepatic NK cells but without changing NKG2D expression. Conclusion: CMCS allevi¬ates CCl4-induced liver inflammation and fibrosis via promoting activation of hepatic

  17. Loss of lysosomal membrane protein NCU-G1 in mice results in spontaneous liver fibrosis with accumulation of lipofuscin and iron in Kupffer cells

    PubMed Central

    Kong, Xiang Y.; Nesset, Cecilie Kasi; Damme, Markus; Løberg, Else-Marit; Lübke, Torben; Mæhlen, Jan; Andersson, Kristin B.; Lorenzo, Petra I.; Roos, Norbert; Thoresen, G. Hege; Rustan, Arild C.; Kase, Eili T.; Eskild, Winnie

    2014-01-01

    Human kidney predominant protein, NCU-G1, is a highly conserved protein with an unknown biological function. Initially described as a nuclear protein, it was later shown to be a bona fide lysosomal integral membrane protein. To gain insight into the physiological function of NCU-G1, mice with no detectable expression of this gene were created using a gene-trap strategy, and Ncu-g1gt/gt mice were successfully characterized. Lysosomal disorders are mainly caused by lack of or malfunctioning of proteins in the endosomal-lysosomal pathway. The clinical symptoms vary, but often include liver dysfunction. Persistent liver damage activates fibrogenesis and, if unremedied, eventually leads to liver fibrosis/cirrhosis and death. We demonstrate that the disruption of Ncu-g1 results in spontaneous liver fibrosis in mice as the predominant phenotype. Evidence for an increased rate of hepatic cell death, oxidative stress and active fibrogenesis were detected in Ncu-g1gt/gt liver. In addition to collagen deposition, microscopic examination of liver sections revealed accumulation of autofluorescent lipofuscin and iron in Ncu-g1gt/gt Kupffer cells. Because only a few transgenic mouse models have been identified with chronic liver injury and spontaneous liver fibrosis development, we propose that the Ncu-g1gt/gt mouse could be a valuable new tool in the development of novel treatments for the attenuation of fibrosis due to chronic liver damage. PMID:24487409

  18. Wharton's jelly-derived mesenchymal stem cells combined with praziquantel as a potential therapy for Schistosoma mansoni-induced liver fibrosis.

    PubMed

    Hammam, Olfat A; Elkhafif, Nagwa; Attia, Yasmeen M; Mansour, Mohamed T; Elmazar, Mohamed M; Abdelsalam, Rania M; Kenawy, Sanaa A; El-Khatib, Aiman S

    2016-02-15

    Liver fibrosis is one of the most serious consequences of S. mansoni infection. The aim of the present study was to investigate the potential anti-fibrotic effect of human Wharton's jelly-derived mesenchymal stem cells (WJMSCs) combined with praziquantel (PZQ) in S. mansoni-infected mice. S. mansoni-infected mice received early (8(th) week post infection) and late (16(th) week post infection) treatment with WJMSCs, alone and combined with oral PZQ. At the 10(th) month post infection, livers were collected for subsequent flow cytometric, histopathological, morphometric, immunohistochemical, gene expression, and gelatin zymographic studies. After transplantation, WJMSCs differentiated into functioning liver-like cells as evidenced by their ability to express human hepatocyte-specific markers. Regression of S. mansoni-induced liver fibrosis was also observed in transplanted groups, as evidenced by histopathological, morphometric, and gelatin zymographic results besides decreased expression of three essential contributors to liver fibrosis in this particular model; alpha smooth muscle actin, collagen-I, and interleukin-13. PZQ additionally enhanced the beneficial effects observed in WJMSCs-treated groups. Our results suggest that combining WJMSCs to PZQ caused better enhancement in S. mansoni-induced liver fibrosis, compared to using each alone.

  19. Cyanidin-3-O-β-glucoside Purified from Black Rice Protects Mice against Hepatic Fibrosis Induced by Carbon Tetrachloride via Inhibiting Hepatic Stellate Cell Activation.

    PubMed

    Jiang, Xinwei; Guo, Honghui; Shen, Tianran; Tang, Xilan; Yang, Yan; Ling, Wenhua

    2015-07-15

    This study investigated whether cyanidin-3-O-β-glucoside (Cy-3-G), a predominant anthocyanin, could exert a protective role on liver injury and its further mechanisms of the anti-fibrosis actions in mice. The results demonstrated that the treatment of Cy-3-G (800 mg/kg diet) for 8 weeks significantly attenuated hepatotoxicity and fibrosis in carbon tetrachloride (CCl4) administered mice. Cy-3-G strongly down-regulated the expression of α-smooth muscle actin (α-SMA), desmin, and matrix metalloproteinase (MMPs), which showed its suppression effect on the activation of hepatic stellate cells (HSCs). In addition, Cy-3-G favorably regulated oxidative stress and apoptosis in liver. Furthermore, Cy-3-G ameliorated the infiltration of inflammatory cells such as neutrophils and leukocytes and meanwhile suppressed the production of pro-inflammatory cytokines and growth factors. In conclusion, daily intake of Cy-3-G could prevent liver fibrosis progression in mice induced by CCl4 through inhibiting HSC activation, which provides a basis for clinical practice of liver fibrosis prevention.

  20. Maresin 1 Mitigates High Glucose-Induced Mouse Glomerular Mesangial Cell Injury by Inhibiting Inflammation and Fibrosis

    PubMed Central

    Tang, Shi; Long, Yang; Huang, Wei; Chen, Jiao; Fan, Fang; Jiang, Chunxia

    2017-01-01

    Background. Inflammation and fibrosis are the important pathophysiologic processes in diabetic nephropathy (DN). Maresin 1 is a potential anti-inflammatory lipid mediator, which has displayed powerful proresolving activities. Aim. We determine whether maresin 1 has protective effect on mouse glomerular mesangial cells (GMCs) induced by high glucose. Methods. We cultured GMCs stimulated by high glucose and categorized as follows: normal glucose group (5.6 mmol/L), high glucose group (30 mmol/L), mannitol group, maresin 1 intervention group (1, 10, and 100 nmol/L), maresin 1 and normal glucose group, and the N-acetylcysteine (NAC) intervention group (10 μmol/L NAC). After 24 h, the expression of ROS, NLRP3, caspase-1, procaspase-1, IL-1β, and pro-IL-1β was detected by western-blot, RT-PCR, and immunofluorescence. After 48 h, the expression of TGF-β1 and FN was detected by RT-PCR and ELISA. Results. Compared with normal glucose group, the expression of ROS, NLRP3, caspase-1, IL-1β, TGF-β1, and FN increased in high glucose group (P < 0.05), but it decreased after the treatment of maresin 1 in different concentrations. On the contrary, the expression of procaspase-1 and pro-IL-1β protein was restrained by high glucose and enhanced by maresin 1 in a dose-dependent manner (P < 0.05). Conclusion. Maresin 1 can inhibit NLRP3 inflammasome, TGF-β1, and FN in GMCs; it may have protective effect on DN by mitigating the inflammation and early fibrosis. PMID:28182085

  1. Mesenchymal stem cells modulate albumin-induced renal tubular inflammation and fibrosis.

    PubMed

    Wu, Hao Jia; Yiu, Wai Han; Li, Rui Xi; Wong, Dickson W L; Leung, Joseph C K; Chan, Loretta Y Y; Zhang, Yuelin; Lian, Qizhou; Lin, Miao; Tse, Hung Fat; Lai, Kar Neng; Tang, Sydney C W

    2014-01-01

    Bone marrow-derived mesenchymal stem cells (BM-MSCs) have recently shown promise as a therapeutic tool in various types of chronic kidney disease (CKD) models. However, the mechanism of action is incompletely understood. As renal prognosis in CKD is largely determined by the degree of renal tubular injury that correlates with residual proteinuria, we hypothesized that BM-MSCs may exert modulatory effects on renal tubular inflammation and epithelial-to-mesenchymal transition (EMT) under a protein-overloaded milieu. Using a co-culture model of human proximal tubular epithelial cells (PTECs) and BM-MSCs, we showed that concomitant stimulation of BM-MSCs by albumin excess was a prerequisite for them to attenuate albumin-induced IL-6, IL-8, TNF-α, CCL-2, CCL-5 overexpression in PTECs, which was partly mediated via deactivation of tubular NF-κB signaling. In addition, albumin induced tubular EMT, as shown by E-cadherin loss and α-SMA, FN and collagen IV overexpression, was also prevented by BM-MSC co-culture. Albumin-overloaded BM-MSCs per se retained their tri-lineage differentiation capacity and overexpressed hepatocyte growth factor (HGF) and TNFα-stimulating gene (TSG)-6 via P38 and NF-κB signaling. Albumin-induced tubular CCL-2, CCL-5 and TNF-α overexpression were suppressed by recombinant HGF treatment, while the upregulation of α-SMA, FN and collagen IV was attenuated by recombinant TSG-6. Neutralizing HGF and TSG-6 abolished the anti-inflammatory and anti-EMT effects of BM-MSC co-culture in albumin-induced PTECs, respectively. In vivo, albumin-overloaded mice treated with mouse BM-MSCs had markedly reduced BUN, tubular CCL-2 and CCL-5 expression, α-SMA and collagen IV accumulation independent of changes in proteinuria. These data suggest anti-inflammatory and anti-fibrotic roles of BM-MSCs on renal tubular cells under a protein overloaded condition, probably mediated via the paracrine action of HGF and TSG-6.

  2. Dicliptera Chinensis polysaccharides target TGF-β/Smad pathway and inhibit stellate cells activation in rats with dimethylnitrosamine-induced hepatic fibrosis.

    PubMed

    Zhang, X; Zhang, J; Jia, L; Xiao, S

    2016-01-27

    This study aims to study impact of Dicliptera chinensis polysaccharide (DCP) on hepatic fibrosis (HF) and activation of hepatic stellate cells (HSCs). Liver fibrosis model was induced by intraperitoneal injection of dimethyl nitrosamines (DMN) in rat. Rats in treatment group were administrated with different concentrations of DCP (0, 100, 300 mg/kg) by intraperitoneal injection. Hematoxylin and eosin (H&E) and Masson's trichrome staining were used to assess histo-pathological change. α-SMA, TGF-β1 and pSmad 2/3 were assayed by immuno-histochemistry. HSC-T6 cells were stimulated by recombined rat TGF-β1 (1 ng/mL) to simulate an activating model in vitro and then interfered with DCP (concentration of 0, 25, 50, 100, 200, 400 µg/ml). MTT assay was used to determine cell proliferation and western blotting was used to detect α-SMA and pSmad 2/3 expression. Results demonstrated that DCP alleviated DMN-induced liver fibrosis in rat and significantly down-regulated TGF-β1 expression, pSmad2/3 and α-SMA in liver tissue in a dose-dependent way. DCP inhibited proliferation and activation of TGF-β1-stimulated HSC-T6 in vitro and significantly down-regulated α-SMA and pSmad2/3 expression. In conclusion, this study revealed that DCP attenuates progression of liver fibrosis through suppressing TGF-β/Smad pathway. DCP is a potential botanical polysaccharide to management liver fibrosis.

  3. Allelic drop-out and preferential amplification in single cells and human blastomeres: implications for preimplantation diagnosis of sex and cystic fibrosis.

    PubMed

    Findlay, I; Ray, P; Quirke, P; Rutherford, A; Lilford, R

    1995-06-01

    Previously the diagnosis of sex and cystic fibrosis status has been studied on single cells using the polymerase chain reaction (PCR). It has been suggested that allelic drop-out (PCR failure of one allele) and/or preferential amplification (hypo-amplification of one allele) may contribute to poor reliability and misdiagnosis, although this remains controversial as some reports suggest that allelic drop-out does not occur. We investigated an improved method of diagnosing sex and cystic fibrosis in single cells using a new technology (fluorescent PCR) to determine the base level of PCR artefacts (allelic drop-out and preferential amplification) which, in combination with improved sensitivity, should improve PCR reliability and accuracy. Fluorescent PCR gives high reliability (approximately 97%) and accuracy rates (approximately 97%) in somatic cells for both sex and cystic fibrosis diagnosis and its lower detection threshold allows allelic drop-out and preferential amplification to be easily distinguished. We also achieved high reliability and accuracy in diagnosing cystic fibrosis in human blastomeres. This study confirms earlier reports of both allelic drop-out and preferential amplification in single cell analysis. We demonstrate that both allelic drop-out and preferential amplification occur in somatic cells and suggest these are separate phenomena. Preferential amplification appeared common in single cell PCR while allelic drop-out apparently occurred at random in each allele. Preferential amplification was mainly amplification of the larger allele. We suggest that some inaccuracy/misdiagnosis may be due to both preferential amplification as well as allelic drop-out. Other findings were variability in drop-out between PCR and that amplification of signals from human blastomeres may be linked to embryo quality. We suggest that allelic drop-out is dependent on the number of cells within the sample.

  4. Long non-coding RNA APTR promotes the activation of hepatic stellate cells and the progression of liver fibrosis

    SciTech Connect

    Yu, Fujun; Zheng, Jianjian; Mao, Yuqing; Dong, Peihong; Li, Guojun; Lu, Zhongqiu; Guo, Chuanyong; Liu, Zhanju; Fan, Xiaoming

    2015-08-07

    In this study, we aimed at assessing a role of Alu-mediated p21 transcriptional regulator (APTR) in hepatofibrogenesis. APTR was upregulated in fibrotic liver samples and activated hepatic stellate cells (HSCs). Knockdown of APTR inhibited the activation of HSCs in vitro and mitigated the accumulation of collagen in vivo. Importantly, APTR silencing could abrogate TGF-β{sub 1}-induced upregulation of α-SMA in HSCs. In addition, inhibition of cell cycle and cell proliferation by APTR knockdown was attenuated by p21 siRNA1 in primary HSCs. Finally, serum APTR levels were increased in patients with liver cirrhosis, indicating a potential biomarker for liver cirrhosis. Collectively, evidence is proposed for a new biological role of APTR in hepatofibrogenesis. - Highlights: • APTR is upregulated in fibrotic liver tissues and activated HSCs. • APTR silencing inhibits HSC activation and the progression of liver fibrosis. • Antifibrotic effect of APTR silencing is achieved by increasing p21.

  5. [Retroperitoneal fibrosis].

    PubMed

    Babski, Paweł; Wojtuń, Stanisław; Gil, Jerzy

    2007-05-01

    Retroperitoneal fibrosis is a rare clinical entity characterised by the presence of patologic collagen tissue in a retroperitoneal space. The fibrous mass covers abdominal organs causing their disfunctions. RPF was described at the begining of XX century but its etiology is not clear yet. Usually it causes an ureter obstuction and hydronephrosis, that is why most commonly is diagnosed by urologists and nephrologists. However, retroperitoneal fibrosis can be multifacial disease. In some patients localisation of fibrosis is atypical and manifestationns can be varied. Gastrological symptoms like jaundice, bowel obstuction, ascites can occure. Besides, some early signs of RPF are nonspecific and can imitate alarming symptoms of neoplasma, e.g.: weight loss, anemia, malaise, anorexia, fever. This force us to initiate gastrological investigation. The awareness of this disease is important. The early diagnosis and treatment improves prognosis and alows to avoid heavy complications. In typical cases radiology is often enough for diagnosis. However, histological examination is needed in many cases, especialy when patological mass is located atypical. A treatment is made up of farmacology and surgery. The first one is based on steroids, immunossuppressant and tamoxifen. Surgery is needed to eliminate organs obstruction.

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

    PubMed

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

    2017-02-01

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

  7. Mouse Mast Cell Protease-4 Deteriorates Renal Function by Contributing to Inflammation and Fibrosis in Immune Complex-Mediated Glomerulonephritis

    PubMed Central

    Scandiuzzi, Lisa; Beghdadi, Walid; Daugas, Eric; Åbrink, Magnus; Tiwari, Neeraj; Brochetta, Cristiana; Claver, Julien; Arouche, Nassim; Zang, Xingxing; Pretolani, Marina; Monteiro, Renato C.; Pejler, Gunnar; Blank, Ulrich

    2010-01-01

    Mast cells exert protective effects in experimental antiglomerular basement membrane-induced glomerulonephritis (GN), yet the responsible mediators have not been identified. In this study, we investigated the role of mouse mast cell protease (mMCP)-4, the functional homolog of human chymase, using mMCP-4–deficient mice. Compared with wild type animals, mMCP-4–deficient mice exhibited lower proteinuria, blood creatinine, and blood urea nitrogen levels, indicating an aggravating role of mMCP-4. Kidney histology confirmed less severe renal damage in mMCP-4–deficient mice with reduced deposits, glomerular and interstitial cellularity, and fibrosis scores. High amounts of mMCP-4 were detected in renal capsules, but not in the whole kidney, from wild type mice. Its expression in renal capsules was markedly decreased after GN induction, suggesting that locally released enzyme by degranulated mast cells could contribute to the functional and physiopathological hallmarks of GN. Supporting a proinflammatory role, glomerular and interstitial macrophage and T cell infiltration, levels of proinflammatory TNF and MCP-1 mRNA, and the expression of the profibrotic peptide angiotensin II together with type I collagen were markedly down-regulated in kidneys of mMCP-4–deficient mice. We conclude that mMCP-4 chymase, contrary to the global anti-inflammatory action of mast cells, aggravates GN by promoting kidney inflammation. These results highlight the complexity of mast cell-mediated inflammatory actions and suggest that chymase inhibition may represent a novel therapeutic target in GN. PMID:20530261

  8. Hepatic stellate cell-targeted delivery of hepatocyte growth factor transgene via bile duct infusion enhances its expression at fibrotic foci to regress dimethylnitrosamine-induced liver fibrosis.

    PubMed

    Narmada, Balakrishnan Chakrapani; Kang, Yuzhan; Venkatraman, Lakshmi; Peng, Qiwen; Sakban, Rashidah Binte; Nugraha, Bramasta; Jiang, Xuan; Bunte, Ralph M; So, Peter T C; Tucker-Kellogg, Lisa; Mao, Hai-Quan; Yu, Hanry

    2013-05-01

    Liver fibrosis generates fibrotic foci with abundant activated hepatic stellate cells and excessive collagen deposition juxtaposed with healthy regions. Targeted delivery of antifibrotic therapeutics to hepatic stellate cells (HSCs) might improve treatment outcomes and reduce adverse effects on healthy tissue. We delivered the hepatocyte growth factor (HGF) gene specifically to activated hepatic stellate cells in fibrotic liver using vitamin A-coupled liposomes by retrograde intrabiliary infusion to bypass capillarized hepatic sinusoids. The antifibrotic effects of DsRed2-HGF vector encapsulated within vitamin A-coupled liposomes were validated by decreases in fibrotic markers in vitro. Fibrotic cultures transfected with the targeted transgene showed a significant decrease in fibrotic markers such as transforming growth factor-β1. In rats, dimethylnitrosamine-induced liver fibrosis is manifested by an increase in collagen deposition and severe defenestration of sinusoidal endothelial cells. The HSC-targeted transgene, administered via retrograde intrabiliary infusion in fibrotic rats, successfully reduced liver fibrosis markers alpha-smooth muscle actin and collagen, accompanied by an increase in the expression of DsRed2-HGF near the fibrotic foci. Thus, targeted delivery of HGF gene to hepatic stellate cells increased the transgene expression at the fibrotic foci and strongly enhanced its antifibrotic effects.

  9. Lack of optimal T-cell reactivity against the hepatitis C virus is associated with the development of fibrosis/cirrhosis during chronic hepatitis.

    PubMed

    Sreenarasimhaiah, Jayaprakash; Jaramillo, Andrés; Crippin, Jeffrey; Lisker-Melman, Mauricio; Chapman, William C; Mohanakumar, T

    2003-02-01

    Chronic hepatitis C virus (HCV) infection develops in 85% of exposed individuals and 20% develop cirrhosis. However, the pathogenesis of this process is not well-understood. The objective of this study was to determine whether HCV-reactive T cells play a role in the process of development of cirrhosis during chronic HCV infection. We analyzed 21 human leukocyte antigen (HLA)-A2 patients with chronic HCV infection (9 with histology of inflammation and 12 with histology of fibrosis/cirrhosis). The frequency of CD8(+) T cells reactive to 12 HCV-derived epitopes was determined by an interferon-gamma enzyme-linked immunospot (ELISPOT) assay. The frequency of CD4(+) Th1 and Th2 cells reactive to the HCV core antigen was determined by interferon-gamma and interleukin-5 ELISPOT assays, respectively. Patients with histology of inflammation showed a significantly higher CD8(+) T-cell response to five HCV-derived epitopes (YLLPRRGPRL [core], CINGVCWTV [NS3], LLCPAGHAV [NS3], ILAGYGAGV [NS4B], and GLQDCTMLV [NS5B]) as compared with patients with histology of fibrosis/cirrhosis. Also, patients with histology of inflammation showed a significantly higher CD4(+) Th1 response to the HCV core antigen as compared to patients with histology of fibrosis/cirrhosis. These results indicate that a lack of an optimal T-cell response to HCV is associated with the development of cirrhosis during chronic HCV infection.

  10. Epithelial transglutaminase 2 is needed for T cell interleukin-17 production and subsequent pulmonary inflammation and fibrosis in bleomycin-treated mice

    PubMed Central

    Oh, Keunhee; Park, Hyung-Bae; Byoun, Ok-Jin; Shin, Dong-Myung; Jeong, Eui Man; Kim, Young Whan; Kim, Yon Su; Melino, Gerry

    2011-01-01

    Pulmonary fibrosis is a potentially life-threatening disease that may be caused by overt or asymptomatic inflammatory responses. However, the precise mechanisms by which tissue injury is translated into inflammation and consequent fibrosis remain to be established. Here, we show that in a lung injury model, bleomycin induced the secretion of IL-6 by epithelial cells in a transglutaminase 2 (TG2)–dependent manner. This response represents a key step in the differentiation of IL-17–producing T cells and subsequent inflammatory amplification in the lung. The essential role of epithelial cells, but not inflammatory cells, TG2 was confirmed in bone marrow chimeras; chimeras made in TG2-deficient recipients showed reduced inflammation and fibrosis, compared with those in wild-type mice, regardless of the bone marrow cell phenotype. Epithelial TG2 thus appears to be a critical inducer of inflammation after noninfectious pulmonary injury. We further demonstrated that fibroblast-derived TG2, acting downstream of transforming growth factor-β, is also important in the effector phase of fibrogenesis. Therefore, TG2 represents an interesting potential target for therapeutic intervention. PMID:21746810

  11. ADAM17 substrate release in proximal tubule drives kidney fibrosis

    PubMed Central

    Kefaloyianni, Eirini; Muthu, Muthu Lakshmi; Kaeppler, Jakob; Sun, Xiaoming; Sabbisetti, Venkata; Chalaris, Athena; Rose-John, Stefan; Wong, Eitan; Sagi, Irit; Waikar, Sushrut S.; Rennke, Helmut; Bonventre, Joseph V.

    2016-01-01

    Kidney fibrosis following kidney injury is an unresolved health problem and causes significant morbidity and mortality worldwide. In a study into its molecular mechanism, we identified essential causative features. Acute or chronic kidney injury causes sustained elevation of a disintegrin and metalloprotease 17 (ADAM17); of its cleavage-activated proligand substrates, in particular of pro-TNFα and the EGFR ligand amphiregulin (pro-AREG); and of the substrates’ receptors. As a consequence, EGFR is persistently activated and triggers the synthesis and release of proinflammatory and profibrotic factors, resulting in macrophage/neutrophil ingress and fibrosis. ADAM17 hypomorphic mice, specific ADAM17 inhibitor–treated WT mice, or mice with inducible KO of ADAM17 in proximal tubule (Slc34a1-Cre) were significantly protected against these effects. In vitro, in proximal tubule cells, we show that AREG has unique profibrotic actions that are potentiated by TNFα-induced AREG cleavage. In vivo, in acute kidney injury (AKI) and chronic kidney disease (CKD, fibrosis) patients, soluble AREG is indeed highly upregulated in human urine, and both ADAM17 and AREG expression show strong positive correlation with fibrosis markers in related kidney biopsies. Our results indicate that targeting of the ADAM17 pathway represents a therapeutic target for human kidney fibrosis. PMID:27642633

  12. Role of bone marrow-derived CD11c(+) dendritic cells in systolic overload-induced left ventricular inflammation, fibrosis and hypertrophy.

    PubMed

    Wang, Huan; Kwak, Dongmin; Fassett, John; Liu, Xiaohong; Yao, Wu; Weng, Xinyu; Xu, Xin; Xu, Yawei; Bache, Robert J; Mueller, Daniel L; Chen, Yingjie

    2017-05-01

    Inflammatory responses play an important role in the development of left ventricular (LV) hypertrophy and dysfunction. Recent studies demonstrated that increased T-cell infiltration and T-cell activation contribute to LV hypertrophy and dysfunction. Dendritic cells (DCs) are professional antigen-presenting cells that orchestrate immune responses, especially by modulating T-cell function. In this study, we investigated the role of bone marrow-derived CD11c(+) DCs in transverse aortic constriction (TAC)-induced LV fibrosis and hypertrophy in mice. We observed that TAC increased the number of CD11c(+) cells and the percentage of CD11c(+) MHCII(+) (major histocompatibility complex class II molecule positive) DCs in the LV, spleen and peripheral blood in mice. Using bone marrow chimeras and an inducible CD11c(+) DC ablation model, we found that depletion of bone marrow-derived CD11c(+) DCs significantly attenuated LV fibrosis and hypertrophy in mice exposed to 24 weeks of moderate TAC. CD11c(+) DC ablation significantly reduced TAC-induced myocardial inflammation as indicated by reduced myocardial CD45(+) cells, CD11b(+) cells, CD8(+) T cells and activated effector CD8(+)CD44(+) T cells in LV tissues. Moreover, pulsing of autologous DCs with LV homogenates from TAC mice promoted T-cell proliferation. These data indicate that bone marrow-derived CD11c(+) DCs play a maladaptive role in hemodynamic overload-induced cardiac inflammation, hypertrophy and fibrosis through the presentation of cardiac self-antigens to T cells.

  13. Familial Pulmonary Fibrosis

    MedlinePlus

    ... Training Home Conditions Familial Pulmonary Fibrosis Familial Pulmonary Fibrosis Make an Appointment Find a Doctor Ask a ... members within the same family have Idiopathic Pulmonary Fibrosis (IPF) or any other form of Idiopathic Interstitial ...

  14. Single-cell analysis reveals IGF-1 potentiation of inhibition of the TGF-β/Smad pathway of fibrosis in human keratocytes in vitro

    PubMed Central

    Sarenac, Tomislav; Trapecar, Martin; Gradisnik, Lidija; Rupnik, Marjan Slak; Pahor, Dusica

    2016-01-01

    Corneal wound healing is often affected by TGF-β–mediated fibrosis and scar formation. Guided fibrosis with IGF-1 and antifibrotic substances might maintain corneal transparency. Primary human corneal keratocytes under serum-free conditions were used as a model of corneal stromal wounding, with markers of corneal fibrosis and opacity studied under TGF-β2 stimulation. Single-cell imaging flow cytometry was used to determine nuclearization of Smad3, and intracellular fluorescence intensity of Smad7 and the corneal crystallin aldehyde dehydrogenase 3A1. Extracellular matrix proteoglycans keratocan and biglycan were quantified using ELISAs. On the TGF-β2 background, the keratocytes were treated with IGF-1, and suberoylanilidehydroxamic acid (SAHA) or halofuginone ± IGF-1. IGF-1 alone decreased Smad3 nuclearization and increased aldehyde dehydrogenase 3A1 expression, with favorable extracellular matrix proteoglycan composition. SAHA induced higher Smad7 levels and inhibited translocation of Smad3 to the nucleus, also when combined with IGF-1. Immunofluorescence showed that myofibroblast transdifferentiation is attenuated and appearance of fibroblasts is favored by IGF-1 alone and in combination with the antifibrotic substances. The TGF-β/Smad pathway of fibrosis and opacity was inhibited by IGF-1, and further with SAHA in particular, and with halofuginone. IGF-1 is thus a valid aid to antifibrotic treatment, with potential for effective and transparent corneal wound healing. PMID:27687492

  15. Dense transcript profiling in single cells by image correlation decoding

    PubMed Central

    Coskun, Ahmet F.; Cai, Long

    2016-01-01

    Recent work in sequential fluorescent in-situ hybridization (FISH) has demonstrated the ability to uniquely encode a large number of molecular species in single cells. However, the multiplexing capacity is practically limited by the density of the barcoded objects in the cell. Here, we present a general method using image correlation to resolve the temporal barcodes in sequential hybridization experiments, allowing high density objects to be decoded. Using this correlation FISH (corrFISH) approach, we profiled the gene expression of ribosomal proteins in single cells in cell cultures and in mouse thymus tissue sections. In tissues, corrFISH revealed cell type specific gene expression of ribosomal proteins. The combination of sequential barcoding FISH and correlation analyses provides a general strategy for multiplexing a large number of RNA molecules and potentially other high copy number molecules in single cells. PMID:27271198

  16. Weak correlation of starch and volume in synchronized photosynthetic cells

    NASA Astrophysics Data System (ADS)

    Rading, M. Michael; Sandmann, Michael; Steup, Martin; Chiarugi, Davide; Valleriani, Angelo

    2015-01-01

    In cultures of unicellular algae, features of single cells, such as cellular volume and starch content, are thought to be the result of carefully balanced growth and division processes. Single-cell analyses of synchronized photoautotrophic cultures of the unicellular alga Chlamydomonas reinhardtii reveal, however, that the cellular volume and starch content are only weakly correlated. Likewise, other cell parameters, e.g., the chlorophyll content per cell, are only weakly correlated with cell size. We derive the cell size distributions at the beginning of each synchronization cycle considering growth, timing of cell division and daughter cell release, and the uneven division of cell volume. Furthermore, we investigate the link between cell volume growth and starch accumulation. This work presents evidence that, under the experimental conditions of light-dark synchronized cultures, the weak correlation between both cell features is a result of a cumulative process rather than due to asymmetric partition of biomolecules during cell division. This cumulative process necessarily limits cellular similarities within a synchronized cell population.

  17. Role of bone marrow cells in the development of pancreatic fibrosis in a rat model of pancreatitis induced by a choline-deficient/ethionine-supplemented diet

    SciTech Connect

    Akita, Shingo; Kubota, Koji; Kobayashi, Akira; Misawa, Ryosuke; Shimizu, Akira; Nakata, Takenari; Yokoyama, Takahide; Takahashi, Masafumi; Miyagawa, Shinichi

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer BMC-derived PSCs play a role in a rat CDE diet-induced pancreatitis model. Black-Right-Pointing-Pointer BMC-derived PSCs contribute mainly to the early stage of pancreatic fibrosis. Black-Right-Pointing-Pointer BMC-derived activated PSCs can produce PDGF and TGF {beta}1. -- Abstract: Bone marrow cell (BMC)-derived myofibroblast-like cells have been reported in various organs, including the pancreas. However, the contribution of these cells to pancreatic fibrosis has not been fully discussed. The present study examined the possible involvement of pancreatic stellate cells (PSCs) originating from BMCs in the development of pancreatic fibrosis in a clinically relevant rat model of acute pancreatitis induced by a choline-deficient/ethionine-supplemented (CDE) diet. BMCs from female transgenic mice ubiquitously expressing green fluorescent protein (GFP) were transplanted into lethally irradiated male rats. Once chimerism was established, acute pancreatitis was induced by a CDE diet. Chronological changes in the number of PSCs originating from the donor BMCs were examined using double immunofluorescence for GFP and markers for PSCs, such as desmin and alpha smooth muscle actin ({alpha}SMA), 1, 3 and 8 weeks after the initiation of CDE feeding. We also used immunohistochemical staining to evaluate whether the PSCs from the BMCs produce growth factors, such as platelet-derived growth factor (PDGF) and transforming growth factor (TGF) {beta}1. The percentage of BMC-derived activated PSCs increased significantly, peaking after 1 week of CDE treatment (accounting for 23.3 {+-} 0.9% of the total population of activated PSCs) and then decreasing. These cells produced both PDGF and TGF{beta}1 during the early stage of pancreatic fibrosis. Our results suggest that PSCs originating from BMCs contribute mainly to the early stage of pancreatic injury, at least in part, by producing growth factors in a rat CDE diet-induced pancreatitis model.

  18. HuR mediates motility of human bone marrow-derived mesenchymal stem cells triggered by sphingosine 1-phosphate in liver fibrosis.

    PubMed

    Chang, Na; Ge, Jingjing; Xiu, Lei; Zhao, Zhongxin; Duan, Xianghui; Tian, Lei; Xie, Jieshi; Yang, Lin; Li, Liying

    2017-01-01

    Sphingosine 1-phosphate (S1P) participates in migration of bone marrow (BM)-derived mesenchymal stem cells (BMSCs) toward damaged liver via upregulation of S1P receptor 3 (S1PR3) during mouse liver fibrogenesis. But, the molecular mechanism is still unclear. HuR, as an RNA-binding protein, regulates tumor cell motility. Here, we examined the role of HuR in migration of human BMSCs (hBMSCs) in liver fibrosis. Results showed that HuR messenger RNA (mRNA) level was increased in human or mouse fibrotic livers, and correlated with S1PR3 mRNA expression. Using immunofluorescence, we found that HuR mainly localized in the nuclei of hepatocytes and non-parenchymal cells in normal livers. However, in fibrotic livers, we detected an increased HuR cytoplasmic localization in non-parenchymal cells. In chimeric mice of BM cell-labeled by EGFP, significant numbers of EGFP-positive cells (BM origin) were positive for HuR in fibrotic areas. Meanwhile, HuR-positive cells were also positive for α-SMA (myofibroblasts). In vitro, S1P induced hBMSCs migration via S1PR3 upregulation. HuR involved in S1P-induced hBMSCs migration and increased stabilization of S1PR3 mRNA via competing with miR-30e. RNA immunoprecipitation showed that HuR interacted with S1PR3 mRNA 3'UTR. Moreover, S1P resulted in phosphorylation and cytoplasmic translocation of HuR via S1PR3 and p38MAPK. Furthermore, we transplanted EGFP(+) BMSCs with or without HuR small interfering RNA (siRNA) into carbon tetrachloride-treated mice and found that knockdown of HuR inhibited the migration of BMSCs toward injured livers by flow cytometric analysis in vivo. We identified a positive feedback regulation mechanism between HuR and S1PR3 in S1P-induced BMSCs migration. HuR participates in upregulation of S1PR3 induced by S1P. S1P results in phosphorylation and translocation of HuR via S1PR3. Our results provide a new regulatory manner to the mechanism of liver fibrogenesis.

  19. Effect of apical hyperosmotic sodium challenge and amiloride on sodium transport in human bronchial epithelial cells from cystic fibrosis donors.

    PubMed

    Rasgado-Flores, Hector; Krishna Mandava, Vamsi; Siman, Homayoun; Van Driessche, Willy; Pilewski, Joseph M; Randell, Scott H; Bridges, Robert J

    2013-12-01

    Hypertonic saline (HS) inhalation therapy benefits cystic fibrosis (CF) patients [Donaldson SH, Bennet WD, Zeman KL, Knowles MR, Tarran R, Boucher RC. N Engl J Med 354: 241-250, 2006; Elkins MR, Robinson M, Rose BR, Harbour C, Moriarty CP, Marks GB, Belousova EG, Xuan W, Bye PT; the National Hypertonic Saline in Cystic Fibrosis (NHSCF) Study Group. N Engl J Med 354: 229-240, 2006]. Surprisingly, these benefits are long-lasting and are diminished by the epithelial Na(+) channel blocker amiloride (Donaldson SH, Bennet WD, Zeman KL, Knowles MR, Tarran R, Boucher RC. N Engl J Med 354: 241-250, 2006). Our aim was to explain these effects. Human bronchial epithelial (hBE) cells from CF lungs were grown in inserts and were used in three experimental approaches: 1) Ussing chambers to measure amiloride-sensitive short-circuit currents (INa); 2) continuous perfusion Ussing chambers; and 3) near "thin-film" conditions in which the airway surface of the inserts was exposed to a small volume (30 μl) of isosmotic or HS solution as the inserts were kept in their incubation tray and were subsequently used to measure INa under isosmotic conditions (near thin-film experiments; Tarran R, Boucher RC. Methods Mol Med 70: 479-492, 2002). HS solutions (660 mosmol/kgH2O) were prepared by adding additional NaCl to the isosmotic buffer. The transepithelial short-circuit current (ISC), conductance (GT), and capacitance (CT) were measured by transepithelial impedance analysis (Danahay H, Atherton HC, Jackson AD, Kreindler JL, Poll CT, Bridges RJ. Am J Physiol Lung Cell Mol Physiol 290: L558-L569, 2006; Singh AK, Singh S, Devor DC, Frizzell RA, van Driessche W, Bridges RJ. Methods Mol Med 70: 129-142, 2002). Exposure to apical HS inhibited INa, GT, and CT. The INa inhibition required 60 min of reexposure to the isosmotic solution to recover 75%. The time of exposure to HS required to inhibit INa was <2.5 min. Under near thin-film conditions, apical exposure to HS inhibited INa, but as

  20. Epigenetic regulation of cardiac fibrosis

    PubMed Central

    Stratton, Matthew S.; McKinsey, Timothy A.

    2016-01-01

    Fibrosis is defined as excess deposition of extracellular matrix (ECM), resulting in tissue scarring and organ dysfunction. In the heart, fibrosis may be reparative, replacing areas of myocyte loss with a structural scar following infarction, or reactive, which is triggered in the absence of cell death and involves interstitial ECM deposition in response to long-lasting stress. Interstitial fibrosis can increase the passive stiffness of the myocardium, resulting in impaired relaxation and diastolic dysfunction. Additionally, fibrosis can lead to disruption of electrical conduction in the heart, causing arrhythmias, and can limit myocyte oxygen availability and thus exacerbate myocardial ischemia. Here, we review recent studies that have illustrated key roles for epigenetic events in the control of pro-fibrotic gene expression, and highlight the potential of small molecules that target epigenetic regulators as a means of treating fibrotic cardiac diseases. PMID:26876451

  1. Role of CTGF gene promoter methylation in the development of hepatic fibrosis

    PubMed Central

    Shi, Cuicui; Li, Guangming; Tong, Yanyan; Deng, Yilin; Fan, Jiangao

    2016-01-01

    Connective tissue growth factor (CTGF) plays a critical role in the hepatic stellate cells (HSCs)-mediated development of hepatic fibrosis. Nevertheless, the effects of CTGF gene promoter methylation in the pathogenesis of hepatic fibrosis remain largely unknown. In the current study, we isolated and overexpressed CTGF in primary HSCs. We analyzed the CTGF gene promoter methylation inHSCs that undergo a phenotypic change into myofibroblast-like cellsthat express α-smooth muscle actin (α-SMA) in vitro and in vivo in a CCl4-induced rat hepatic fibrosis model. We found that CTGF promoted the phenotypic changes of HSCs into myofibroblasts in vitro, while inhibition of CTGF promoter methylation augmented the process, suggesting that CTGF gene promoter methylation may negatively regulate hepatic fibrosis. In vivo, CCl4 induced hepatic fibrosis in rats, and the severity of hepatic fibrosis inversely correlated with the levels of CTGF gene promoter methylation in HSCs. Together, our data demonstrate that CTGF gene promoter methylation may prevent the development of hepatic fibrosis, and low level of CTGF gene promoter methylation in HSCs may be a predisposing factor for developing liver fibrotic disease. PMID:27069546

  2. TGF-β1–Containing Exosomes from Injured Epithelial Cells Activate Fibroblasts to Initiate Tissue Regenerative Responses and Fibrosis

    PubMed Central

    Borges, Fernanda T.; Melo, Sonia A.; Özdemir, Berna C.; Kato, Noritoshi; Revuelta, Ignacio; Miller, Caroline A.; Gattone, Vincent H.; LeBleu, Valerie S.

    2013-01-01

    Hypoxia is associated with tissue injury and fibrosis but its functional role in fibroblast activation and tissue repair/regeneration is unknown. Using kidney injury as a model system, we demonstrate that injured epithelial cells produce an increased number of exosomes with defined genetic information to activate fibroblasts. Exosomes released by injured epithelial cells promote proliferation, α-smooth muscle actin expression, F-actin expression, and type I collagen production in fibroblasts. Fibroblast activation is dependent on exosomes delivering TGF-β1 mRNA among other yet to be identified moieties. This study suggests that TGF-β1 mRNA transported by exosomes constitutes a rapid response to initiate tissue repair/regenerative responses and activation of fibroblasts when resident parenchyma is injured. The results also inform potential utility of exosome-targeted therapies to control tissue fibrosis. PMID:23274427

  3. Liver fibrosis progression is related to CD4 cell depletion in patients coinfected with hepatitis C virus and human immunodeficiency virus.

    PubMed

    Puoti, M; Bonacini, M; Spinetti, A; Putzolu, V; Govindarajan, S; Zaltron, S; Favret, M; Callea, F; Gargiulo, F; Donato, F; Carosi, G

    2001-01-01

    A total of 204 patients with liver biopsy-proven hepatitis C virus (HCV) infection, 84 with and 120 without human immunodeficiency virus (HIV) coinfection, were studied, to evaluate variables possibly associated with the stage of liver fibrosis. All patients were injection drugs users, with a mean age of 32 years and an estimated duration of HCV infection of 12 years. Twenty-four patients (11%) had many fibrous septa with (5%) or without (6%) cirrhosis, 56 (27%) had few fibrous septa, and 124 (60%) had no fibrous septa. In all patients, an association was found between CD4 cell counts <500 cells/mm(3)and the presence of many fibrous septa (odds ratio, 3.2; P=.037), independent of HIV infection and other factors. These results suggest that HIV infection-induced CD4 depletion is independently associated with the severity of liver fibrosis in chronic HCV infection.

  4. Computer vision approach to morphometric feature analysis of basal cell nuclei for evaluating malignant potentiality of oral submucous fibrosis.

    PubMed

    Muthu Rama Krishnan, M; Pal, Mousumi; Paul, Ranjan Rashmi; Chakraborty, Chandan; Chatterjee, Jyotirmoy; Ray, Ajoy K

    2012-06-01

    This research work presents a quantitative approach for analysis of histomorphometric features of the basal cell nuclei in respect to their size, shape and intensity of staining, from surface epithelium of Oral Submucous Fibrosis showing dysplasia (OSFD) to that of the Normal Oral Mucosa (NOM). For all biological activity, the basal cells of the surface epithelium form the proliferative compartment and therefore their morphometric changes will spell the intricate biological behavior pertaining to normal cellular functions as well as in premalignant and malignant status. In view of this, the changes in shape, size and intensity of staining of the nuclei in the basal cell layer of the NOM and OSFD have been studied. Geometric, Zernike moments and Fourier descriptor (FD) based as well as intensity based features are extracted for histomorphometric pattern analysis of the nuclei. All these features are statistically analyzed along with 3D visualization in order to discriminate the groups. Results showed increase in the dimensions (area and perimeter), shape parameters and decreasing mean nuclei intensity of the nuclei in OSFD in respect to NOM. Further, the selected features are fed to the Bayesian classifier to discriminate normal and OSFD. The morphometric and intensity features provide a good sensitivity of 100%, specificity of 98.53% and positive predicative accuracy of 97.35%. This comparative quantitative characterization of basal cell nuclei will be of immense help for oral onco-pathologists, researchers and clinicians to assess the biological behavior of OSFD, specially relating to their premalignant and malignant potentiality. As a future direction more extensive study involving more number of disease subjects is observed.

  5. Using dendritic cells to evaluate how Burkholderia cenocepacia clonal isolates from a chronically infected cystic fibrosis patient subvert immune functions.

    PubMed

    Guadalupe Cabral, M; Pereira, Marília; Silva, Zélia; Iria, Inês; Coutinho, Carla; Lopes, Andreia; Sá-Correia, Isabel; Videira, Paula A

    2017-04-01

    Infection with Burkholderia cepacia complex (Bcc) bacteria is a threat to cystic fibrosis (CF) patients, commonly leading to a fatal pneumonia, the cepacia syndrome. It causes a massive production of pro-inflammatory cytokines and leucocyte recruitment to airway epithelium without resolving infection and contributing to tissue lesion. To dissect how Bcc bacteria subvert the immune response, we developed a co-culture model with human dendritic cells (DCs) and B. cenocepacia clonal variants isolated from a chronically infected CF patient, who died with cepacia syndrome. We demonstrated that the two late variants were sevenfold and 17-fold (respectively) more internalized by DCs than the variant that initiated infection. The late variants showed improved survival within DCs (60.29 and 52.82 CFU/DC) compared to the initial variant (0.38 CFU/DC). All clonal isolates induced high expression of inflammatory cytokines IL-8, IL-6, IL-1β, IL-12, IL-23, TNF-α and IL-1β. This pro-inflammatory trait was significantly more pronounced in DCs infected with the late variants than in DCs infected with the variant that initiated patient's infection. All infected DCs failed to upregulate maturation markers, HLA-DR, CD80, CD86 and CD83. Nevertheless, these infected DCs activated approximately twice more T cells than non-infected DCs. Similar T cell activation was observable with respective conditioned media, suggesting a non-antigen-specific activation. Our data indicate that during prolonged infection, B. cenocepacia acquires ability to survive intracellularly, inducing inflammation, while refraining DC's maturation and stimulating non-antigen-specific T cell responses. The co-culture model here developed may be broadly applied to study B. cenocepacia-induced immunomodulation.

  6. Activation of TGF-β1-CD147 positive feedback loop in hepatic stellate cells promotes liver fibrosis.

    PubMed

    Li, Hai-Yan; Ju, Di; Zhang, Da-Wei; Li, Hao; Kong, Ling-Min; Guo, Yanhai; Li, Can; Wang, Xi-Long; Chen, Zhi-Nan; Bian, Huijie

    2015-11-12

    Activation of hepatic stellate cells (HSCs) by transforming growth factor-β1 (TGF-β1) initiates HBV-associated fibrogenesis. The mechanism of TGF-β1 modulating HSC activation is not fully uncovered. We hypothesized a positive feedback signaling loop of TGF-β1-CD147 promoting liver fibrogenesis by activation of HSCs. Human HSC cell line LX-2 and spontaneous liver fibrosis model derived from HBV transgenic mice were used to evaluate the activation of molecules in the signaling loop. Wound healing and cell contraction assay were performed to detect the CD147-overexpressed HSC migration and contraction. The transcriptional regulation of CD147 by TGF-β1/Smad4 was determined using dual-luciferase reporter assay and chromatin immunoprecipitation. We found that a positive reciprocal regulation between TGF-β1 and CD147 mediated HSC activation. CD147 over-expression promoted HSC migration and accelerated TGF-β1-induced cell contraction. Phosphorylation of Smad2 and Smad3 in cooperation with Smad4 mediated the TGF-β1-regulated CD147 expression. Smad4 activated the transcription by direct interaction with CD147 promoter. Meanwhile, CD147 modulated the activated phenotype of HSCs through the ERK1/2 and Sp1 which up-regulated α-SMA, collagen I, and TGF-β1 synthesis. These findings indicate that TGF-β1-CD147 loop plays a key role in regulating the HSC activation and combination of TGF-β receptor inhibitor and anti-CD147 antibody might be promised to reverse fibrogenesis.

  7. A Novel Matrine Derivative WM130 Inhibits Activation of Hepatic Stellate Cells and Attenuates Dimethylnitrosamine-Induced Liver Fibrosis in Rats.

    PubMed

    Xu, Yang; Peng, Zhangxiao; Ji, Weidan; Li, Xiang; Lin, Xuejing; Qian, Liqiang; Li, Xiaoya; Chai, Xiaoyun; Wu, Qiuye; Gao, Quangen; Su, Changqing

    2015-01-01

    Activation of hepatic stellate cells (HSCs) is a critical event in process of hepatic fibrogenesis and cirrhosis. Matrine, the active ingredient of Sophora, had been used for clinical treatment of acute/chronic liver disease. However, its potency was low. We prepared a high potency and low toxicity matrine derivate, WM130 (C30N4H40SO5F), which exhibited better pharmacological activities on antihepatic fibrosis. This study demonstrated that WM130 results in a decreased proliferative activity of HSC-T6 cells, with the half inhibitory concentration (IC50) of 68 μM. WM130 can inhibit the migration and induce apoptosis in HSC-T6 cells at both concentrations of 68 μM (IC50) and 34 μM (half IC50). The expression of α-SMA, Collagen I, Collagen III, and TGF-β1 could be downregulated, and the protein phosphorylation levels of EGFR, AKT, ERK, Smad, and Raf (p-EGFR, p-AKT, p-ERK, p-Smad, and p-Raf) were also decreased by WM130. On the DMN-induced rat liver fibrosis model, WM130 can effectively reduce the TGF-β1, AKT, α-SMA, and p-ERK levels, decrease the extracellular matrix (ECM) formation, and inhibit rat liver fibrosis progression. In conclusion, this study demonstrated that WM130 can significantly inhibit the activation of HSC-T6 cells and block the rat liver fibrosis progression by inducing apoptosis, suppressing the deposition of ECM, and inhibiting TGF-β/Smad and Ras/ERK pathways.

  8. The antioxidant effect of β-caryophyllene protects rat liver from carbon tetrachloride-induced fibrosis by inhibiting hepatic stellate cell activation.

    PubMed

    Calleja, Miguel Angel; Vieites, Jose María; Montero-Meléndez, Trinidad; Montero-Meterdez, Trinidad; Torres, María Isabel; Faus, María José; Gil, Angel; Suárez, Antonio

    2013-02-14

    Plant-based whole foods provide thousands of bioactive metabolites to the human diet that reduce the risk of developing chronic diseases. β-Caryophyllene (CAR) is a common constituent of the essential oil of numerous plants, vegetables, fruits and medicinal herbs, and has been used as a flavouring agent since the 1930 s. Here, we report the antioxidant activity of CAR, its protective effect on liver fibrosis and its inhibitory capacity on hepatic stellate cell (HSC) activation. CAR was tested for the inhibition of lipid peroxidation and as a free radical scavenger. CAR had higher inhibitory capacity on lipid peroxidation than probucol, α-humulene and α-tocopherol. Also, CAR showed high scavenging activities against hydroxyl radical and superoxide anion. The activity of 5-lipoxygenase, an enzyme that actively participates in fibrogenesis, was significantly inhibited by CAR. Carbon tetrachloride-treated rats received CAR at 2, 20 and 200 mg/kg. CAR significantly improved liver structure, and reduced fibrosis and the expression of Col1a1, Tgfb1 and Timp1 genes. Oxidative stress was used to establish a model of HSC activation with overproduction of extracellular matrix proteins. CAR (1 and 10 μm) increased cell viability and significantly reduced the expression of fibrotic marker genes. CAR, a sesquiterpene present in numerous plants and foods, is as a natural antioxidant that reduces carbon tetrachloride-mediated liver fibrosis and inhibits hepatic cell activation.

  9. Spatial organization and correlations of cell nuclei in brain tumors.

    PubMed

    Jiao, Yang; Berman, Hal; Kiehl, Tim-Rasmus; Torquato, Salvatore

    2011-01-01

    Accepting the hypothesis that cancers are self-organizing, opportunistic systems, it is crucial to understand the collective behavior of cancer cells in their tumorous heterogeneous environment. In the present paper, we ask the following basic question: Is this self-organization of tumor evolution reflected in the manner in which malignant cells are spatially distributed in their heterogeneous environment? We employ a variety of nontrivial statistical microstructural descriptors that arise in the theory of heterogeneous media to characterize the spatial distributions of the nuclei of both benign brain white matter cells and brain glioma cells as obtained from histological images. These descriptors, which include the pair correlation function, structure factor and various nearest neighbor functions, quantify how pairs of cell nuclei are correlated in space in various ways. We map the centroids of the cell nuclei into point distributions to show that while commonly used local spatial statistics (e.g., cell areas and number of neighboring cells) cannot clearly distinguish spatial correlations in distributions of normal and abnormal cell nuclei, their salient structural features are captured very well by the aforementioned microstructural descriptors. We show that the tumorous cells pack more densely than normal cells and exhibit stronger effective repulsions between any pair of cells. Moreover, we demonstrate that brain gliomas are organized in a collective way rather than randomly on intermediate and large length scales. The existence of nontrivial spatial correlations between the abnormal cells strongly supports the view that cancer is not an unorganized collection of malignant cells but rather a complex emergent integrated system.

  10. Arsenic promotes ubiquitinylation and lysosomal degradation of cystic fibrosis transmembrane conductance regulator (CFTR) chloride channels in human airway epithelial cells.

    PubMed

    Bomberger, Jennifer M; Coutermarsh, Bonita A; Barnaby, Roxanna L; Stanton, Bruce A

    2012-05-18

    Arsenic exposure significantly increases respiratory bacterial infections and reduces the ability of the innate immune system to eliminate bacterial infections. Recently, we observed in the gill of killifish, an environmental model organism, that arsenic exposure induced the ubiquitinylation and degradation of cystic fibrosis transmembrane conductance regulator (CFTR), a chloride channel that is essential for the mucociliary clearance of respiratory pathogens in humans. Accordingly, in this study, we tested the hypothesis that low dose arsenic exposure reduces the abundance and function of CFTR in human airway epithelial cells. Arsenic induced a time- and dose-dependent increase in multiubiquitinylated CFTR, which led to its lysosomal degradation, and a decrease in CFTR-mediated chloride secretion. Although arsenic had no effect on the abundance or activity of USP10, a deubiquitinylating enzyme, siRNA-mediated knockdown of c-Cbl, an E3 ubiquitin ligase, abolished the arsenic-stimulated degradation of CFTR. Arsenic enhanced the degradation of CFTR by increasing phosphorylated c-Cbl, which increased its interaction with CFTR, and subsequent ubiquitinylation of CFTR. Because epidemiological studies have shown that arsenic increases the incidence of respiratory infections, this study suggests that one potential mechanism of this effect involves arsenic-induced ubiquitinylation and degradation of CFTR, which decreases chloride secretion and airway surface liquid volume, effects that would be proposed to reduce mucociliary clearance of respiratory pathogens.

  11. Modulation of Stem Cells Differentiation and Myostatin as an Approach to Counteract Fibrosis in Muscle Dystrophy and Regeneration after Injury

    DTIC Science & Technology

    2008-03-01

    Renal fibrosis and glomerulosclerosis in a new mouse model of diabetic nephropathy and its regression by bone morphogenic protein-7 and advanced...whether long-term treatment with pioglitazone reduces vascular and renal fibrosis in an animal model of type 2 diabetes 8. Pilot grant (Bathia/Ho...1), will be subjected to anti-myostatin treatments and tested for differentiation as in Task 1: a) anti-myostatin antibody; b) follistatin; c

  12. The Therapeutic Potential of Human Umbilical Mesenchymal Stem Cells From Wharton’s Jelly in the Treatment of Rat Peritoneal Dialysis-Induced Fibrosis

    PubMed Central

    Fan, Yu-Pei; Hsia, Ching-Chih; Tseng, Kuang-Wen; Liao, Chih-Kai; Fu, Tz-Win; Ko, Tsui-Ling; Chiu, Mei-Miao; Shih, Yang-Hsin; Huang, Pei-Yu; Chiang, Yi-Chia

    2016-01-01

    A major complication in continuous, ambulatory peritoneal dialysis in patients with end-stage renal disease who are undergoing long-term peritoneal dialysis (PD) is peritoneal fibrosis, which can result in peritoneal structural changes and functional ultrafiltration failure. Human umbilical mesenchymal stem cells (HUMSCs) in Wharton’s jelly possess stem cell properties and are easily obtained and processed. This study focuses on the effects of HUMSCs on peritoneal fibrosis in in vitro and in vivo experiments. After 24-hour treatment with mixture of Dulbecco’s modified Eagle’s medium and PD solution at a 1:3 ratio, primary human peritoneal mesothelial cells became susceptible to PD-induced cell death. Such cytotoxic effects were prevented by coculturing with primary HUMSCs. In a rat model, intraperitoneal injections of 20 mM methylglyoxal (MGO) in PD solution for 3 weeks (the PD/MGO 3W group) markedly induced abdominal cocoon formation, peritoneal thickening, and collagen accumulation. Immunohistochemical analyses indicated neoangiogenesis and significant increase in the numbers of ED-1- and α-smooth muscle actin (α-SMA)-positive cells in the thickened peritoneum in the PD/MGO 3W group, suggesting that PD/MGO induced an inflammatory response. Furthermore, PD/MGO treatment for 3 weeks caused functional impairments in the peritoneal membrane. However, in comparison with the PD/MGO group, intraperitoneal administration of HUMSCs into the rats significantly ameliorated the PD/MGO-induced abdominal cocoon formation, peritoneal fibrosis, inflammation, neoangiogenesis, and ultrafiltration failure. After 3 weeks of transplantation, surviving HUMSCs were found in the peritoneum in the HUMSC-grafted rats. Thus, xenografts of HUMSCs might provide a potential therapeutic strategy in the prevention of peritoneal fibrosis. Significance This study demonstrated that direct intraperitoneal transplantation of human umbilical mesenchymal stem cells into the rat effectively

  13. Severe deficiency of cystic fibrosis transmembrane conductance regulator messenger RNA carrying nonsense mutations R553X and W1316X in respiratory epithelial cells of patients with cystic fibrosis.

    PubMed Central

    Hamosh, A; Trapnell, B C; Zeitlin, P L; Montrose-Rafizadeh, C; Rosenstein, B J; Crystal, R G; Cutting, G R

    1991-01-01

    Cystic fibrosis (CF) is the most common, lethal inherited disorder in the Caucasian population. We have recently reported two African-American patients with nonsense mutations in each CF gene and severe pancreatic disease, but mild pulmonary disease. In order to examine the effect of these nonsense mutations on CF gene expression, bronchial and nasal epithelial cells were obtained from one of these patients (no. 246), a compound heterozygote for nonsense mutations R553X and W1316X; a healthy normal individual; a patient (no. 528) homozygous for the common CF mutation (delta F508); and a CF patient (no. 272) who carries the R553X mutation and a missense mutation, S549N. When mRNA from bronchial cells of the normal individual, the delta F508 homozygote, and the S549N/R553X compound heterozygote was reverse transcribed and amplified by polymerase chain reaction using primers derived from the CF gene, DNA fragments of the predicted size were observed. However, patient no. 246 with nonsense mutations in each CF gene has no detectable cystic fibrosis transmembrane conductance regulator (CFTR) messenger RNA, and therefore should have severely diminished, and possibly absent, CFTR protein. Furthermore, less than 2% of the CFTR transcripts in nasal epithelial cells from patient no. 272 (S549N/R553X) were derived from the gene with the nonsense mutation. We conclude that severe reduction in CFTR mRNA causes CF, but can have different consequences in the lung and pancreas. Images PMID:1721624

  14. Connective tissue cells expressing fibro/adipogenic progenitor markers increase under chronic damage: relevance in fibroblast-myofibroblast differentiation and skeletal muscle fibrosis.

    PubMed

    Contreras, Osvaldo; Rebolledo, Daniela L; Oyarzún, Juan Esteban; Olguín, Hugo C; Brandan, Enrique

    2016-06-01

    Fibrosis occurs in skeletal muscle under various pathophysiological conditions such as Duchenne muscular dystrophy (DMD), a devastating disease characterized by fiber degeneration that results in progressive loss of muscle mass, weakness and increased extracellular matrix (ECM) accumulation. Fibrosis is also observed after skeletal muscle denervation and repeated cycles of damage followed by regeneration. The ECM is synthesized largely by fibroblasts in the muscle connective tissue under normal conditions. Myofibroblasts, cells that express α-smooth muscle actin (α-SMA), play a role in many tissues affected by fibrosis. In skeletal muscle, fibro/adipogenic progenitors (FAPs) that express cell-surface platelet-derived growth factor receptor-α (PDGFR-α) and the transcription factor Tcf4 seem to be responsible for connective tissue synthesis and are good candidates for the origin of myofibroblasts. We show that cells positive for Tcf4 and PDGFR-α are expressed in skeletal muscle under normal conditions and are increased in various skeletal muscles of mdx mice, a murine model for DMD, wild type muscle after sciatic denervation and muscle subjected to chronic damage. These cells co-label with the myofibroblast marker α-SMA in dystrophic muscle but not in normal tissue. The Tcf4-positive cells lie near macrophages mainly concentrated in dystrophic necrotic-regenerating foci. The close proximity of Tcf4-positive cells to inflammatory cells and their previously described role in muscle regeneration might reflect an active interaction between these cell types and growth factors, possibly resulting in a muscular regenerative or fibrotic condition.

  15. Fibrocytes are associated with the fibrosis of coronary heart disease.

    PubMed

    Lei, Pu-Ping; Qu, Yong-Qiang; Shuai, Qun; Tao, Si-Ming; Bao, Yu-Xia; Wang, Yu; Wang, Shang-Wen; Wang, Dian-Hua

    2013-01-15

    Fibrocytes contribute significantly to fibrosis in many cardiac diseases. However, it is not clear whether fibrocytes are associated with the fibrosis in coronary heart disease (CHD). The aim of this study was to determine whether fibrocytes are involved in cardiac fibrosis in CHD. We identified the presence of fibrocytes in CHD heart by immunofluorescence and confocal microscopy, examined the collagen volume fraction by Masson's Trichrome staining, and evaluated the correlation between fibrocytes and cardiac fibrosis. In conjunction, we examined the location of CXCL12, a homing factor and specific ligand for CXCR4, by immunohistochemistry. Fibrocytes were identified in 26 out of 27 CHD hearts and in 10 out of 11 normal hearts. Combinations, including CD34/αSMA, CD34/procollagen-I, CD45/αSMA, CXCR4/procollagen-I and CXCR4/αSMA, stained significantly more fibrocytes in CHD hearts as compared with those in normal hearts (p<0.05). There were positive correlations between the collagen volume fraction and the amount of fibrocytes (r=0.558; p=0.003<0.01) and between the number of CXCR4(+) fibrocytes and the CXCL12(+) cells (r=0.741; p=0.000<0.01) in CHD hearts. Based upon these findings, we conclude that fibrocytes, likely recruited through the CXCR4/CXCL12 axis, may contribute to the increase in the fibroblast population in CHD heart.

  16. Interferon gamma-secreting HCV-specific CD8+ T cells in the liver of patients with chronic C hepatitis: relation to liver fibrosis--ANRS HC EP07 study.

    PubMed

    Bonilla, N; Barget, N; Andrieu, M; Roulot, D; Letoumelin, P; Grando, V; Trinchet, J C; Ganne-Carrié, N; Beaugrand, M; Deny, P; Choppin, J; Guillet, J; Ziol, M

    2006-07-01

    Little is known about the role of specific hepatitis C virus (HCV) CD8+ T cells in liver damage, especially for the progression of fibrosis, during the highly variable course of chronic C hepatitis. The aim of this study was to investigate the presence of HCV-specific CD8+ T cells in the liver of patients with chronic C hepatitis and to examine their clinical significance by relating the response to liver fibrosis and progression rate, serum viral load, serum aminotransferase levels, inflammatory activity and in situ characteristics of the intrahepatic infiltrate. Fifteen patients were prospectively included in the study. Intrahepatic lymphocytes were tested for interferon gamma (IFNg) production in response to HCV class I-restricted epitopic peptides using enzyme-linked immunospot analysis. Liver biopsy samples were evaluated for fibrosis, fibrosis progression rate, activity, and in situ number of CD8+ cytotoxic lymphocytes and apoptotic cells. An IFNg-specific CD8+ T-cell response was detected in the liver samples of 47% of patients which was significantly related to a lower stage of fibrosis (P = 0.02) and a lower progression rate of fibrosis (P = 0.01). It was neither related to the number of cytotoxic lymphocytes infiltrating the liver nor to hepatocyte apoptosis. In conclusion, our results indicate that the presence of HCV-specific IFNg-secreting T cells in the liver of patients with chronic C hepatitis is associated with low liver fibrosis and fibrosis progression rate, suggesting that these IFNg-secreting T cells might limit the progression of liver damage.

  17. Genomic analysis of lung cell lines exposures to space radiation and the effect of lunar dust on selected fibrosis gene using RT2 PCR Array

    NASA Astrophysics Data System (ADS)

    Yeshitla, Samrawit

    In the United States (U.S.), lung cancer is the number one cause of cancer death among men and women. Previous studies on human and animal epithelial lung cells showed that ionizing radiation and certain environmental pollutants are carcinogens. The surface area of the lungs and the slow turnover rate of the epithelial cells are suggested to play a role in the vulnerability of the cells, which lead to increase in the progenitor cell of the lung. It has been proposed that these progenitor cells, when exposed to radiation undergo multiple alterations that cause the cells to become cancerous. The current thought is that the lungs contain several facultative progenitor cells that are situated throughout the lung epithelium and are regionally restricted in their regenerative capacity. In this study, normal Human Bronchial Epithelial Cells (HBECs) were immortalized through the expression of Cdk4 and hTERT and evaluated for the effects radiation using in vitro study. The HBECs retained its novel multipotent capacity in vitro and represented unrestricted progenitor cells of the adult lungs, which resemble an embryonic progenitor. Analysis of the transformed clones of human bronchial epithelial cell line, HEBC3KT exposed to Fe ions and gamma rays revealed chromosomal abnormality, which was detected with the Multi-color Fluorescent In Situ Hybridization (mFish). In Part two of this study the F344 rats exposed to lunar dust, for 4 weeks (6h/d; 5d/wk.) in nose-only inhalation chambers at concentrations of 0 (control air), 2.1, 6.8, 20.8, and 61 mg/m3 of lunar dust, were used to determine the lunar dust toxicity on the lung tissues and total RNA were prepared from the tissues and used for gene expression. Analysis of gene expression data using Ingenuity Pathway Analysis tool identified multiple pathways of which fibrosis was one of the pathways. The Rat Fibrosis RT 2 Profile PCR Array was used to profile the expression of 84 genes that are relevant to fibrosis in the lung

  18. Hydrodynamics-based transfection of rat interleukin-10 gene attenuates porcine serum-induced liver fibrosis in rats by inhibiting the activation of hepatic stellate cells.

    PubMed

    Huang, Yue-Hong; Chen, Yun-Xin; Zhang, Li-Juan; Chen, Zhi-Xin; Wang, Xiao-Zhong

    2014-09-01

    Liver fibrosis is the common pathological outcome for the majority of chronic liver diseases. Interleukin-10 (IL-10) is a cytokine that downregulates proinflammatory responses and has a modulatory effect on liver fibrogenesis. However, little is known regarding the effect of rat interleukin‑10 (rIL‑10) gene by hydrodynamics-based transfection (HBT) on liver fibrosis in rats. The aim of this study was to investigate the effect of the rIL-10 gene by HBT on the progression of liver fibrosis induced by porcine serum (PS) in rats and explore its possible mechanism. Plasmid‑expressing rIL-10 was transferred into rats by HBT and immunohistochemistry and RT-PCR were used to detect the major organ expressing rIL-10. Liver fibrosis was induced in rats by intraperitoneal administration of PS for 8 weeks. Plasmid pcDNA3-rIL-10 solution was administered weekly by HBT starting at the 5th week. Liver function and hepatic histology were examined. The possible molecular mechanisms of rIL-10 gene therapy were assessed in liver tissue and hepatic stellate cells (HSCs) co-cultured with BRL cells (a hepatocyte line) in vitro. The results showed rIL-10 expression occurred mainly in the liver following rIL-10 gene transfer by HBT. Maintaining a stable expression of rIL-10 in serum was assessed by repeated administration. The rIL-10 gene treatment attenuated liver inflammation and fibrosis in PS-induced fibrotic rats, reduced the deposition of collagen and the expression of α-smooth muscle actin (α-SMA) in fibrotic rats. The in vitro experiment showed that the expression of a-SMA and procollagen type I in HSCs co-cultured with the BRL‑transfected rIL-10 gene were significantly decreased. These findings indicate that rIL-10 gene therapy by HBT attenuates PS-induced liver fibrosis in rats and that its mechanism is associated with rIL-10 inhibiting the activation of HSCs and promoting the degeneration of collagen.

  19. Human Immunodeficiency Virus (HIV)-1 Infects Human Hepatic Stellate Cells and Promotes Collagen I and Monocyte Chemoattractant Protein-1 Expression: Implications for the Pathogenesis of HIV/Hepatitis C Virus–Induced Liver Fibrosis

    PubMed Central

    Tuyama, Ana C.; Hong, Feng; Saiman, Yedidya; Wang, Chuansheng; Ozkok, Derya; Mosoian, Arevik; Chen, Ping; Chen, Benjamin K.; Klotman, Mary E.; Bansal, Meena B.

    2010-01-01

    Patients coinfected with human immunodeficiency virus (HIV) and hepatitis C virus (HCV) develop more rapid fibrosis than those infected with HCV only. In HIV/HCV-coinfected patients, fibrosis progression correlates with HIV RNA levels, suggesting a direct role of HIV in liver fibrogenesis. Chemokine (C-C motif) receptor 5 (CCR5) and cysteine-X-cysteine receptor 4 (CXCR4), the two major coreceptors required for HIV entry into cells, are expressed on activated hepatic stellate cells (HSCs), the principle fibrogenic cell type in the liver. We therefore examined whether HIV can infect HSCs, explored the potential mechanisms of viral entry, and assessed the impact of infection as reflected by the ability of HSCs to transfer virus to T lymphocytes and elicit a proinflammatory and profibrogenic response. We report that the laboratory-adapted viruses HIV-IIIB (CXCR4-tropic or X4) and HIV-BaL (CCR5-tropic or R5) and primary HIV isolates can infect both a human stellate cell line, LX-2, and primary human HSCs. HIV entry and gene expression in HSCs was confirmed using HIV–green fluorescent protein (GFP) expression viral constructs in the presence or absence of the reverse-transcriptase inhibitor azidothymidine. CD4 expression on a subset of primary HSCs was demonstrated using fluorescence-activated cell sorting and immunofluorescence staining. Blocking experiments in the presence of anti-CD4, anti-CXCR4, and anti-CCR5 revealed that HIV entry into HSCs is predominantly CD4/chemokine coreceptor-independent. HIV infection promoted HSC collagen I expression and secretion of the proinflammatory cytokine monocyte chemoattractant protein-1. Furthermore, infected LX-2 cells were capable of transferring GFP-expressing virus to T lymphocytes in a coculture system. Conclusion Taken together, our results suggest a potential role of HIV in liver fibrosis/inflammation mediated through effects on HSCs. The role of early highly active antiretroviral therapy initiation in patients with HIV

  20. Human Muse cells, non-tumorigenic pluripotent-like stem cells, have the capacity for liver regeneration by specific homing and replenishment of new hepatocytes in liver fibrosis mouse model.

    PubMed

    Iseki, Masahiro; Kushida, Yoshihiro; Wakao, Shohei; Akimoto, Takahiro; Mizuma, Masamichi; Motoi, Fuyuhiko; Asada, Ryuta; Shimizu, Shinobu; Unno, Michiaki; Chazenbalk, Gregorio; Dezawa, Mari

    2016-11-02

    Muse cells, a novel type of non-tumorigenic pluripotent-like stem cells reside in the bone marrow, skin and adipose tissue, are collectable as cells positive for pluripotent surface marker SSEA-3. They are able to differentiate into cells representative of all three germ layers. The capacity of intravenously injected human bone marrow-Muse cells to repair the liver fibrosis model of immunodeficient mice was evaluated in this study. They exhibited the ability for differentiation spontaneously into hepatoblast/hepatocyte-lineage cells and high migration toward the serum and liver tissue of carbon tetrachloride-treated mice in vitro. In vivo, they specifically accumulated into the liver, but not into other organs except the lower rate in the lung at 2 weeks after intravenous injection into the liver fibrosis model. After homing, Muse cells spontaneously differentiated in vivo into HepPar-1 (71.1±15.2%), human albumin (54.3±8.2%) and anti-trypsin (47.9±4.6%)-positive cells without fusing with host hepatocytes, and expressed mature functional markers such as human-CYP1A2, and human-Glc-6-Pase, at 8 weeks. Recovery in serum total bilirubin and albumin, and significant attenuation of fibrosis were recognized with statistical differences between the Muse group and control groups which received the vehicle or the same number of non-Muse cells, namely cells other than Muse cells in bone marrow mesenchymal stem cells. Thus, unlike ES and iPS cells, Muse cells are unique in their efficient migration and integration into damaged liver only by intravenous injection, nontumorigenicity, and spontaneous differentiation into hepatocytes, rendering induction into hepatocytes prior to transplantation unnecessary. They are suggested to repair liver fibrosis in two simple steps; expansion after collection from the bone marrow and intravenous injection. Such feasible strategy might provide impressive regenerative performance to liver disease patients.

  1. The advancements of heparanase in fibrosis

    PubMed Central

    Lv, Qianying; Zeng, Ji; He, Long

    2016-01-01

    Fibrosis is the endpoint in many chronic inflammatory diseases and is defined as an abnormal accumulation of extracellular matrix components. Fibrosis can affect almost any tissue, especially heart, lung, liver, and kidney, and numerous studies have been conducted to find satisfactory treatments. Since heparanase is a kind of endo-β-D-glucuronidase that is capable of cleaving heparan sulfate side chains of heparan sulfate proteoglycans on cell surfaces and the extracellular matrix, which further regulate the bioavailability of growth factors (FGF-2, TGF-β). Meanwhile, FGF-2 and TGF-β play a major role in the fibrosis process. Recent studies including ours have consistently demonstrated that heparanase could promote fibrosis process in different organs. Thus in this mini-review, we updated the advancement of heparanase in the regulation of fibrosis generation, and discussed its impact on several critical signaling pathways relevant to fibrosis. PMID:28078057

  2. Biomarkers for liver fibrosis

    DOEpatents

    Jacobs, Jon M.; Burnum-Johnson, Kristin E.; Baker, Erin M.; Smith, Richard D.; Gritsenko, Marina A.; Orton, Daniel

    2015-09-15

    Methods and systems for diagnosing or prognosing liver fibrosis in a subject are provided. In some examples, such methods and systems can include detecting liver fibrosis-related molecules in a sample obtained from the subject, comparing expression of the molecules in the sample to controls representing expression values expected in a subject who does not have liver fibrosis or who has non-progressing fibrosis, and diagnosing or prognosing liver fibrosis in the subject when differential expression of the molecules between the sample and the controls is detected. Kits for the diagnosis or prognosis of liver fibrosis in a subject are also provided which include reagents for detecting liver fibrosis related molecules.

  3. Circulating fibrocytes traffic to the lungs in response to CXCL12 and mediate fibrosis.

    PubMed

    Phillips, Roderick J; Burdick, Marie D; Hong, Kurt; Lutz, Marin A; Murray, Lynne A; Xue, Ying Ying; Belperio, John A; Keane, Michael P; Strieter, Robert M

    2004-08-01

    Previous reports have identified a circulating pool of CD45(+) collagen I(+) CXCR4(+) (CD45(+)Col I(+)CXCR4(+)) cells, termed fibrocytes, that traffic to areas of fibrosis. No studies have demonstrated that these cells actually contribute to fibrosis, however. Pulmonary fibrosis was originally thought to be mediated solely by resident lung fibroblasts. Here we show that a population of human CD45(+)Col I(+)CXCR4(+) circulating fibrocytes migrates in response to CXCL12 and traffics to the lungs in a murine model of bleomycin-induced pulmonary fibrosis. Next, we demonstrated that murine CD45(+)Col I(+)CXCR4(+) fibrocytes also traffic to the lungs in response to a bleomycin challenge. Maximal intrapulmonary recruitment of CD45(+)Col I(+)CXCR4(+) fibrocytes directly correlated with increased collagen deposition in the lungs. Treatment of bleomycin-exposed animals with specific neutralizing anti-CXCL12 Ab's inhibited intrapulmonary recruitment of CD45(+)Col I(+)CXCR4(+) circulating fibrocytes and attenuated lung fibrosis. Thus, our results demonstrate, we believe for the first time, that circulating fibrocytes contribute to the pathogenesis of pulmonary fibrosis.

  4. Tubular Dickkopf-3 promotes the development of renal atrophy and fibrosis.

    PubMed

    Federico, Giuseppina; Meister, Michael; Mathow, Daniel; Heine, Gunnar H; Moldenhauer, Gerhard; Popovic, Zoran V; Nordström, Viola; Kopp-Schneider, Annette; Hielscher, Thomas; Nelson, Peter J; Schaefer, Franz; Porubsky, Stefan; Fliser, Danilo; Arnold, Bernd; Gröne, Hermann-Josef

    2016-01-21

    Renal tubular atrophy and interstitial fibrosis are common hallmarks of etiologically different progressive chronic kidney diseases (CKD) that eventually result in organ failure. Even though these pathological manifestations constitute a major public health problem, diagnostic tests, as well as therapeutic options, are currently limited. Members of the dickkopf (DKK) family, DKK1 and -2, have been associated with inhibition of Wnt signaling and organ fibrosis. Here, we identify DKK3 as a stress-induced, tubular epithelia-derived, secreted glycoprotein that mediates kidney fibrosis. Genetic as well as antibody-mediated abrogation of DKK3 led to reduced tubular atrophy and decreased interstitial matrix accumulation in two mouse models of renal fibrosis. This was facilitated by an amplified, antifibrogenic, inflammatory T cell response and diminished canonical Wnt/β-catenin signaling in stressed tubular epithelial cells. Moreover, in humans, urinary DKK3 levels specifically correlated with the extent of tubular atrophy and interstitial fibrosis in different glomerular and tubulointerstitial diseases. In summary, our data suggest that DKK3 constitutes an immunosuppressive and a profibrotic epithelial protein that might serve as a potential therapeutic target and diagnostic marker in renal fibrosis.

  5. Cystic Fibrosis

    PubMed Central

    Asay, Lyal D.

    1965-01-01

    Cystic fibrosis, a disease thought to be transmitted as a recessive genetic trait, is found as a disease in about one in 1,000 to one in 10,000 births. It involves all of the exocrine glands with presenting symptoms dependent upon the extent of involvement of any group of glands. Many aspects of the disease can be corrected by substitution therapy. This applies particularly to the use of animal pancreas for the steatorrhea and salt for prevention of heat prostration. Unfortunately, the obstructive pulmonary disease with secondary bronchial infections can only be treated symptomatically by the use of mucus thinning agents, postural drainage, and antibiotics. Nevertheless, longevity can be increased and a great deal of hope offered to the families of these unfortunate children by careful supervision of their medical care. ImagesFigure 1.Figure 2.Figure 3.Figure 4.Figure 5.Figure 6.Figure 7.Figure 8.Figure 9.Figure 10.Figure 11. PMID:14288148

  6. Chemotaxis and Binding of Pseudomonas aeruginosa to Scratch-Wounded Human Cystic Fibrosis Airway Epithelial Cells

    PubMed Central

    Schwarzer, Christian; Fischer, Horst; Machen, Terry E.

    2016-01-01

    Confocal imaging was used to characterize interactions of Pseudomonas aeruginosa (PA, expressing GFP or labeled with Syto 11) with CF airway epithelial cells (CFBE41o-, grown as confluent monolayers with unknown polarity on coverglasses) in control conditions and following scratch wounding. Epithelia and PAO1-GFP or PAK-GFP (2 MOI) were incubated with Ringer containing typical extracellular salts, pH and glucose and propidium iodide (PI, to identify dead cells). PAO1 and PAK swam randomly over and did not bind to nonwounded CFBE41o- cells. PA migrated rapidly (began within 20 sec, maximum by 5 mins) and massively (10–80 fold increase, termed “swarming”), but transiently (random swimming after 15 mins), to wounds, particularly near cells that took up PI. Some PA remained immobilized on cells near the wound. PA swam randomly over intact CFBE41o- monolayers and wounded monolayers that had been incubated with medium for 1 hr. Expression of CFTR and altered pH of the media did not affect PA interactions with CFBE41o- wounds. In contrast, PAO1 swarming and immobilization along wounds was abolished in PAO1 (PAO1ΔcheYZABW, no expression of chemotaxis regulatory components cheY, cheZ, cheA, cheB and cheW) and greatly reduced in PAO1 that did not express amino acid receptors pctA, B and C (PAO1ΔpctABC) and in PAO1 incubated in Ringer containing a high concentration of mixed amino acids. Non-piliated PAKΔpilA swarmed normally towards wounded areas but bound infrequently to CFBE41o- cells. In contrast, both swarming and binding of PA to CFBE41o- cells near wounds were prevented in non-flagellated PAKΔfliC. Data are consistent with the idea that (i) PA use amino acid sensor-driven chemotaxis and flagella-driven swimming to swarm to CF airway epithelial cells near wounds and (ii) PA use pili to bind to epithelial cells near wounds. PMID:27031335

  7. Somatostatin analogue (octreotide) inhibits bile duct epithelial cell proliferation and fibrosis after extrahepatic biliary obstruction.

    PubMed Central

    Tracy, T. F.; Tector, A. J.; Goerke, M. E.; Kitchen, S.; Lagunoff, D.

    1993-01-01

    Extrahepatic biliary obstruction leads to bile duct epithelial cell proliferation. Somatostatin and its analogue, octreotide, have been shown to inhibit DNA synthesis and proliferation in hepatocytes. We investigated the effect of octreotide on the biliary epithelial cell proliferative responses to biliary obstruction. Male Sprague-Dawley rats underwent common bile duct ligation and subcutaneous injection of either saline or octreotide (6 micrograms/kg) twice daily for 7 days. Morphometric analysis of hepatocytes, bile duct epithelial cells, and periportal connective tissue was performed by computerized point counting. Hepatocyte volume was preserved with octreotide treatment, which also significantly decreased bile duct proliferation and periportal extracellular matrix deposition in response to biliary obstruction compared with saline treated, duct-ligated animals. These results indicate that octreotide prevents the morphological changes that accompany extrahepatic biliary obstruction. Images Figure 1 PMID:8256850

  8. Targeting sphingosine kinase 1 attenuates bleomycin-induced pulmonary fibrosis.

    PubMed

    Huang, Long Shuang; Berdyshev, Evgeny; Mathew, Biji; Fu, Panfeng; Gorshkova, Irina A; He, Donghong; Ma, Wenli; Noth, Imre; Ma, Shwu-Fan; Pendyala, Srikanth; Reddy, Sekhar P; Zhou, Tong; Zhang, Wei; Garzon, Steven A; Garcia, Joe G N; Natarajan, Viswanathan

    2013-04-01

    Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive interstitial lung disease, wherein transforming growth factor β (TGF-β) and sphingosine-1-phosphate (S1P) contribute to the pathogenesis of fibrosis. However, the in vivo contribution of sphingosine kinase (SphK) in fibrotic processes has not been documented. Microarray analysis of blood mononuclear cells from patients with IPF and SphK1- or SphK2-knockdown mice and SphK inhibitor were used to assess the role of SphKs in fibrogenesis. The expression of SphK1/2 negatively correlated with lung function and survival in patients with IPF. Also, the expression of SphK1 was increased in lung tissues from patients with IPF and bleomycin-challenged mice. Knockdown of SphK1, but not SphK2, increased survival and resistance to pulmonary fibrosis in bleomycin-challenged mice. Administration of SphK inhibitor reduced bleomycin-induced mortality and pulmonary fibrosis in mice. Knockdown of SphK1 or treatment with SphK inhibitor attenuated S1P generation and TGF-β secretion in a bleomycin-induced lung fibrosis mouse model that was accompanied by reduced phosphorylation of Smad2 and MAPKs in lung tissue. In vitro, bleomycin-induced expression of SphK1 in lung fibroblast was found to be TGF-β dependent. Taken together, these data indicate that SphK1 plays a critical role in the pathology of lung fibrosis and is a novel therapeutic target.

  9. Amniotic fluid-derived mesenchymal stem cells prevent fibrosis and preserve renal function in a preclinical porcine model of kidney transplantation.

    PubMed

    Baulier, Edouard; Favreau, Frederic; Le Corf, Amélie; Jayle, Christophe; Schneider, Fabrice; Goujon, Jean-Michel; Feraud, Olivier; Bennaceur-Griscelli, Annelise; Hauet, Thierry; Turhan, Ali G

    2014-07-01

    It is well known that ischemia/reperfusion injuries strongly affect the success of human organ transplantation. Development of interstitial fibrosis and tubular atrophy is the main deleterious phenomenon involved. Stem cells are a promising therapeutic tool already validated in various ischemic diseases. Amniotic fluid-derived mesenchymal stem cells (af-MSCs), a subpopulation of multipotent cells identified in amniotic fluid, are known to secrete growth factors and anti-inflammatory cytokines. In addition, these cells are easy to collect, present higher proliferation and self-renewal rates compared with other adult stem cells (ASCs), and are suitable for banking. Consequently, af-MSCs represent a promising source of stem cells for regenerative therapies in humans. To determine the efficiency and the safety of af-MSC infusion in a preclinical porcine model of renal autotransplantation, we injected autologous af-MSCs in the renal artery 6 days after transplantation. The af-MSC injection improved glomerular and tubular functions, leading to full renal function recovery and abrogated fibrosis development at 3 months. The strong proof of concept generated by this translational porcine model is a first step toward evaluation of af-MSC-based therapies in human kidney transplantation.

  10. The Mitochondrial Cardiolipin Remodeling Enzyme Lysocardiolipin Acyltransferase Is a Novel Target in Pulmonary Fibrosis

    PubMed Central

    Huang, Long Shuang; Mathew, Biji; Zhao, Yutong; Noth, Imre; Reddy, Sekhar P.; Harijith, Anantha; Usatyuk, Peter V.; Berdyshev, Evgeny V.; Kaminski, Naftali; Zhou, Tong; Zhang, Wei; Zhang, Yanmin; Rehman, Jalees; Kotha, Sainath R.; Gurney, Travis O.; Parinandi, Narasimham L.; Lussier, Yves A.; Garcia, Joe G. N.

    2014-01-01

    Rationale: Lysocardiolipin acyltransferase (LYCAT), a cardiolipin-remodeling enzyme regulating the 18:2 linoleic acid pattern of mammalian mitochondrial cardiolipin, is necessary for maintaining normal mitochondrial function and vascular development. We hypothesized that modulation of LYCAT expression in lung epithelium regulates development of pulmonary fibrosis. Objectives: To define a role for LYCAT in human and murine models of pulmonary fibrosis. Methods: We analyzed the correlation of LYCAT expression in peripheral blood mononuclear cells (PBMCs) with the outcomes of pulmonary functions and overall survival, and used the murine models to establish the role of LYCAT in fibrogenesis. We studied the LYCAT action on cardiolipin remodeling, mitochondrial reactive oxygen species generation, and apoptosis of alveolar epithelial cells under bleomycin challenge. Measurements and Main Results: LYCAT expression was significantly altered in PBMCs and lung tissues from patients with idiopathic pulmonary fibrosis (IPF), which was confirmed in two preclinical murine models of IPF, bleomycin- and radiation-induced pulmonary fibrosis. LYCAT mRNA expression in PBMCs directly and significantly correlated with carbon monoxide diffusion capacity, pulmonary function outcomes, and overall survival. In both bleomycin- and radiation-induced pulmonary fibrosis murine models, hLYCAT overexpression reduced several indices of lung fibrosis, whereas down-regulation of native LYCAT expression by siRNA accentuated fibrogenesis. In vitro studies demonstrated that LYCAT modulated bleomycin-induced cardiolipin remodeling, mitochondrial membrane potential, reactive oxygen species generation, and apoptosis of alveolar epithelial cells, potential mechanisms of LYCAT-mediated lung protection. Conclusions: This study is the first to identify modulation of LYCAT expression in fibrotic lungs and offers a novel therapeutic approach for ameliorating lung inflammation and pulmonary fibrosis. PMID

  11. Protective Role of Insulin-Like Growth Factor-1 Receptor in Endothelial Cells against Unilateral Ureteral Obstruction–Induced Renal Fibrosis

    PubMed Central

    Liang, Ming; Woodard, Lauren E.; Liang, Anlin; Luo, Jinlong; Wilson, Matthew H.; Mitch, William E.; Cheng, Jizhong

    2016-01-01

    Insulin-like growth factor-1 receptor (IGF-1R) can regulate vascular homeostasis and endothelial function. We studied the role of IGF-1R in oxidative stress–induced endothelial dysfunction. Unilateral ureteral obstruction (UUO) was performed in wild-type (WT) mice and mice with endothelial cell (EC)–specific IGF-1R knockout (KO). After UUO in endothelial IGF-1R KO mice, endothelial barrier dysfunction was more severe than in WT mice, as seen by increased inflammatory cell infiltration and vascular endothelial (VE)–cadherin phosphorylation. UUO in endothelial IGF-1R KO mice increased interstitial fibroblast accumulation and enhanced extracellular protein deposition as compared with the WT mice. Endothelial barrier function measured by transendothelial migration in response to hydrogen peroxide (H2O2) was impaired in ECs. Silencing IGF-1R enhanced the influence of H2O2 in disrupting the VE–protein tyrosine phosphatase/VE-cadherin interaction. Overexpression of IGF-1R suppressed H2O2-induced endothelial barrier dysfunction. Furthermore, by using the piggyBac transposon system, we expressed IGF-1R in VE cells in mice. The expression of IGF-1R in ECs also suppressed the inflammatory cell infiltration and renal fibrosis induced by UUO. IGF-1R KO in the VE-cadherin lineage of bone marrow cells had no significant effect on the UUO-induced fibrosis, as compared with control mice. Our results indicate that IGF-1R in the endothelium maintains the endothelial barrier function by stabilization of the VE–protein tyrosine phosphatase/VE-cadherin complex. Decreased expression of IGF-1R impairs endothelial function and increases the fibrosis of kidney disease. PMID:25783760

  12. Quercetin attenuates the activation of hepatic stellate cells and liver fibrosis in mice through modulation of HMGB1-TLR2/4-NF-κB signaling pathways.

    PubMed

    Li, Xi; Jin, Qianwen; Yao, Qunyan; Xu, Beili; Li, Zheng; Tu, Chuantao

    2016-11-02

    This study aimed to investigate the effects of quercetin on liver fibrogenesis in mice and to elucidate the underlying molecular mechanisms. Mice were administered with carbon tetrachloride (CCl4) for eight weeks to induce liver fibrosis and concomitantly orally treated with quercetin (50mgkg(-1)day(-1)). Here, we demonstrated that quercetin dramatically ameliorated liver injury, inflammation, and hepatic fibrogenesis induced by CCl4. Quercetin also inhibited the activation of hepatic stellate cells (HSC) in vivo and in vitro, as evaluated by α-smooth muscle actin (α-SMA) expression, which is a specific marker of HSC activation. Moreover, reduced fibrosis was associated with decreased high-mobility group box 1 (HMGB1), toll like receptor (TLR) 2 and TLR4 genes, and protein expression. Quercetin also inhibited the cytoplasmic translocation of HMGB1 in hepatocytes of fibrotic livers. Further investigation demonstrated that quercetin treatment significantly attenuated CCl4-induced nuclear translocation of the nuclear factor-κB (NF-κB) p65 and inhibited degradation of IκBα (an inhibitor of NF-κB) expression in the liver compared with vehicle-treated fibrotic mice. Considered together, our data indicate that quercetin has hepatoprotective and anti-fibrotic effects in animal models of liver fibrosis, the mechanism of which may be involved in modulating the HMGB1-TLR2/4-NF-κB signaling pathways.

  13. TSG-6 released from intradermally injected mesenchymal stem cells accelerates wound healing and reduces tissue fibrosis in murine full-thickness skin wounds.

    PubMed

    Qi, Yu; Jiang, Dongsheng; Sindrilaru, Anca; Stegemann, Agatha; Schatz, Susanne; Treiber, Nicolai; Rojewski, Markus; Schrezenmeier, Hubert; Vander Beken, Seppe; Wlaschek, Meinhard; Böhm, Markus; Seitz, Andreas; Scholz, Natalie; Dürselen, Lutz; Brinckmann, Jürgen; Ignatius, Anita; Scharffetter-Kochanek, Karin

    2014-02-01

    Proper activation of macrophages (Mφ) in the inflammatory phase of acute wound healing is essential for physiological tissue repair. However, there is a strong indication that robust Mφ inflammatory responses may be causal for the fibrotic response always accompanying adult wound healing. Using a complementary approach of in vitro and in vivo studies, we here addressed the question of whether mesenchymal stem cells (MSCs)-due to their anti-inflammatory properties-would control Mφ activation and tissue fibrosis in a murine model of full-thickness skin wounds. We have shown that the tumor necrosis factor-α (TNF-α)-stimulated protein 6 (TSG-6) released from MSCs in co-culture with activated Mφ or following injection into wound margins suppressed the release of TNF-α from activated Mφ and concomitantly induced a switch from a high to an anti-fibrotic low transforming growth factor-β1 (TGF-β1)/TGF-β3 ratio. This study provides insight into what we believe to be a previously undescribed multifaceted role of MSC-released TSG-6 in wound healing. MSC-released TSG-6 was identified to improve wound healing by limiting Mφ activation, inflammation, and fibrosis. TSG-6 and MSC-based therapies may thus qualify as promising strategies to enhance tissue repair and to prevent excessive tissue fibrosis.

  14. Inhibition of airway epithelial-to-mesenchymal transition and fibrosis by kaempferol in endotoxin-induced epithelial cells and ovalbumin-sensitized mice.

    PubMed

    Gong, Ju-Hyun; Cho, In-Hee; Shin, Daekeun; Han, Seon-Young; Park, Sin-Hye; Kang, Young-Hee

    2014-03-01

    Chronic airway remodeling is characterized by structural changes within the airway wall, including smooth muscle hypertrophy, submucosal fibrosis and epithelial shedding. Epithelial-to-mesenchymal transition (EMT) is a fundamental mechanism of organ fibrosis, which can be induced by TGF-β. In the in vitro study, we investigated whether 1-20 μM kaempferol inhibited lipopolysaccharide (LPS)-induced bronchial EMT in BEAS-2B cells. The in vivo study explored demoting effects of 10-20 mg/kg kaempferol on airway fibrosis in BALB/c mice sensitized with ovalbumin (OVA). LPS induced airway epithelial TGF-β1 signaling that promoted EMT with concurrent loss of E-cadherin and induction of α-smooth muscle actin (α-SMA). Nontoxic kaempferol significantly inhibited TGF-β-induced EMT process through reversing E-cadherin expression and retarding the induction of N-cadherin and α-SMA. Consistently, OVA inhalation resulted in a striking loss of epithelial morphology by displaying myofibroblast appearance, which led to bronchial fibrosis with submucosal accumulation of collagen fibers. Oral administration of kaempferol suppressed collagen deposition, epithelial excrescency and goblet hyperplasia observed in the lung of OVA-challenged mice. The specific inhibition of TGF-β entailed epithelial protease-activated receptor-1 (PAR-1) as with 20 μM kaempferol. The epithelial PAR-1 inhibition by SCH-79797 restored E-cadherin induction and deterred α-SMA induction, indicating that epithelial PAR-1 localization was responsible for resulting in airway EMT. These results demonstrate that dietary kaempferol alleviated fibrotic airway remodeling via bronchial EMT by modulating PAR1 activation. Therefore, kaempferol may be a potential therapeutic agent targeting asthmatic airway constriction.

  15. The ΔF508 Mutation in the Cystic Fibrosis Transmembrane Conductance Regulator Is Associated With Progressive Insulin Resistance and Decreased Functional β-Cell Mass in Mice.

    PubMed

    Fontés, Ghislaine; Ghislain, Julien; Benterki, Isma; Zarrouki, Bader; Trudel, Dominique; Berthiaume, Yves; Poitout, Vincent

    2015-12-01

    Cystic fibrosis (CF) is the result of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR). CF-related diabetes affects 50% of adult CF patients. How CFTR deficiency predisposes to diabetes is unknown. Herein, we examined the impact of the most frequent cftr mutation in humans, deletion of phenylalanine at position 508 (ΔF508), on glucose homeostasis in mice. We compared ΔF508 mutant mice with wild-type (WT) littermates. Twelve-week-old male ΔF508 mutants had lower body weight, improved oral glucose tolerance, and a trend toward higher insulin tolerance. Glucose-induced insulin secretion was slightly diminished in ΔF508 mutant islets, due to reduced insulin content, but ΔF508 mutant islets were not more sensitive to proinflammatory cytokines than WT islets. Hyperglycemic clamps confirmed an increase in insulin sensitivity with normal β-cell function in 12- and 18-week-old ΔF508 mutants. In contrast, 24-week-old ΔF508 mutants exhibited insulin resistance and reduced β-cell function. β-Cell mass was unaffected at 11 weeks of age but was significantly lower in ΔF508 mutants versus controls at 24 weeks. This was not associated with gross pancreatic pathology. We conclude that the ΔF508 CFTR mutation does not lead to an intrinsic β-cell secretory defect but is associated with insulin resistance and a β-cell mass deficit in aging mutants.

  16. Fat-storing cells and myofibroblasts are involved in the initial phase of carbon tetrachloride-induced hepatic fibrosis in BN/BiRij rats.

    PubMed Central

    Seifert, W. F.; Roholl, P. J.; Blauw, B.; van der Ham, F.; van Thiel-De Ruiter, C. F.; Seifert-Bock, I.; Bosma, A.; Knook, D. L.; Brouwer, A.

    1994-01-01

    This study on the appearance, distribution and kinetics of fibroblast-like cells (fat-storing cells, transitional cells, myofibroblasts and fibroblasts) after CCl4-treatment was undertaken to delineate further the respective roles of these cell types in liver fibrogenesis. The different cell types were distinguished on the basis of their immunophenotypic pattern with a combination of marker antibodies and on the basis of ultrastructural characteristics. Combined staining for alpha-smooth muscle actin (sma) and desmin (Des) revealed perisinusoidal fat-storing cells (FSC) as d+ sma- and myofibroblasts around the central veins of the normal rat liver as d+ sma+. During the initial phase of CCl4-induced hepatic fibrosis (week 1 and 2), the number of d+ sma+ cells increased in the degenerating area around the central veins and d+ sma+ cells appeared in the very thin fibrotic septa at week 2. Ultrastructural examination of the affected central areas showed the presence of myofibroblasts. These sma+ cells proliferated, as shown by double staining for bromodeoxyuridine (BrdU) and sma. In degenerating parenchymal areas, d+ sma- FSC were present. The FSC in the perisinusoidal space of areas which were not affected by CCl4 intoxification, remained d+ sma-. These immunostaining findings support the electron microscopical results, which show the presence of cells with the typical ultrastructural characteristics of FSC in both the degenerating areas and the perisinusoidal space of unaffected areas. After one week of CCl4-treatment, enhanced deposition of procollagen type III was observed around the central veins. Enhanced deposition of collagen type IV was seen subendothelially along the sinusoids, notably in degenerating parenchymal areas where the septa were later formed. FSC appear to be the principal source of collagen type IV during fibrogenesis. These observations further support and specify the role of FSC in early fibrogenesis. With the progression of the CCl4-induced

  17. Hypoxia-Induced Epithelial-Mesenchymal Transition Is Involved in Bleomycin-Induced Lung Fibrosis.

    PubMed

    Guo, Liang; Xu, Jun-mei; Liu, Lei; Liu, Su-mei; Zhu, Rong

    2015-01-01

    Pulmonary fibrosis is a severe disease that contributes to the morbidity and mortality of a number of lung diseases. However, the molecular and cellular mechanisms leading to lung fibrosis are poorly understood. This study investigated the roles of epithelial-mesenchymal transition (EMT) and the associated molecular mechanisms in bleomycin-induced lung fibrosis. The bleomycin-induced fibrosis animal model was established by intratracheal injection of a single dose of bleomycin. Protein expression was measured by Western blot, immunohistochemistry, and immunofluorescence. Typical lesions of lung fibrosis were observed 1 week after bleomycin injection. A progressive increase in MMP-2, S100A4, α-SMA, HIF-1α, ZEB1, CD44, phospho-p44/42 (p-p44/42), and phospho-p38 MAPK (p-p38) protein levels as well as activation of EMT was observed in the lung tissues of bleomycin mice. Hypoxia increased HIF-1α and ZEB1 expression and activated EMT in H358 cells. Also, continuous incubation of cells under mild hypoxic conditions increased CD44, p-p44/42, and p-p38 protein levels in H358 cells, which correlated with the increase in S100A4 expression. In conclusion, bleomycin induces progressive lung fibrosis, which may be associated with activation of EMT. The fibrosis-induced hypoxia may further activate EMT in distal alveoli through a hypoxia-HIF-1α-ZEB1 pathway and promote the differentiation of lung epithelial cells into fibroblasts through phosphorylation of p38 MAPK and Erk1/2 proteins.

  18. C/EBP homologous protein (CHOP) deficiency ameliorates renal fibrosis in unilateral ureteral obstructive kidney disease.

    PubMed

    Liu, Shing-Hwa; Wu, Cheng-Tien; Huang, Kuo-How; Wang, Ching-Chia; Guan, Siao-Syun; Chen, Li-Ping; Chiang, Chih-Kang

    2016-04-19

    Renal tubulointerstitial fibrosis is an important pathogenic feature in chronic kidney disease and end-stage renal disease, regardless of the initiating insults. A recent study has shown that CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP) is involved in acute ischemia/reperfusion-related acute kidney injury through oxidative stress induction. However, the influence of CHOP on chronic kidney disease-correlated renal fibrosis remains unclear. Here, we investigated the role of CHOP in unilateral ureteral obstruction (UUO)-induced experimental chronic tubulointerstital fibrosis. The CHOP knockout and wild type mice with or without UUO were used. The results showed that the increased expressions of renal fibrosis markers collagen I, fibronectin, α-smooth muscle actin, and plasminogen activator inhibitor-1 in the kidneys of UUO-treated wild type mice were dramatically attenuated in the kidneys of UUO-treated CHOP knockout mice. CHOP deficiency could also ameliorate lipid peroxidation and endogenous antioxidant enzymes depletion, tubular apoptosis, and inflammatory cells infiltration in the UUO kidneys. These results suggest that CHOP deficiency not only attenuates apoptotic death and oxidative stress in experimental renal fibrosis, but also reduces local inflammation, leading to diminish UUO-induced renal fibrosis. Our findings support that CHOP may be an important signaling molecule in the progression of chronic kidney disease.

  19. Genetics Home Reference: retroperitoneal fibrosis

    MedlinePlus

    ... 1 link) Retroperitoneal fibrosis Educational Resources (3 links) Disease InfoSearch: Retroperitoneal fibrosis MalaCards: retroperitoneal fibrosis Orphanet: IgG4-related retroperitoneal fibrosis Patient Support and Advocacy Resources (2 ...

  20. Modulation of Stem Cells Differentiation and Myostatin as an approach to Counteract fibrosis in Muscle Dystrophy and Regeneration after Injury

    DTIC Science & Technology

    2010-03-01

    as advanced age and diabetes ) and Dupuytren contracture, another localized fibrotic process. Most of the current pharmacological treatments for PD...also seen in rat models of diabetic nephropathy , experimental glomerulonephritis, myocardial infarction and hypertrophy, and pulmonary fibrosis;91...valderrama, E. Chronic diabetic nephropathy : role of inducible nitric oxide synthase. Pediatr. Nephrol. 17, 20–29 (2002). 67. Chen, Y. et al. Deficiency of

  1. COMPASS identifies T-cell subsets correlated with clinical outcomes.

    PubMed

    Lin, Lin; Finak, Greg; Ushey, Kevin; Seshadri, Chetan; Hawn, Thomas R; Frahm, Nicole; Scriba, Thomas J; Mahomed, Hassan; Hanekom, Willem; Bart, Pierre-Alexandre; Pantaleo, Giuseppe; Tomaras, Georgia D; Rerks-Ngarm, Supachai; Kaewkungwal, Jaranit; Nitayaphan, Sorachai; Pitisuttithum, Punnee; Michael, Nelson L; Kim, Jerome H; Robb, Merlin L; O'Connell, Robert J; Karasavvas, Nicos; Gilbert, Peter; C De Rosa, Stephen; McElrath, M Juliana; Gottardo, Raphael

    2015-06-01

    Advances in flow cytometry and other single-cell technologies have enabled high-dimensional, high-throughput measurements of individual cells as well as the interrogation of cell population heterogeneity. However, in many instances, computational tools to analyze the wealth of data generated by these technologies are lacking. Here, we present a computational framework for unbiased combinatorial polyfunctionality analysis of antigen-specific T-cell subsets (COMPASS). COMPASS uses a Bayesian hierarchical framework to model all observed cell subsets and select those most likely to have antigen-specific responses. Cell-subset responses are quantified by posterior probabilities, and human subject-level responses are quantified by two summary statistics that describe the quality of an individual's polyfunctional response and can be correlated directly with clinical outcome. Using three clinical data sets of cytokine production, we demonstrate how COMPASS improves characterization of antigen-specific T cells and reveals cellular 'correlates of protection/immunity' in the RV144 HIV vaccine efficacy trial that are missed by other methods. COMPASS is available as open-source software.

  2. COMPASS identifies T-cell subsets correlated with clinical outcomes

    PubMed Central

    Lin, Lin; Finak, Greg; Ushey, Kevin; Seshadri, Chetan; Hawn, Thomas R.; Frahm, Nicole; Scriba, Thomas J.; Mahomed, Hassan; Hanekom, Willem; Bart, Pierre-Alexandre; Pantaleo, Giuseppe; Tomaras, Georgia D.; Rerks-Ngarm, Supachai; Kaewkungwal, Jaranit; Nitayaphan, Sorachai; Pitisuttithum, Punnee; Michael, Nelson L.; Kim, Jerome H.; Robb, Merlin L.; O’Connell, Robert J.; Karasavvas, Nicos; Gilbert, Peter; DeRosa, Stephen; McElrath, M. Juliana

    2015-01-01

    Advances in flow cytometry and other single-cell technologies have enabled high-dimensional, high-throughput measurements of individual cells and allowed interrogation of cell population heterogeneity. Computational tools to take full advantage of these technologies are lacking. Here, we present COMPASS, a computational framework for unbiased polyfunctionality analysis of antigen-specific T-cell subsets. COMPASS uses a Bayesian hierarchical framework to model all observed functional cell subsets and select those most likely to exhibit antigen-specific responses. Cell-subset responses are quantified by posterior probabilities, while subject-level responses are quantified by two novel summary statistics that can be correlated directly with clinical outcome, and describe the quality of an individual’s (poly)functional response. Using three clinical datasets of cytokine production we demonstrate how COMPASS improves characterization of antigen-specific T cells and reveals novel cellular correlates of protection in the RV144 HIV vaccine efficacy trial that are missed by other methods. COMPASS is available as open-source software. PMID:26006008

  3. Chemokine receptor Cxcr4 contributes to kidney fibrosis via multiple effectors.

    PubMed

    Yuan, Amy; Lee, Yashang; Choi, Uimook; Moeckel, Gilbert; Karihaloo, Anil

    2015-03-01

    Kidney fibrosis is the final common pathway for virtually every type of chronic kidney disease and is a consequence of a prolonged healing response that follows tissue inflammation. Chronic kidney inflammation ultimately leads to progressive tissue injury and scarring/fibrosis. Several pathways have been implicated in the progression of kidney fibrosis. In the present study, we demonstrate that G protein-coupled chemokine (C-X-C motif) receptor (CXCR)4 was significantly upregulated after renal injury and that sustained activation of Cxcr4 expression augmented the fibrotic response. We demonstrate that after unilateral ureteral obstruction (UUO), both gene and protein expression of Cxcr4 were highly upregulated in tubular cells of the nephron. The increased Cxcr4 expression in tubules correlated with their increased dedifferentiated state, leading to increased mRNA expression of platelet-derived growth factor (PDGF)-α, transforming growth factor (TGF)-β1, and concurrent loss of bone morphogenetic protein 7 (Bmp7). Ablation of tubular Cxcr4 attenuated UUO-mediated fibrotic responses, which correlated with a significant reduction in PDGF-α and TGF-β1 levels and preservation of Bmp7 expression after UUO. Furthermore, Cxcr4(+) immune cells infiltrated the obstructed kidney and further upregulate their Cxcr4 expression. Genetic ablation of Cxcr4 from macrophages was protective against UUO-induced fibrosis. There was also reduced total kidney TGF-β1, which correlated with reduced Smad activation and α-smooth muscle actin levels. We conclude that chronic high Cxcr4 expression in multiple effector cell types can contribute to the pathogenesis of renal fibrosis by altering their biological profile. This study uncovered a novel cross-talk between Cxcr4-TGF-β1 and Bmp7 pathways and may provide novel targets for interrupting the progression of fibrosis.

  4. Preliminary Evidence for Cell Membrane Amelioration in Children with Cystic Fibrosis by 5-MTHF and Vitamin B12 Supplementation: A Single Arm Trial

    PubMed Central

    Scambi, Cinzia; De Franceschi, Lucia; Guarini, Patrizia; Poli, Fabio; Siciliano, Angela; Pattini, Patrizia; Biondani, Andrea; La Verde, Valentina; Bortolami, Oscar; Turrini, Francesco; Carta, Franco; D'Orazio, Ciro; Assael, Baroukh M.; Faccini, Giovanni; Bambara, Lisa M.

    2009-01-01

    Background Cystic fibrosis (CF) is one of the most common fatal autosomal recessive disorders in the Caucasian population caused by mutations of gene for the cystic fibrosis transmembrane conductance regulator (CFTR). New experimental therapeutic strategies for CF propose a diet supplementation to affect the plasma membrane fluidity and to modulate amplified inflammatory response. The objective of this study was to evaluate the efficacy of 5-methyltetrahydrofolate (5-MTHF) and vitamin B12 supplementation for ameliorating cell plasma membrane features in pediatric patients with cystic fibrosis. Methodology and Principal Findings A single arm trial was conducted from April 2004 to March 2006 in an Italian CF care centre. 31 children with CF aged from 3 to 8 years old were enrolled. Exclusion criteria were diabetes, chronic infections of the airways and regular antibiotics intake. Children with CF were supplemented for 24 weeks with 5-methyltetrahydrofolate (5-MTHF, 7.5 mg /day) and vitamin B12 (0.5 mg/day). Red blood cells (RBCs) were used to investigate plasma membrane, since RBCs share lipid, protein composition and organization with other cell types. We evaluated RBCs membrane lipid composition, membrane protein oxidative damage, cation content, cation transport pathways, plasma and RBCs folate levels and plasma homocysteine levels at baseline and after 24 weeks of 5-MTHF and vitamin B12 supplementation. In CF children, 5-MTHF and vitamin B12 supplementation (i) increased plasma and RBC folate levels; (ii) decreased plasma homocysteine levels; (iii) modified RBC membrane phospholipid fatty acid composition; (iv) increased RBC K+ content; (v) reduced RBC membrane oxidative damage and HSP70 membrane association. Conclusion and Significance 5-MTHF and vitamin B12 supplementation might ameliorate RBC membrane features of children with CF. Trial Registration ClinicalTrials.gov NCT00730509 PMID:19277125

  5. Diagnostic Values For Club Cell Secretory Protein (CC16) in Serum of Patients of Combined Pulmonary Fibrosis and Emphysema.

    PubMed

    Kokuho, Nariaki; Ishii, Takeo; Kamio, Koichiro; Hayashi, Hiroki; Kurahara, Misuzu; Hattori, Kumiko; Motegi, Takashi; Azuma, Arata; Gemma, Akihiko; Kida, Kozui

    2015-08-01

    Combined pulmonary fibrosis and emphysema (CPFE) is an under-recognized syndrome for which the diagnostic use of serum biomarkers is an attractive possibility. We hypothesized that CC16 and/or TGF-β1 or combinations with other biomarkers are useful for diagnosing CPFE. Patients with respiratory symptoms and a smoking history, with or without chronic obstructive pulmonary disease, were divided into the following three groups according to findings of high-resolution computed tomography of the chest: controls without either emphysema or fibrosis, patients with emphysema alone, and patients compatible with the diagnosis of CPFE. Serum concentrations of CC16, TGF-β1, SP-D, and KL-6 were measured in patients whose condition was stable for at least 3 months. To investigate changes in biomarkers of lung fibrosis in patients with a life-long smoking history, additional measurements were performed on the patients with idiopathic pulmonary fibrosis (IPF) of smoking history. The mean age of the first three groups was 68.0 years, whereas that of the IPF group was 71.8 years, and the groups contained 36, 115, 27, and 10 individuals, respectively. The serum concentration of CC16 in the four groups was 5.67 ± 0.42, 5.66 ± 0.35, 9.38 ± 1.04 and 22.15 ± 4.64 ng/ml, respectively, indicating that those patients with lung fibrosis had a significantly higher concentration. The combined use of CC16, SP-D, and KL-6 provided supportive diagnosis in conjunction with radiological imaging in diagnosis of CPFE. We conclude that a combination of biomarkers including CC16 could provide useful information to screen and predict the possible diagnosis of CPFE.

  6. Macrophage-to-Myofibroblast Transition Contributes to Interstitial Fibrosis in Chronic Renal Allograft Injury.

    PubMed

    Wang, Ying-Ying; Jiang, Hong; Pan, Jun; Huang, Xiao-Ru; Wang, Yu-Cheng; Huang, Hong-Feng; To, Ka-Fai; Nikolic-Paterson, David J; Lan, Hui-Yao; Chen, Jiang-Hua

    2017-02-16

    Interstitial fibrosis is an important contributor to graft loss in chronic renal allograft injury. Inflammatory macrophages are associated with fibrosis in renal allografts, but how these cells contribute to this damaging response is not clearly understood. Here, we investigated the role of macrophage-to-myofibroblast transition in interstitial fibrosis in human and experimental chronic renal allograft injury. In biopsy specimens from patients with active chronic allograft rejection, we identified cells undergoing macrophage-to-myofibroblast transition by the coexpression of macrophage (CD68) and myofibroblast (α-smooth muscle actin [α-SMA]) markers. CD68(+)/α-SMA(+) cells accounted for approximately 50% of the myofibroblast population, and the number of these cells correlated with allograft function and the severity of interstitial fibrosis. Similarly, in C57BL/6J mice with a BALB/c renal allograft, cells coexpressing macrophage markers (CD68 or F4/80) and α-SMA composed a significant population in the interstitium of allografts undergoing chronic rejection. Fate-mapping in Lyz2-Cre/Rosa26-Tomato mice showed that approximately half of α-SMA(+) myofibroblasts in renal allografts originated from recipient bone marrow-derived macrophages. Knockout of Smad3 protected against interstitial fibrosis in renal allografts and substantially reduced the number of macrophage-to-myofibroblast transition cells. Furthermore, the majority of macrophage-to-myofibroblast transition cells in human and experimental renal allograft rejection coexpressed the M2-type macrophage marker CD206, and this expression was considerably reduced in Smad3-knockout recipients. In conclusion, our studies indicate that macrophage-to-myofibroblast transition contributes to interstitial fibrosis in chronic renal allograft injury. Moreover, the transition of bone marrow-derived M2-type macrophages to myofibroblasts in the renal allograft is regulated via a Smad3-dependent mechanism.

  7. The proportion of different interleukin-17-producing T-cell subsets is associated with liver fibrosis in chronic hepatitis C.

    PubMed

    Cachem, Fabio C O F; Dias, Aleida S; Monteiro, Clarice; Castro, José Roberto; Fernandes, Gabriel; Delphim, Letícia; Almeida, Adilson J; Tavares, Felipe; Maciel, Alessandra M A; Amendola-Pires, Marcia M; Brandão-Mello, Carlos E; Bento, Cleonice A M

    2017-01-31

    Studies have suggested the pivotal role of T helper type 1 (Th1) -related cytokines on the outcome of hepatitis C virus (HCV) infection. Nevertheless, the role of different interleukin-17 (IL-17) -secreting T cells on chronic hepatitis C (CHC) is less clear. Here, the in vivo IL-1β, IL-6, and IL-17 levels were positively correlated with both alanine transaminase (ALT) levels and hepatic lesions. When compared with the control group, CHC patients showed a lower proportion of IL-17-secreting (CD4(+) and CD8(+) ) T cells capable of simultaneously producing IL-21. Moreover, the percentage of IL-10-secreting Th17 cells was also lower in CHC patients. Notably, advanced liver lesions were observed among those patients with lower percentage levels of IL-17-producing T cells positive for IL-21, interferon-γ (IFN-γ) and IL-10. In contrast, the severity of hepatic damage was associated with peripheral single IL-17(+) T cells. The percentage of IL-17(+) IL-21(-) IFN-γ(+) (CD4(+) and CD8(+) ) T-cell phenotypes was positively associated with plasma CD14 levels. Finally, elevated levels of circulating CD14 were detected among CHC patients with extensive liver damage. In summary, although preliminary, our results suggest that a balance between different IL-17-producing T cells, associated with peripheral levels of CD14, may be a progress marker for liver disease in chronically HCV-infected patients.

  8. Fibrosis and Simple Cysts

    MedlinePlus

    ... caffeine and other stimulants found in coffee, tea, chocolate, and many soft drinks. Studies have not found ... side effects. How do fibrosis and simple cysts affect your risk for breast cancer? Neither fibrosis nor ...

  9. What Causes Cystic Fibrosis?

    MedlinePlus

    ... page from the NHLBI on Twitter. What Causes Cystic Fibrosis? A defect in the CFTR gene causes cystic ... in the severity of the disease. How Is Cystic Fibrosis Inherited? Every person inherits two CFTR genes—one ...

  10. Pulmonary Fibrosis Foundation

    MedlinePlus

    ... for a friend. Ways to Give Announcements CHICAGO BEAR JORDAN HOWARD BRINGS HIS FIGHT TO PULMONARY FIBROSIS ... 22-year-old lead rusher for the Chicago Bears, will announce his commitment to fight pulmonary fibrosis ( ...

  11. Nucleoplasmic viscosity of living cells investigated by fluorescence correlation spectroscopy

    NASA Astrophysics Data System (ADS)

    Liang, Lifang; Xing, Da; Chen, Tongshen; Pei, Yihui

    2007-11-01

    Fluorescence correlation spectroscopy (FCS) is a new kind of real-time, high-speed and single-molecule technique. It is used to detect the kinetic characteristics of fluorescent dye such as diffusion coefficient in the aqueous solution. Combined with confocal microscope optics, it has been now widely applied in cell biological research. Through a time correlation analysis of spontaneous intensity fluctuations, this technique with EGFP as a probe is capable of determining viscosity of fluids according to Stokes-Einstein equation. Nucleoplasmic viscosity is an important physical parameter to quantify the rheological characteristics of the nucleoplasm. Investigation on nucleoplasmic viscosity plays an important role in further understanding intranuclear environment. In this paper, FCS is introduced to noninvasively investigate nucleoplasmic viscosity of living cells. The results show that nucleoplasmic viscosity of lung adenocarcinoma (ASTC-a-1) cells is 2.55+/-0.61 cP and nucleoplasmic viscosity is larger than cytoplasmic viscosity at 37 °C (pH 7.4). In addition, significant changes in nucleoplasmic viscosity are detected by FCS when cells are exposed to hyper or hypotonic medium. Our study suggests that FCS can be used to detect the kinetic characteristics of biomolecules in living cells and thus helps to investigate the dynamic changes of the microenvironment in the cell.

  12. Cross Talk Free Fluorescence Cross Correlation Spectroscopy in Live Cells

    PubMed Central

    Thews, Elmar; Gerken, Margarita; Eckert, Reiner; Zäpfel, Johannes; Tietz, Carsten; Wrachtrup, Jörg

    2005-01-01

    Fluorescence correlation spectroscopy (FCS) is now a widely used technique to measure small ensembles of labeled biomolecules with single molecule detection sensitivity (e.g., low endogenous concentrations). Fluorescence cross correlation spectroscopy (FCCS) is a derivative of this technique that detects the synchronous movement of two biomolecules with different fluorescence labels. Both methods can be applied to live cells and, therefore, can be used to address a variety of unsolved questions in cell biology. Applications of FCCS with autofluorescent proteins (AFPs) have been hampered so far by cross talk between the detector channels due to the large spectral overlap of the fluorophores. Here we present a new method that combines advantages of these techniques to analyze binding behavior of proteins in live cells. To achieve this, we have used dual color excitation of a common pair of AFPs, ECFP and EYFP, being discriminated in excitation rather than in emission. This is made possible by pulsed excitation and detection on a shorter timescale compared to the average residence time of particles in the FCS volume element. By this technique we were able to eliminate cross talk in the detector channels and obtain an undisturbed cross correlation signal. The setup was tested with ECFP/EYFP lysates as well as chimeras as negative and positive controls and demonstrated to work in live HeLa cells coexpressing the two fusion proteins ECFP-connexin and EYFP-connexin. PMID:15951373

  13. Comparison of the therapeutic effectiveness of human CD34+ and rat bone marrow mesenchymal stem cells on improvement of experimental liver fibrosis in Wistar rats

    PubMed Central

    Sayyed, Hayam G; Osama, Amany; Idriss, Naglaa K; Sabry, Dina; Abdelrhim, Azza S; Bakry, Rania

    2016-01-01

    Background and objective: Human umbilical cord blood (UCB) cells and bone marrow mesenchymal stem cells (BM-MSCs) have numerous advantages as grafts for cell transplantation. We hypothesized differing impacts of human UCB cells and rat BM-MSCs on reversal of hepatic injury and revival of liver function in carbon tetrachloride (CCl4)-induced liver fibrosis. Methods: Forty rats were divided into 4 groups; control group, CCl4 group, CCl4/CD34+ group and CCl4/BM-MSCs group. Blood samples were driven from rats at 4, 8 and 12 weeks to measure serum concentration of albumin and alanine aminotransferase (ALT). Quantitative expression of collagen Iα, TGF-β, α-SMA, albumin, MMP-2, MMP-9 and TNF-α were assessed by polymerase chain reaction. Histopathological examination of the liver tissue was performed. GFP labeled cells were detected in groups injected with stem cells. Results: Regarding liver function, CD34+ were more efficient than BM-MSCs in elevating albumin (P<0.05) and reducing ALT (P<0.05) concentrations. Concerning gene expression, CD34+ were more effective than BM-MSCs in reducing gene expressions of collagen Iα (P<0.01), TGF-β1 (P<0.01) and α-SMA (P<0.01). Both CD34+ and BM-MSCs have the same efficacy in reducing TNF-α (P<0.001 and P<0.01, respectively). Furthermore, CD34+ were more valuable than BM-MSCs in increasing gene expression of albumin (P<0.05) and MMP-9 (P<0.01). Conclusion: Taken together; human UCB CD34+ stem cells were more efficient in improvement of experimental liver injury than BM-MSCs. This study highlighted an important role of human UCB CD34+ stem cells in liver fibrosis therapy. PMID:27785340

  14. Pancreatic Fat Accumulation, Fibrosis, and Acinar Cell Injury in the Zucker Diabetic Fatty Rat Fed a Chronic High-Fat Diet

    PubMed Central

    Matsuda, Akiko; Makino, Naohiko; Tozawa, Tomohiro; Shirahata, Nakao; Honda, Teiichiro; Ikeda, Yushi; Sato, Hideyuki; Ito, Miho; Kakizaki, Yasuharu; Akamatsu, Manabu; Ueno, Yoshiyuki; Kawata, Sumio

    2014-01-01

    Objective The histological alteration of the exocrine pancreas in obesity has not been clarified. In the present study, we investigated biochemical and histological changes in the exocrine pancreas of obese model rats. Methods Zucker lean rats were fed a standard diet, and Zucker diabetic fatty (ZDF) rats were divided into 2 groups fed a standard diet and a high-fat diet, respectively. These experimental groups were fed each of the diets from 6 weeks until 12, 18, 24 weeks of age. We performed blood biochemical assays and histological analysis of the pancreas. Results In the ZDF rats fed a high-fat diet, the ratio of accumulated pancreatic fat area relative to exocrine gland area was increased significantly at 18 weeks of age in comparison with the other 2 groups (P < 0.05), and lipid droplets were observed in acinar cells. Subsequently, at 24 weeks of age in this group, pancreatic fibrosis and the serum exocrine pancreatic enzyme levels were increased significantly relative to the other 2 groups (P < 0.01). Conclusions In ZDF rats fed a chronic high-fat diet, fat accumulates in pancreatic acinar cells, and this fatty change seems to be related to subsequent pancreatic fibrosis and acinar cell injury. PMID:24717823

  15. Hsian-tsao (Mesona procumbens Heml.) prevents against rat liver fibrosis induced by CCl(4) via inhibition of hepatic stellate cells activation.

    PubMed

    Shyu, Ming-Huan; Kao, Tzu-Chien; Yen, Gow-Chin

    2008-12-01

    In this study, the protective effect of extract of Hsian-tsao (Mesona procumbens) (EHT) against liver fibrogenesis in carbon tetrachloride (CCl(4))-injured rats was evaluated. The inhibitory effect of oleanolic acid (OA) and ursolic acid (UA), which are the active compounds in EHT, on the activation of hepatic stellate cells (HSC) was also determined. The results showed that EHT at a dosage of 1.2g/kg of b.w. significantly reduced the liver injury induced by CCl(4) in rats. It also decreased the activity of serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) and the deposition of collagen in the liver. Oral administration of EHT reduced the levels of alpha-smooth muscle actin (alpha-SMA) and the activity of metalloproteinases (MMPs) in rats injured by treatment with CCl(4). In addition, we performed experiments with the rat hepatic stellate cell line HSC-T6 in which we induced the expression of MMP-2 and alpha-SMA with phorbol-12-myristate-13-acetate (PMA). Treating these cells with OA (20microM) or UA (10microM) caused a decrease in the levels of both proteins. Taken together, our data indicate that EHT can efficiently inhibit CCl(4)-induced liver fibrosis in rats. EHT may therefore be a useful functional food for preventing liver fibrosis.

  16. TNF-alpha, but not IFN-gamma, regulates CCN2 (CTGF), collagen type I, and proliferation in mesangial cells: possible roles in the progression of renal fibrosis.

    PubMed

    Cooker, Laurinda A; Peterson, Darryl; Rambow, Joann; Riser, Melisa L; Riser, Rebecca E; Najmabadi, Feridoon; Brigstock, David; Riser, Bruce L

    2007-07-01

    Connective tissue growth factor (CCN2) is a profibrotic factor acting downstream and independently of TGF-beta to mediate renal fibrosis. Although inflammation is often involved in the initiation and/or progression of fibrosis, the role of inflammatory cytokines in regulation of glomerular CCN2 expression, cellular proliferation, and extracellular matrix accumulation is unknown. We studied two such cytokines, TNF-alpha and IFN-gamma, for their effects on cultured mesangial cells in the presence or absence of TGF-beta, as a model for progressive renal fibrosis. Short-term treatment with TNF-alpha, like TGF-beta, significantly increased secreted CCN2 per cell, but unlike TGF-beta inhibited cellular replication. TNF-alpha combined with TGF-beta further increased CCN2 secretion and mRNA levels and reduced proliferation. Surprisingly, however, TNF-alpha treatment decreased baseline collagen type I protein and mRNA levels and largely blocked their stimulation by TGF-beta. Long-term treatment with TGF-beta or TNF-alpha alone no longer increased CCN2 protein levels. However, the combination synergistically increased CCN2. IFN-gamma had no effect on either CCN2 or collagen activity and produced a mild inhibition of TGF-beta-induced collagen only at a high concentration (500 U/ml). In summary, we report a strong positive regulatory role for TNF-alpha, but not IFN-gamma, in CCN2 production and secretion, including that driven by TGF-beta. The stimulation of CCN2 release by TNF-alpha, unlike TGF-beta, is independent of cellular proliferation and not linked to increased collagen type I accumulation. This suggests that the paradigm of TGF-beta-driven CCN2 with subsequent collagen production may be overridden by an as yet undefined inhibitory mechanism acting either directly or indirectly on matrix metabolism.

  17. Bergamot (Citrus bergamia Risso) fruit extracts and identified components alter expression of interleukin 8 gene in cystic fibrosis bronchial epithelial cell lines

    PubMed Central

    2011-01-01

    Background Cystic fibrosis (CF) airway pathology is a fatal, autosomal, recessive genetic disease characterized by extensive lung inflammation. After induction by TNF-α, elevated concentrations of several pro-inflammatory cytokines (i.e. IL-6, IL-1β) and chemokines (i.e. IL-8) are released from airway epithelial cells. In order to reduce the excessive inflammatory response in the airways of CF patients, new therapies have been developed and in this respect, medicinal plant extracts have been studied. In this article we have investigated the possible use of bergamot extracts (Citrus bergamia Risso) and their identified components to alter the expression of IL-8 associated with the cystic fibrosis airway pathology. Methods The extracts were chemically characterized by 1H-NMR (nuclear magnetic resonance), GC-FID (gas chromatography-flame ionization detector), GC-MS (gas chromatography-mass spectrometry) and HPLC (high pressure liquid chromatography). Both bergamot extracts and main detected chemical constituents were assayed for their biological activity measuring (a) cytokines and chemokines in culture supernatants released from cystic fibrosis IB3-1 cells treated with TNF-α by Bio-Plex cytokine assay; (b) accumulation of IL-8 mRNA by real-time PCR. Results The extracts obtained from bergamot (Citrus bergamia Risso) epicarps contain components displaying an inhibitory activity on IL-8. Particularly, the most active molecules were bergapten and citropten. These effects have been confirmed by analyzing mRNA levels and protein release in the CF cellular models IB3-1 and CuFi-1 induced with TNF-α or exposed to heat-inactivated Pseudomonas aeruginosa. Conclusions These obtained results clearly indicate that bergapten and citropten are strong inhibitors of IL-8 expression and could be proposed for further studies to verify possible anti-inflammatory properties to reduce lung inflammation in CF patients. PMID:21496221

  18. Detecting Nanodomains in Living Cell Membrane by Fluorescence Correlation Spectroscopy

    NASA Astrophysics Data System (ADS)

    He, Hai-Tao; Marguet, Didier

    2011-05-01

    Cell membranes actively participate in numerous cellular functions. Inasmuch as bioactivities of cell membranes are known to depend crucially on their lateral organization, much effort has been focused on deciphering this organization on different length scales. Within this context, the concept of lipid rafts has been intensively discussed over recent years. In line with its ability to measure diffusion parameters with great precision, fluorescence correlation spectroscopy (FCS) measurements have been made in association with innovative experimental strategies to monitor modes of molecular lateral diffusion within the plasma membrane of living cells. These investigations have allowed significant progress in the characterization of the cell membrane lateral organization at the suboptical level and have provided compelling evidence for the in vivo existence of raft nanodomains. We review these FCS-based studies and the characteristic structural features of raft nanodomains. We also discuss the findings in regards to the current view of lipid rafts as a general membrane-organizing principle.

  19. [Nucleoplasmic viscosity of living cells investigated by fluorescence correlation spectroscopy].

    PubMed

    Liang, Li-Fang; Da, Xing; Chen, Tong-Sheng; Pei, Yi-Hui

    2009-02-01

    In order to non-invasively investigate nucleoplasmic viscosity in real time with good temporal resolution, the present study firstly introduced a new method based on fluorescence correlation spectroscopy (FCS). FCS is a kind of single-molecule technique with high temporal and spatial resolution to analyze the dynamics of fluorescent molecules in nanomolar concentration. Through a time correlation analysis of spontaneous intensity fluctuations, this technique in conjunction with EGFP as a probe is capable of determining nucleoplasmic viscosity in terms of Stokes-Einstein equation as well as its corresponding analysis of the diffusion coefficient for EGFP in the nucleus. The results showed that nucleoplasmic viscosity of ASTC-a-1 cells and HeLa cells were respectively (2.55 +/- 0.61) cP and (2.04 +/- 0.49) cP at pH 7.4 and 37 degrees C, consistent with the results by traditional methods, and nucleoplasmic viscosity was found to be larger than cytoplasmic viscosity. Meanwhile, the real-time analysis of nucleoplasmic viscosity in living cells exposed to hypotonic media proved that FCS could be used to track the changing rheological characteristics of the nucleoplasm in living cells. Taken together, this study suggests that FCS provides an accurate and non-invasive method to investigate the microenvironment in living cells on the femtoliter scale and it can be used as a powerful tool in researches on the dynamical processes of intracellular molecules.

  20. Puerarin attenuates renal fibrosis by reducing oxidative stress induced-epithelial cell apoptosis via MAPK signal pathways in vivo and in vitro.

    PubMed

    Zhou, Xiangjun; Bai, Chen; Sun, Xinbo; Gong, Xiaoxin; Yang, Yong; Chen, Congbo; Shan, Guang; Yao, Qisheng

    2017-11-01

    Puerarin (PR) is an isoflavonoid isolated from the root of the plant Pueraria lobata and has been widely used in traditional Chinese herbal medicine for the treatment of various diseases. Oxidative stress and epithelial cell apoptosis play important roles in the renal fibrotic process. The present study aimed to determine whether or not PR inhibits renal fibrosis by reducing oxidative stress induced-epithelial cell apoptosis. In vivo, unilateral ureteral obstruction (UUO) induced renal fibrosis, and epithelial cell apoptosis. A total of 24 mice were randomly assigned to four experimental groups: sham, UUO alone, UUO +50 mg/kg PR, and UUO +100 mg/kg PR. In vitro, 200 μM hydrogen peroxide (H2O2) induced epithelial cell apoptosis. The experiments were dived into four groups: control, H2O2 alone, H2O2+50 μM PR, and H2O2+100 μM PR. Tubular injury was measured in the renal cortex of the mice through periodic acid-Schiff (PAS) staining, and the extracellular matrix (ECM) was measured through Sirius red (SR), immunohistochemistry (IHC) staining, and Western blot. Renal epithelial cell apoptosis was measured through terminal deoxynucleotidyl transferase-mediated dUTP Nick-End labeling (TUNEL), flow cytometry (FCM), and Hoechst assays. The protein expression of NOX4, caspase3, ERK, P38, and JNK was assessed through Western blot. PAS staining showed that PR decreased renal tubular injury in UUO mice. SR and IHC staining demonstrated that PR decreased the accumulation of ECM. PR treatment significantly inhibited epithelial cell apoptosis according to the results of TUNEL, FCM, Hoechst, and Western blot. Furthermore, NOX4 increased in UUO mice and decreased with PR treatment. H2O2-derived oxidative stress activated epithelial apoptosis and mitogen-activated protein kinases (MAPK), and PR treatment significantly reversed it. These results suggest that PR treatment ameliorates renal fibrosis by inhibiting oxidative stress induced-epithelial cell apoptosis through

  1. Transforming growth factor-β-sphingosine kinase 1/S1P signaling upregulates microRNA-21 to promote fibrosis in renal tubular epithelial cells.

    PubMed

    Liu, Xiujuan; Hong, Quan; Wang, Zhen; Yu, Yanyan; Zou, Xin; Xu, Lihong

    2016-02-01

    Renal fibrosis is a progressive pathological change characterized by tubular cell apoptosis, tubulointerstitial fibroblast proliferation, and excessive deposition of extracellular matrix (ECM). miR-21 has been implicated in transforming growth factor-β (TGF-β)-stimulated tissue fibrosis. Recent studies showed that sphingosine kinase/sphingosine-1-phosphate (SphK/S1P) are also critical for TGF-β-stimulated tissue fibrosis; however, it is not clear whether SphK/S1P interacts with miR-21 or not. In this study, we hypothesized that SphK/S1P signaling is linked to upregulation of miR-21 by TGF-β. To verify this hypothesis, we first determined that miR-21 was highly expressed in renal tubular epithelial cells (TECs) stimulated with TGF-β by using qRT-PCR and Northern blotting. Simultaneously, inhibition of miR-21, mediated by the corresponding antimir, markedly decreased the expression and deposition of type I collagen, fibronectin (Fn), cysteine-rich protein 61 (CCN1), α-smooth muscle actin, and fibroblast-specific protein1 in TGF-β-treated TECs. ELISA and qRT-PCR were used to measure the S1P and SphK1 levels in TECs. S1P production was induced by TGF-β through activation of SphK1. Furthermore, it was observed that TGF-β-stimulated upregulation of miR-21 was abolished by SphK1 siRNA and was restored by the addition of exogenous S1P. Blocking S1PR2 also inhibited upregulation of miR-21. Additionally, miR-21 overexpression attenuated the repression of TGF-β-stimulated ECM deposition and epithelial-mesenchymal transition by SphK1 and S1PR2 siRNA. In summary, our study demonstrates a link between SphK1/S1P and TGF-β-induced miR-21 in renal TECs and may represent a novel therapeutic target in renal fibrosis.

  2. Antibiotic Tolerance Induced by Lactoferrin in Clinical Pseudomonas aeruginosa Isolates from Cystic Fibrosis Patients

    PubMed Central

    Andrés, María T.; Viejo-Diaz, Mónica; Pérez, Francisco; Fierro, José F.

    2005-01-01

    Lactoferrin-induced cell depolarization and a delayed tobramycin-killing effect on Pseudomonas aeruginosa cells were correlated. This antibiotic tolerance effect (ATE) reflects the ability of a defense protein to modify the activity of an antibiotic as a result of its modulatory effect on bacterial physiology. P. aeruginosa isolates from cystic fibrosis patients showed higher ATE values (≤6-fold) than other clinical strains. PMID:15793153

  3. Chloride and potassium channels in cystic fibrosis airway epithelia

    NASA Astrophysics Data System (ADS)

    Welsh, Michael J.; Liedtke, Carole M.

    1986-07-01

    Cystic fibrosis, the most common lethal genetic disease in Caucasians, is characterized by a decreased permeability in sweat gland duct and airway epithelia. In sweat duct epithelium, a decreased Cl- permeability accounts for the abnormally increased salt content of sweat1. In airway epithelia a decreased Cl- permeability, and possibly increased sodium absorption, may account for the abnormal respiratory tract fluid2,3. The Cl- impermeability has been localized to the apical membrane of cystic fibrosis airway epithelial cells4. The finding that hormonally regulated Cl- channels make the apical membrane Cl- permeable in normal airway epithelial cells5 suggested abnormal Cl- channel function in cystic fibrosis. Here we report that excised, cell-free patches of membrane from cystic fibrosis epithelial cells contain Cl- channels that have the same conductive properties as Cl- channels from normal cells. However, Cl- channels from cystic fibrosis cells did not open when they were attached to the cell. These findings suggest defective regulation of Cl- channels in cystic fibrosis epithelia; to begin to address this issue, we performed two studies. First, we found that isoprenaline, which stimulates Cl- secretion, increases cellular levels of cyclic AMP in a similar manner in cystic fibrosis and non-cystic fibrosis epithelial cells. Second, we show that adrenergic agonists open calcium-activated potassium channels, indirectly suggesting that calcium-dependent stimulus-response coupling is intact in cystic fibrosis. These data suggest defective regulation of Cl- channels at a site distal to cAMP accumulation.

  4. Lineage correlations of single cell division time as a probe of cell-cycle dynamics.

    PubMed

    Sandler, Oded; Mizrahi, Sivan Pearl; Weiss, Noga; Agam, Oded; Simon, Itamar; Balaban, Nathalie Q

    2015-03-26

    Stochastic processes in cells are associated with fluctuations in mRNA, protein production and degradation, noisy partition of cellular components at division, and other cell processes. Variability within a clonal population of cells originates from such stochastic processes, which may be amplified or reduced by deterministic factors. Cell-to-cell variability, such as that seen in the heterogeneous response of bacteria to antibiotics, or of cancer cells to treatment, is understood as the inevitable consequence of stochasticity. Variability in cell-cycle duration was observed long ago; however, its sources are still unknown. A central question is whether the variance of the observed distribution originates from stochastic processes, or whether it arises mostly from a deterministic process that only appears to be random. A surprising feature of cell-cycle-duration inheritance is that it seems to be lost within one generation but to be still present in the next generation, generating poor correlation between mother and daughter cells but high correlation between cousin cells. This observation suggests the existence of underlying deterministic factors that determine the main part of cell-to-cell variability. We developed an experimental system that precisely measures the cell-cycle duration of thousands of mammalian cells along several generations and a mathematical framework that allows discrimination between stochastic and deterministic processes in lineages of cells. We show that the inter- and intra-generation correlations reveal complex inheritance of the cell-cycle duration. Finally, we build a deterministic nonlinear toy model for cell-cycle inheritance that reproduces the main features of our data. Our approach constitutes a general method to identify deterministic variability in lineages of cells or organisms, which may help to predict and, eventually, reduce cell-to-cell heterogeneity in various systems, such as cancer cells under treatment.

  5. Imaging characteristics of local recurrences after stereotactic body radiation therapy for stage I non-small cell lung cancer: Evaluation of mass-like fibrosis

    PubMed Central

    Hayashi, Shinya; Tanaka, Hidekazu; Hoshi, Hiroaki

    2015-01-01

    Background This study aimed to evaluate stereotactic body radiation therapy (SBRT) in patients with stage I non-small cell lung cancer (NSCLC) in terms of radiation-induced changes and computed tomography (CT) features of local recurrence by 18F-fluorodeoxyglucose positron emission tomography (18F-FDG-PET). Methods From January 2006 to December 2012, 81 patients with NSCLC received SBRT. Follow-up consisted of non-contrast enhanced CT scans performed before and every four months after SBRT. In addition, 18F-FDG-PET/CT was conducted before SBRT for each patient, and one year later for each case suspected of recurrence. The CT findings were classified into two categories: mass-like fibrosis and others. The mass-like fibrosis category was subdivided into two patterns: mass-like consolidation (with air bronchogram) and mass-like opacity. Results Six patients had histologically confirmed local recurrence, including 83% (5/6) with mass-like opacity pattern and one case of modified conventional pattern (P = 0.02). In contrast, the non-recurrent group exhibited only 7% (5/75) with mass-like opacity and 13% (10/75) with mass-like consolidation pattern. Five patients with local recurrence presented with the mass-like opacity pattern, compared with 33% of patients (5/15) from the non-recurrent group (P = 0.01) and showed an increase in maximum diameter at ≥12 months after SBRT. The recurrent group also had a significantly higher standardized uptake value (SUVmax) than the non-recurrent group (P < 0.001), with all values >5 (range: 5.7–25.4). Conclusion The following characteristics of mass-like fibrosis should be considered indicators of local recurrence after SBRT: opacity pattern, increasing maximum diameter, and SUVmax > 5. PMID:26273357

  6. Vitamin D Can Ameliorate Chlorhexidine Gluconate-Induced Peritoneal Fibrosis and Functional Deterioration through the Inhibition of Epithelial-to-Mesenchymal Transition of Mesothelial Cells

    PubMed Central

    Lee, Yi-Che; Hung, Shih-Yuan; Liou, Hung-Hsiang; Lin, Tsun-Mei; Tsai, Chu-Hung; Lin, Sheng-Hsiang; Tsai, Yau-Sheng; Chang, Min-Yu; Wang, Hsi-Hao; Ho, Li-Chun; Chen, Yi-Ting; Wu, Ching-Fang; Chen, Ho-Ching; Chen, Hsin-Pao; Liu, Kuang-Wen; Chen, Chih-I.; She, Kuan Min; Wang, Hao-Kuang; Lin, Chi-Wei; Chiou, Yuan-Yow

    2015-01-01

    Background. Peritoneal dialysis (PD) can induce fibrosis and functional alterations in PD patients' peritoneal membranes, due to long-term unphysiological dialysate exposure, partially occurring via triggering of epithelial-to-mesenchymal transition (EMT) in peritoneal mesothelial cells (MCs). Vitamin D can ameliorate these negative effects; however, the mechanism remains unexplored. Therefore, we investigated its possible links to MCs EMT inhibition. Methods. Peritoneal fibrosis was established in Sprague-Dawley rats by chlorhexidine gluconate (CG) intraperitoneal injection for 21 days, with and without 1α,25(OH)2D3 treatment. Morphological and functional evaluation and western blot analysis of EMT marker were performed upon peritoneum tissue. In vitro study was also performed in a primary human peritoneal MC culture system; MCs were incubated with transforming growth factor-β1 (TGF-β1) in the absence or presence of 1α,25(OH)2D3. EMT marker expression, migration activities, and cytoskeleton redistribution of MCs were determined. Results. 1α,25(OH)2D3 ameliorated CG-induced morphological and functional deterioration in animal model, along with CG-induced upregulation of α-SMA and downregulation of E-cadherin expression. Meanwhile, 1α,25(OH)2D3 also ameliorated TGF-β1-induced decrease in E-cadherin expression, increase in Snai1 and α-SMA expression, intracellular F-actin redistribution, and migration activity in vitro. Conclusion. 1α,25(OH)2D3 can ameliorate CG-induced peritoneal fibrosis and attenuate functional deterioration through inhibiting MC EMT. PMID:26495304

  7. Relationships among CFTR expression, HCO3− secretion, and host defense may inform gene- and cell-based cystic fibrosis therapies

    PubMed Central

    Shah, Viral S.; Ernst, Sarah; Tang, Xiao Xiao; Karp, Philip H.; Parker, Connor P.; Ostedgaard, Lynda S.; Welsh, Michael J.

    2016-01-01

    Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel. Airway disease is the major source of morbidity and mortality. Successful implementation of gene- and cell-based therapies for CF airway disease requires knowledge of relationships among percentages of targeted cells, levels of CFTR expression, correction of electrolyte transport, and rescue of host defense defects. Previous studies suggested that, when ∼10–50% of airway epithelial cells expressed CFTR, they generated nearly wild-type levels of Cl− secretion; overexpressing CFTR offered no advantage compared with endogenous expression levels. However, recent discoveries focused attention on CFTR-mediated HCO3− secretion and airway surface liquid (ASL) pH as critical for host defense and CF pathogenesis. Therefore, we generated porcine airway epithelia with varying ratios of CF and wild-type cells. Epithelia with a 50:50 mix secreted HCO3− at half the rate of wild-type epithelia. Likewise, heterozygous epithelia (CFTR+/− or CFTR+/∆F508) expressed CFTR and secreted HCO3− at ∼50% of wild-type values. ASL pH, antimicrobial activity, and viscosity showed similar relationships to the amount of CFTR. Overexpressing CFTR increased HCO3− secretion to rates greater than wild type, but ASL pH did not exceed wild-type values. Thus, in contrast to Cl− secretion, the amount of CFTR is rate-limiting for HCO3− secretion and for correcting host defense abnormalities. In addition, overexpressing CFTR might produce a greater benefit than expressing CFTR at wild-type levels when targeting small fractions of cells. These findings may also explain the risk of airway disease in CF carriers. PMID:27114540

  8. Bone marrow transplantation alters lung antigen presenting cells to promote TH17 response and the development of pneumonitis and fibrosis following gammaherpesvirus infection

    PubMed Central

    Zhou, Xiaofeng; Loomis-King, Hillary; Gurczynski, Stephen J.; Wilke, Carol A.; Konopka, Kristine E.; Ptaschinski, Catherine; Coomes, Stephanie M; Iwakura, Yoichiro; van Dyk, Linda F.; Lukacs, Nicholas W.; Moore, Bethany B.

    2015-01-01

    Hematopoietic stem cell transplantation (HSCT) efficacy is limited by numerous pulmonary complications. We developed a model of syngeneic bone marrow transplant (BMT) followed by infection with murine gamma herpesvirus (γHV-68) that results in pneumonitis and fibrosis and mimics human “non-infectious” HSCT complications. BMT mice experience increased early lytic replication, but establish viral latency by 21 days post infection (dpi). CD4 T cells in BMT mice are skewed towards IL-17A rather than IFN-γ production. Transplantation of bone marrow from Il-17a−/− donors or treatment with anti-IL-17A neutralization antibodies at late stages attenuates pneumonitis and fibrosis in infected BMT mice, suggesting that hematopoietic-derived IL-17A is essential for development of pathology. IL-17A directly influences activation and extracellular matrix production by lung mesenchymal cells. Lung CD11c+ cells of BMT mice secrete more TGF-β1, and pro-TH17 mRNAs for IL-23 and IL-6, and less TH1-promoting cytokine mRNA for IFN-γ but slightly more IL-12 mRNA in response to viral infection. Adoptive transfer of non-BMT lung CD11c-enriched cells restores robust TH1 response and suppresses aberrant TH17 response in BMT mice to improve lung pathology. Our data suggest “non-infectious” HSCT lung complications may reflect preceding viral infections and demonstrate that IL-17A neutralization may offer therapeutic advantage even after disease onset. PMID:26376362

  9. MicroRNA-130a and -130b enhance activation of hepatic stellate cells by suppressing PPARγ expression: A rat fibrosis model study

    SciTech Connect

    Lu, Le; Wang, Jinlong; Lu, Hongwei; Zhang, Guoyu; Liu, Yang; Wang, Jiazhong; Zhang, Yafei; Shang, Hao; Ji, Hong; Chen, Xi; Duan, Yanxia; Li, Yiming

    2015-09-25

    Hepatic stellate cells (HSCs) are the primary sources of extracellular matrix (ECM) in normal and fibrotic liver. Peroxisome proliferator-activated receptor gamma (PPARγ) maintains HSCs in a quiescent state, and its downregulation induces HSC activation. MicroRNAs (miRNAs) can induce PPARγ mRNA degradation, but the mechanism by which miRNAs regulate PPARγ in rat HSCs is unclear. This study aimed to investigate some miRNAs which putatively bind to the 3′-untranslated region (3′-UTR) of PPARγ mRNA, and increase expression of ECM genes in rat HSCs. In carbon tetrachloride injection (CCl{sub 4}) and common bile duct ligation (CBDL) liver fibrosis models, miRNAs miR-130a, miR-130b, miR-301a, miR-27b and miR-340 levels were found to be increased and PPARγ expression decreased. Overexpression of miR-130a and miR-130b enhanced cell proliferation by involving Runx3. MiR-130a and miR-130b decreased PPARγ expression by targeting the 3′-UTR of PPARγ mRNA in rat HSC-T6 cells. Transforming growth factor-β1 (TGF-β1) may mediate miR-130a and miR-130b overexpression, PPARγ downregulation, and ECM genes overexpression in cell culture. These findings suggest that miR-130a and miR-130b are involved in downregulation of PPARγ in liver fibrosis. - Highlights: • MiR-130a and miR-130b are increased and PPARγ is decreased in liver fibrosis models. • MiR-130a and miR-130b decreased PPARγ by targeting the 3′-UTR of PPARγ mRNA. • MiR-130a and miR-130b enhanced HSC cell proliferation by involving Runx3. • TGF-β1 may mediate miR-130a and miR-130b overexpression.

  10. Induction by TNF-α of IL-6 and IL-8 in Cystic Fibrosis Bronchial IB3-1 Epithelial Cells Encapsulated in Alginate Microbeads

    PubMed Central

    Borgatti, Monica; Mazzitelli, Stefania; Breveglieri, Giulia; Gambari, Roberto; Nastruzzi, Claudio

    2010-01-01

    We have developed a microencapsulation procedure for the entrapment and manipulation of IB3-1 cystic fibrosis cells. The applied method is based on generation of monodisperse droplets by a vibrational nozzle. Different experimental parameters were analyzed, including frequency and amplitude of vibration, polymer pumping rate and distance between the nozzle and the gelling bath. We have found that the microencapsulation procedure does not alter the viability of the encapsulated IB3-1 cells. The encapsulated IB3-1 cells were characterized in term of secretomic profile, analyzing the culture medium by Bio-Plex strategy. The experiments demonstrated that most of the analyzed proteins, were secreted both by the free and encapsulated cells, even if in a different extent. In order to determine the biotechnological applications of this procedure, we determined whether encapsulated IB3-1 cells could be induced to pro-inflammatory responses, after treatment with TNF-α. In this experimental set-up, encapsulated and free IB3-1 cells were treated with TNF-α, thereafter the culture media from both cell populations were collected. As expected, TNF-α induced a sharp increase in the secretion of interleukins, chemokines and growth factors. Of great interest was the evidence that induction of interleukin-6 and interleukin-8 occurs also by encapsulated IB3-1 cells. PMID:20936184

  11. Glibenclamide induces apoptosis through inhibition of cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channels and intracellular Ca(2+) release in HepG2 human hepatoblastoma cells.

    PubMed

    Kim, J A; Kang, Y S; Lee, S H; Lee, E H; Yoo, B H; Lee, Y S

    1999-08-11

    Glibenclamide, an inhibitor of cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channels, induced apoptosis in a dose- and time-dependent manner in HepG2 human hepatoblastoma cells. Glibenclamide increased intracellular Ca(2+) concentration, which was significantly inhibited by Ca(2+) release blockers dantrolene and TMB-8. BAPTA/AM, an intracellular Ca(2+) chelator, and the Ca(2+) release blockers significantly inhibited glibenclamide-induced apoptosis. Glibanclamide also increased intracellular Cl(-) concentration, which was significantly blocked by CFTR Cl(-) channel activators levamisole and bromotetramisole. These activators also significantly inhibited both intracellular Ca(2+) release and apoptosis induced by glibenclamide. The expression of CFTR protein in the cells was confirmed by Western blot analysis. These results suggest that glibenclamide induced apoptosis through inhibition of CFTR Cl(-) channels and intracellular Ca(2+) release and that this protein may be a good target for treatment of human hepatomas.

  12. Melatonin: the dawning of a treatment for fibrosis?

    PubMed

    Hu, Wei; Ma, Zhiqiang; Jiang, Shuai; Fan, Chongxi; Deng, Chao; Yan, Xiaolong; Di, Shouyin; Lv, Jianjun; Reiter, Russel J; Yang, Yang

    2016-03-01

    Fibrosis is a common occurrence following organ injury and failure. To date, there is no effective treatment for this condition. Melatonin targets numerous molecular pathways, a consequence of its antioxidant and anti-inflammatory actions that reduce excessive fibrosis. Herein, we review the multiple protective effects of melatonin against fibrosis. There exist four major phases of the fibrogenic response including primary injury to the organ, activation of effector cells, the elaboration of extracellular matrix (ECM) and dynamic deposition. Melatonin regulates each of these phases. Additionally, melatonin reduces fibrosis levels in numerous organs. Melatonin exhibits its anti-fibrosis effects in heart, liver, lung, kidney, and other organs. In addition, adhesions which occur following surgical procedures are also inhibited by melatonin. The information reviewed here should be significant to understanding the protective role of melatonin against fibrosis, contribute to the design of further experimental studies related to melatonin and the fibrotic response and shed light on a potential treatment for fibrosis.

  13. Therapeutic potentials occurring during the early differentiation process of mesenchymal stem cells in a rats model with thioacetamide-induced liver fibrosis

    PubMed Central

    Choi, Sang-Tae; Hong, Hea-Nam; Won, You-Jin; Ahn, Chul-Soo; Ha, Tae-Yong; Song, Gi-Won; Jung, Dong-Hwan; Park, Gil-Chun; Lee, Sung-Gyu

    2013-01-01

    Backgrounds/Aims Mesenchymal stem cells (MSCs) have the capacity to differentiate into hepatocytes, The purpose of this study is to investigate the MSCs' differentiation process and therapeutic potentials by comparing isolated MSCs with HGF-treated MSCs in rat's model with thiacetamide (TAA)-induced cirrhosis. Methods Male Sprague-Dawley (SD) rats, weighing 100-150 g were used in this study. To induce liver fibrosis, recipient rats were taken with 0.04% thioacetamide (TAA) in the drinking water (400 mg TAA/L) for 8 weeks. The rats underlying liver cirrhosis were divided into 3 groups according to the transplanted materials, compared to normal saline as control (I) and isolated MSCs (II) HGF-treated MSCs. Results Severe hepatic fibrosis and hepatocyte destruction were detected in the control group. Less hepatic cirrhosis and collagen formation, more hepatocyte regeneration and glycogen storage were detected in isolated MSCs compared to HGF-treated MSCs group, Distribution of red autofluorescence is mainly localized near the sinusoids in isolated MSCs, scattered away the sinusoids in HGF-treated MSCs group. MSCs transdifferentiated into CK-19 postive Oval cells and then to albulmin-producing hepatocytes, HGF treated MSCs differentiated into hepatocyte without the intermediate oval cells phase. HGF treated MSCs became the CK18-positive, MSCs became CD 90-positive. Conclusions Significant hepatocyte differentiation occurred in not HGF-treated MSCs but isolated MSCs group unexpectedly. These results suggest that the beneficial effect of MSCs on in rat's model with TAA-induced cirrhosis may occur during early differentiation course of MSCs. Mature hepatocyte itself has a little effect on the accelerated differentiation and functional capacity of hepatic lineage cell-line. PMID:26155209

  14. Design, synthesis, and characterization of novel apigenin analogues that suppress pancreatic stellate cell proliferation in vitro and associated pancreatic fibrosis in vivo.

    PubMed

    Chen, Haijun; Mrazek, Amy A; Wang, Xiaofu; Ding, Chunyong; Ding, Ye; Porro, Laura J; Liu, Huiling; Chao, Celia; Hellmich, Mark R; Zhou, Jia

    2014-07-01

    Accumulating evidence suggests that activated pancreatic stellate cells (PSC) play an important role in chronic pancreatitis (CP), and inhibition of the activated PSC is considered as a potential strategy for the treatment and prevention of CP. Herein, we disclose our findings that apigenin and its novel analogues suppress the proliferation and induce apoptosis in PSC, which reduce the PSC-mediated fibrosis in CP. Chemical modifications of apigenin have been directed to build a focused library of O-alkylamino-tethered apigenin derivatives at 4'-O position of the ring C with the attempt to enhance the potency and drug-like properties including aqueous solubility. A number of compounds such as 14, 16, and 24 exhibited potent antiproliferative effects as well as improved aqueous solubility. Intriguingly, apigenin, new analogues 23 and 24 displayed significant efficacy to reduce pancreatic fibrosis even at a low dose of 0.5mg/kg in our proof-of-concept study using a preclinical in vivo mouse model of CP.

  15. The Role of PPARs in Lung Fibrosis

    PubMed Central

    Lakatos, Heather F.; Thatcher, Thomas H.; Kottmann, R. Matthew; Garcia, Tatiana M.; Phipps, Richard P.; Sime, Patricia J.

    2007-01-01

    Pulmonary fibrosis is a group of disorders characterized by accumulation of scar tissue in the lung interstitium, resulting in loss of alveolar function, destruction of normal lung architecture, and respiratory distress. Some types of fibrosis respond to corticosteroids, but for many there are no effective treatments. Prognosis varies but can be poor. For example, patients with idiopathic pulmonary fibrosis (IPF) have a median survival of only 2.9 years. Prognosis may be better in patients with some other types of pulmonary fibrosis, and there is variability in survival even among individuals with biopsy-proven IPF. Evidence is accumulating that the peroxisome proliferator-activated receptors (PPARs) play important roles in regulating processes related to fibrogenesis, including cellular differentiation, inflammation, and wound healing. PPARα agonists, including the hypolidipemic fibrate drugs, inhibit the production of collagen by hepatic stellate cells and inhibit liver, kidney, and cardiac fibrosis in animal models. In the mouse model of lung fibrosis induced by bleomycin, a PPARα agonist significantly inhibited the fibrotic response, while PPARα knockout mice developed more serious fibrosis. PPARβ/δ appears to play a critical role in regulating the transition from inflammation to wound healing. PPARβ/δ agonists inhibit lung fibroblast proliferation and enhance the antifibrotic properties of PPARγ agonists. PPARγ ligands oppose the profibrotic effect of TGF-β, which induces differentiation of fibroblasts to myofibroblasts, a critical effector cell in fibrosis. PPARγ ligands, including the thiazolidinedione class of antidiabetic drugs, effectively inhibit lung fibrosis in vitro and in animal models. The clinical availability of potent and selective PPARα and PPARγ agonists should facilitate rapid development of successful treatment strategies based on current and ongoing research. PMID:17710235

  16. Macrophage Activation in Pediatric Nonalcoholic Fatty Liver Disease (NAFLD) Correlates with Hepatic Progenitor Cell Response via Wnt3a Pathway

    PubMed Central

    Renzi, Anastasia; De Stefanis, Cristiano; Stronati, Laura; Franchitto, Antonio; Alisi, Anna; Onori, Paolo; De Vito, Rita; Alpini, Gianfranco; Gaudio, Eugenio

    2016-01-01

    Non-alcoholic fatty liver disease is one of the most important causes of liver-related morbidity in children. In non-alcoholic fatty liver disease, the activation of liver resident macrophage pool is a central event in the progression of liver injury. The aims of the present study were to evaluate the polarization of liver macrophages and the possible role of Wnt3a production by macrophages in hepatic progenitor cell response in the progression of pediatric non-alcoholic fatty liver disease. 32 children with biopsy-proven non-alcoholic fatty liver disease were included. 20 out of 32 patients were treated with docosahexaenoic acid for 18 months and biopsies at the baseline and after 18 months were included. Hepatic progenitor cell activation, macrophage subsets and Wnt/β-catenin pathway were evaluated by immunohistochemistry and immunofluorescence. Our results indicated that in pediatric non-alcoholic fatty liver disease, pro-inflammatory macrophages were the predominant subset. Macrophage polarization was correlated with Non-alcoholic fatty liver disease Activity Score, ductular reaction, and portal fibrosis; docosahexaenoic acid treatment determined a macrophage polarization towards an anti-inflammatory phenotype in correlation with the reduction of serum inflammatory cytokines, with increased macrophage apoptosis, and with the up-regulation of macrophage Wnt3a expression; macrophage Wnt3a expression was correlated with β-catenin phosphorylation in hepatic progenitor cells and signs of commitment towards hepatocyte fate. In conclusion, macrophage polarization seems to have a key role in the progression of pediatric non-alcoholic fatty liver disease; the modulation of macrophage polarization could drive hepatic progenitor cell response by Wnt3a production. PMID:27310371

  17. Impairment of Excitation-Contraction Coupling in Right Ventricular Hypertrophied Muscle with Fibrosis Induced by Pulmonary Artery Banding

    PubMed Central

    Urashima, Takashi; Shimura, Daisuke; Amemiya, Erika; Miyasaka, Genki; Yokota, Shunsuke; Fujimoto, Yoshitaka; Akaike, Toru; Inoue, Takahiro; Minamisawa, Susumu

    2017-01-01

    Interstitial myocardial fibrosis is one of the factors responsible for dysfunction of the heart. However, how interstitial fibrosis affects cardiac function and excitation-contraction coupling (E-C coupling) has not yet been clarified. We developed an animal model of right ventricular (RV) hypertrophy with fibrosis by pulmonary artery (PA) banding in rats. Two, four, and six weeks after the PA-banding operation, the tension and intracellular Ca2+ concentration of RV papillary muscles were simultaneously measured (n = 33). The PA-banding rats were clearly divided into two groups by the presence or absence of apparent interstitial fibrosis in the papillary muscles: F+ or F- group, respectively. The papillary muscle diameter and size of myocytes were almost identical between F+ and F-, although the RV free wall weight was heavier in F+ than in F-. F+ papillary muscles exhibited higher stiffness, lower active tension, and lower Ca2+ responsiveness compared with Sham and F- papillary muscles. In addition, we found that the time to peak Ca2+ had the highest correlation coefficient to percent of fibrosis among other parameters, such as RV weight and active tension of papillary muscles. The phosphorylation level of troponin I in F+ was significantly higher than that in Sham and F-, which supports the idea of lower Ca2+ responsiveness in F+. We also found that connexin 43 in F+ was sparse and disorganized in the intercalated disk area where interstitial fibrosis strongly developed. In the present study, the RV papillary muscles obtained from the PA-banding rats enabled us to directly investigate the relationship between fibrosis and cardiac dysfunction, the impairment of E-C coupling in particular. Our results suggest that interstitial fibrosis worsens cardiac function due to 1) the decrease in Ca2+ responsiveness and 2) the asynchronous activation of each cardiac myocyte in the fibrotic preparation due to sparse cell-to-cell communication. PMID:28068381

  18. Engineered antifouling microtopographies--correlating wettability with cell attachment.

    PubMed

    Carman, Michelle L; Estes, Thomas G; Feinberg, Adam W; Schumacher, James F; Wilkerson, Wade; Wilson, Leslie H; Callow, Maureen E; Callow, James A; Brennan, Anthony B

    2006-01-01

    Bioadhesion and surface wettability are influenced by microscale topography. In the present study, engineered pillars, ridges and biomimetic topography inspired by the skin of fast moving sharks (Sharklet AF) were replicated in polydimethylsiloxane elastomer. Sessile drop contact angle changes on the surfaces correlated well (R2 = 0.89) with Wenzel and Cassie and Baxter's relationships for wettability. Two separate biological responses, i.e. settlement of Ulva linza zoospores and alignment of porcine cardiovascular endothelial cells, were inversely proportional to the width (between 5 and 20 microm) of the engineered channels. Zoospore settlement was reduced by approximately 85% on the finer (ca 2 microm) and more complex Sharklet AF topographies. The response of both cell types suggests their responses are governed by the same underlying thermodynamic principles as wettability.

  19. Metabolic activity of sperm cells: correlation with sperm cell concentration, viability and motility in the rabbit.

    PubMed

    Sabés-Alsina, Maria; Planell, Núria; Gil, Sílvia; Tallo-Parra, Oriol; Maya-Soriano, Maria José; Taberner, Ester; Piles, Miriam; Sabés, Manel; Lopez-Bejar, Manel

    2016-10-01

    The resazurin reduction test (RRT) is a useful technique to assess the metabolic rate of sperm cells. RRT depends on the ability of metabolically active cells to reduce the non-fluorescent dye resazurin to the fluorescent resorufin. The aim of this study was to develop a vital fluorometric method to evaluate metabolic activity of rabbit sperm cells. Twenty-five rabbit males were included in the study. Viability and morphology, motility and metabolic activity were evaluated using an eosin-nigrosin staining, a computer-assisted semen analysis (CASA) and the RRT, respectively. Spearman rank correlation analysis was used to determine the correlation between RRT and semen parameters. After evaluation, a concentration of 10 × 106 sperm cells/ml was selected for further experiments with RRT. No significant correlation was found between the RRT results and the motility parameters. However, after RRT a significant positive correlation between relative fluorescence units and the percentage of alive spermatozoa (r = 0.62; P = 0.001) and a negative one with the percentage of sperm cells with acrosomic abnormalities (r = -0.45; P < 0.05) were detected. The vital assessment of metabolic rate of sperm cells by RRT could provide more information about semen quality than other routine semen analysis, correlating with sperm viability and acrosome status information.

  20. 22-Oxacalcitriol Prevents Progression of Peritoneal Fibrosis in a Mouse Model

    PubMed Central

    Hirose, Misaki; Nishino, Tomoya; Obata, Yoko; Nakazawa, Masayuki; Nakazawa, Yuka; Furusu, Akira; Abe, Katsushige; Miyazaki, Masanobu; Koji, Takehiko; Kohno, Shigeru

    2013-01-01

    ♦ Objective: Vitamin D plays an important role in calcium homeostasis and is used to treat secondary hyperparathyroidism among dialysis patients. The biologic activity of vitamin D and its analogs is mediated by vitamin D receptor (VDR), which is distributed widely throughout the body. Recent papers have revealed that low vitamin D levels are correlated with severe fibrosis in chronic diseases, including cystic fibrosis and hepatitis. The aim of the present study was to evaluate the protective effects of vitamin D against the progression of peritoneal fibrosis. ♦ Methods: Peritoneal fibrosis was induced by injection of chlorhexidine gluconate (CG) into the peritoneal cavity of mice every other day for 3 weeks. An analog of vitamin D, 22-oxacalcitriol (OCT), was administered subcutaneously daily from initiation of the CG injections. The peritoneal tissue was excised at 3 weeks. Changes in morphology were assessed by hematoxylin and eosin staining. Expression of VDR, alpha smooth muscle actin (as a marker of myofibroblasts), type III collagen, transforming growth factor β(TGF-β), phosphorylated Smad2/3, F4/80 (as a marker of macrophages), and monocyte chemoattractant protein-1 (MCP-1) was examined by immunohistochemistry. Southwestern histochemistry was used to detect activated nuclear factor κB (NF-κB). ♦ Results: In the CG-injected mice, immunohistochemical analysis revealed expression of VDR in mesothelial cells, myofibroblasts, and macrophages in the thickened submesothelial zone. Treatment with OCT significantly prevented peritoneal fibrosis and reduced the accumulation of type III collagen in CG-treated mice. Among the markers of fibrosis, the numbers of myofibroblasts, cells positive for TGF-β, and cells positive for phosphorylated Smad2/3 were significantly decreased in the OCT-treated group compared with the vehicle-treated group. Furthermore, OCT suppressed inflammatory mediators of fibrosis, as shown by the reduced numbers of activated NF

  1. Magnolol Attenuates Concanavalin A-induced Hepatic Fibrosis, Inhibits CD4(+) T Helper 17 (Th17) Cell Differentiation and Suppresses Hepatic Stellate Cell Activation: Blockade of Smad3/Smad4 Signalling.

    PubMed

    Zhang, Hongjun; Ju, Baoling; Zhang, Xiaoli; Zhu, Yanfei; Nie, Ying; Xu, Yuanhong; Lei, Qiuxia

    2016-12-29

    Magnolol is a pharmacological biphenolic compound extracted from Chinese herb Magnolia officinalis, which displays anti-inflammatory and antioxidant effects. This study was aimed at exploring the potential effect of magnolol on immune-related liver fibrosis. Herein, BALB/c mice were injected with concanavalin A (ConA, 8 mg/kg/week) up to 6 weeks to establish hepatic fibrosis, and magnolol (10, 20, 30 mg/kg/day) was given to these mice orally throughout the whole experiment. We found that magnolol preserved liver function and attenuated liver fibrotic injury in vivo. In response to ConA stimulation, the CD4(+) T cells preferred to polarizing towards CD4(+) T helper 17 (Th17) cells in liver. Magnolol was observed to inhibit Th17 cell differentiation in ConA-treated liver in addition to suppressing interleukin (IL)-17A generation. Hepatic stellate cells were activated in fibrotic liver as demonstrated by increased alpha smooth muscle actin (α-SMA) and desmin. More transforming growth factor (TGF)-β1 and activin A were secreted into the serum. Magnolol suppressed this abnormal HSC activation. Furthermore, the phosphorylation of Smad3 in its linker area (Thr179, Ser 204/208/213) was inhibited by magnolol. In vitro, the recombinant IL-17A plus TGF-β1 or activin A induced activation of human LX2 HSCs and promoted their collagen production. Smad3/Smad4 signalling pathway was activated in LX2 cells exposed to the fibrotic stimuli, as illustrated by the up-regulated phospho-Smad3 and the enhanced interaction between Smad3 and Smad4. These alterations were suppressed by magnolol. Collectively, our study reveals a novel antifibrotic effect of magnolol on Th17 cell-mediated fibrosis.

  2. Evaluation of liver fibrosis in patients with thalassemia: the important role of hyaluronic acid.

    PubMed

    Papastamataki, Maria; Delaporta, Polyxeni; Premetis, Evangelos; Kattamis, Antonios; Ladis, Vassilios; Papassotiriou, Ioannis

    2010-10-15

    Patients with transfusion-dependent thalassemia major often develop liver fibrosis due to liver iron overload and/or hepatitis virus C (HCV) infection. Hyaluronic acid (HA) plays a prominent role in the pathogenesis of liver fibrosis and the elevation of serum HA concentration is due to either increased synthesis by inflammatory cells and hepatic stellate cells or impaired degradation by sinusoidal endothelial cells (SECs) and thus is proposed as a non-invasive biomarker of liver fibrosis either by itself and/or included in the Hepascore formula. In this study we evaluated prospectively a screening of liver fibrosis in 201 adult patients aged 19-54 years with transfusion-dependent thalassemia major, based on HA measurements. 41/201 patients were HCV-RNA (+). HA was measured with a turbidimetric assay applied on a clinical chemistry analyzer. The Hepascore was computed from the results by using the model previously published. The main results of the study showed that: a) HA levels were increased in 110/201 (55%) thalassemia patients 85.0 ± 10.3 ng/ml, ranged from 15.0 to 1495.0 μg/l, compared to 20.8 ± 7.4 μg/l reference laboratory values, p<0.001, b) HA levels were significantly higher in HCV-RNA(+) compared to HCV-RNA(-) patients, 171.6 ± 202 vs 53.8 ± 35.5 μg/l, p<0.0001 c) no significant correlations were found between HA levels and/or Hepascore with ferritin and liver iron content (LIC) assessed with MRI (p>0.324 and p>0.270, respectively). Our findings indicate that hyaluronic acid measurements contribute to the assessment of liver fibrosis in patients with thalassemia and might be helpful for further evaluation of patients with liver biopsy if this is truly needed. Furthermore, liver fibrosis in thalassemia seems to be independent from liver siderosis.

  3. Aspirin and some other nonsteroidal anti-inflammatory drugs inhibit cystic fibrosis transmembrane conductance regulator protein gene expression in T-84 cells.

    PubMed

    Tondelier, D; Brouillard, F; Lipecka, J; Labarthe, R; Bali, M; Costa de Beauregard, M A; Torossi, T; Cougnon, M; Edelman, A; Baudouin-Legros, M

    1999-01-01

    Cystic fibrosis (CF) is caused by mutations in the CF gene, which encodes CF transmembrane conductance regulator protein (CFTR), a transmembrane protein that acts as a cAMP-regulated chloride channel The disease is characterized by inflammation but the relationship between inflammation, abnormal transepithelial ion transport, and the clinical manifestations of CF are uncertain. The present study was undertaken to determine whether three nonsteroidal anti-inflammatory drugs (NSAIDs) (aspirin, ibuprofen, and indomethacin) modulate CFTR gene expression in T-84 cells. Treatment with NSAIDs reduced CFTR transcripts, and decreased cAMP-stimulated anion fluxes, an index of CFTR function. However, the two phenomena occurred at different concentrations of both drugs. The results indicate that NSAIDs can regulate both CFTR gene expression and the function of CFTR-related chloride transport, and suggest that NSAIDs act via multiple transduction pathways.

  4. Stage scoring of liver fibrosis using Mueller matrix microscope

    NASA Astrophysics Data System (ADS)

    Zhou, Jialing; He, Honghui; Wang, Ye; Ma, Hui

    2016-10-01

    Liver fibrosis is a common pathological process of varied chronic liver diseases including alcoholic hepatitis, virus hepatitis, and so on. Accurate evaluation of liver fibrosis is necessary for effective therapy and a five-stage grading system was developed. Currently, experienced pathologists use stained liver biopsies to assess the degree of liver fibrosis. But it is difficult to obtain highly reproducible results because of huge discrepancy among different observers. Polarization imaging technique has the potential of scoring liver fibrosis since it is capable of probing the structural and optical properties of samples. Considering that the Mueller matrix measurement can provide comprehensive microstructural information of the tissues, in this paper, we apply the Mueller matrix microscope to human liver fibrosis slices in different fibrosis stages. We extract the valid regions and adopt the Mueller matrix polar decomposition (MMPD) and Mueller matrix transformation (MMT) parameters for quantitative analysis. We also use the Monte Carlo simulation to analyze the relationship between the microscopic Mueller matrix parameters and the characteristic structural changes during the fibrosis process. The experimental and Monte Carlo simulated results show good consistency. We get a positive correlation between the parameters and the stage of liver fibrosis. The results presented in this paper indicate that the Mueller matrix microscope can provide additional information for the detections and fibrosis scorings of liver tissues and has great potential in liver fibrosis diagnosis.

  5. Enhanced growth of endothelial precursor cells on PCG-matrix facilitates accelerated, fibrosis-free, wound healing: a diabetic mouse model.

    PubMed

    Kanitkar, Meghana; Jaiswal, Amit; Deshpande, Rucha; Bellare, Jayesh; Kale, Vaijayanti P

    2013-01-01

    Diabetes mellitus (DM)-induced endothelial progenitor cell (EPC) dysfunction causes impaired wound healing, which can be rescued by delivery of large numbers of 'normal' EPCs onto such wounds. The principal challenges herein are (a) the high number of EPCs required and (b) their sustained delivery onto the wounds. Most of the currently available scaffolds either serve as passive devices for cellular delivery or allow adherence and proliferation, but not both. This clearly indicates that matrices possessing both attributes are 'the need of the day' for efficient healing of diabetic wounds. Therefore, we developed a system that not only allows selective enrichment and expansion of EPCs, but also efficiently delivers them onto the wounds. Murine bone marrow-derived mononuclear cells (MNCs) were seeded onto a PolyCaprolactone-Gelatin (PCG) nano-fiber matrix that offers a combined advantage of strength, biocompatibility wettability; and cultured them in EGM2 to allow EPC growth. The efficacy of the PCG matrix in supporting the EPC growth and delivery was assessed by various in vitro parameters. Its efficacy in diabetic wound healing was assessed by a topical application of the PCG-EPCs onto diabetic wounds. The PCG matrix promoted a high-level attachment of EPCs and enhanced their growth, colony formation, and proliferation without compromising their viability as compared to Poly L-lactic acid (PLLA) and Vitronectin (VN), the matrix and non-matrix controls respectively. The PCG-matrix also allowed a sustained chemotactic migration of EPCs in vitro. The matrix-effected sustained delivery of EPCs onto the diabetic wounds resulted in an enhanced fibrosis-free wound healing as compared to the controls. Our data, thus, highlight the novel therapeutic potential of PCG-EPCs as a combined 'growth and delivery system' to achieve an accelerated fibrosis-free healing of dermal lesions, including diabetic wounds.

  6. Dioscin alleviates alcoholic liver fibrosis by attenuating hepatic stellate cell activation via the TLR4/MyD88/NF-κB signaling pathway

    PubMed Central

    Liu, Min; Xu, Youwei; Han, Xu; Yin, Lianhong; Xu, Lina; Qi, Yan; Zhao, Yanyan; Liu, Kexin; Peng, Jinyong

    2015-01-01

    The present work aimed to investigate the activities and underlying mechanisms of dioscin against alcoholic liver fibrosis (ALF). In vivo liver fibrosis in mice was induced by an alcoholic liquid diet, and in vitro studies were performed on activated HSC-T6 and LX2 cells treated with lipopolysaccharide. Our results showed that dioscin significantly attenuated hepatic stellate cells (HSCs) activation, improved collagen accumulation, and attenuated inflammation through down-regulating the levels of myeloid differentiation factor 88 (MyD88), nuclear factor κB (NF-κB), interleukin (IL)-1, IL-6 and tumour necrosis factor-α by decreasing Toll-like receptor (TLR)4 expression both in vivo and in vitro. TLR4 overexpression was also decreased by dioscin, leading to the markedly down-regulated levels of MyD88, NF-κB, transforming growth factor-β1 (TGF-β1), α-smooth muscle actin (α-SMA) and type I collagen (COL1A1) in cultured HSCs. Suppression of cellular MyD88 by ST2825 or abrogation of NF-κB by pyrrolidine dithiocarbamate eliminated the inhibitory effects of dioscin on the levels of TGF-β1, α-SMA and COL1A1. In a word, dioscin exhibited potent effects against ALF via altering TLR4/MyD88/NF-κB signaling pathway, which provided novel insights into the mechanisms of this compound as an antifibrogenic candidate for the treatment of ALF in the future. PMID:26655640

  7. [Cystic fibrosis in a woman aged seventy].

    PubMed

    Ras, Janneke E; van Velzen, Edwin; van Berkhout, Ferdinand Teding; van den Brand, Joop J G

    2010-01-01

    A seventy-year-old woman was admitted to hospital with a Staphylococcus aureus respiratory tract infection. She had a history of extensive bronchiectasis and allergic bronchopulmonary aspergillosis (ABPA). Cystic fibrosis (CF) was suspected and cystic fibrosis transmembrane conductance regulator (CFTR) gene analysis showed F508del and R117H-7T mutations. In these mutations there is residual activity in the chloride channel in the cell membrane coded by the CFTR gene. This results in a much milder disease pattern varying from no disease at all to isolated organ disease. This type of disease is known as non-classical cystic fibrosis. In our patient the diagnosis of cystic fibrosis was made exceptionally late in life.

  8. What Causes Idiopathic Pulmonary Fibrosis?

    MedlinePlus

    ... the NHLBI on Twitter. What Causes Idiopathic Pulmonary Fibrosis? Sometimes doctors can find out what is causing pulmonary fibrosis (lung scarring). For example, exposure to environmental pollutants ...

  9. MIrExpress: A Database for Gene Coexpression Correlation in Immune Cells Based on Mutual Information and Pearson Correlation.

    PubMed

    Wang, Luman; Mo, Qiaochu; Wang, Jianxin

    2015-01-01

    Most current gene coexpression databases support the analysis for linear correlation of gene pairs, but not nonlinear correlation of them, which hinders precisely evaluating the gene-gene coexpression strengths. Here, we report a new database, MIrExpress, which takes advantage of the information theory, as well as the Pearson linear correlation method, to measure the linear correlation, nonlinear correlation, and their hybrid of cell-specific gene coexpressions in immune cells. For a given gene pair or probe set pair input by web users, both mutual information (MI) and Pearson correlation coefficient (r) are calculated, and several corresponding values are reported to reflect their coexpression correlation nature, including MI and r values, their respective rank orderings, their rank comparison, and their hybrid correlation value. Furthermore, for a given gene, the top 10 most relevant genes to it are displayed with the MI, r, or their hybrid perspective, respectively. Currently, the database totally includes 16 human cell groups, involving 20,283 human genes. The expression data and the calculated correlation results from the database are interactively accessible on the web page and can be implemented for other related applications and researches.

  10. Role of the Ito cell in liver parenchymal fibrosis in rats fed alcohol and a high fat-low protein diet.

    PubMed Central

    French, S. W.; Miyamoto, K.; Wong, K.; Jui, L.; Briere, L.

    1988-01-01

    Eight pairs of young adult rats were pair-fed a high fat-low protein diet and ethanol or isocaloric glucose by permanent intragastric cannula for up to 6 months. Biopsies of the liver were taken monthly and the fibrosis was quantitated morphometrically using the sirius red polarization method of collagen visualization by light microscopy. Morphometric analysis of the sinusoids and scars were performed on electron micrographs made from the liver biopsies. An increase in the collagen in both the central and portal areas was found when the livers of the alcohol-fed rats were compared with controls. The predominant cell in the scars was the Ito cell. An increase in the percentage of the total Ito cell square area made up of rough endoplasmic reticulum (RER) was noted when the sinusoids of the liver of the ethanol-fed rats were compared with controls. No difference in the RER was found when the sinusoidal Ito cells were compared with the Ito cells located within the scars of the ethanol-fed rats. It was concluded that Ito cell "activation" by chronic ethanol feeding in the sinusoids of rats accurately predicts "activation" of the Ito cells within scars. The Ito cells are diffusely activated even though the scarring is localized. This implies that local factors as well as Ito cell activation are necessary for scar formation. In the case of alcoholic liver disease, scar formation may be initiated by centrilobular necrosis. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 PMID:3394803

  11. Epithelial-mesenchymal transition in liver fibrosis

    PubMed Central

    ZHAO, YA-LEI; ZHU, RONG-TAO; SUN, YU-LING

    2016-01-01

    Liver fibrosis is the result of a sustained wound healing response to sustained chronic liver injury, which includes viral, alcoholic and autoimmune hepatitis. Hepatic regeneration is the dominant outcome of liver damage. The outcomes of successful repair are the replacement of dead epithelial cells with healthy epithelial cells, and reconstruction of the normal hepatic structure and function. Prevention of the development of epithelial-mesenchymal transition (EMT) may control and even reverse liver fibrosis. EMT is a critical process for an epithelial cell to undergo a conversion to a mesenchymal phenotype, and is believed to be an inflammation-induced response, which may have a central role in liver fibrosis. The origin of fibrogenic cells in liver fibrosis remains controversial. Numerous studies have investigated the origin of all fibrogenic cells within the liver and the mechanism of the signaling pathways that lead to the activation of EMT programs during numerous chronic liver diseases. The present study aimed to summarize the evidence to explain the possible role of EMT in liver fibrosis. PMID:26998262

  12. [Regeneration and fibrosis of corneal tissues].

    PubMed

    Simirskiĭ, V N

    2014-01-01

    In this review, the features of the regeneration of corneal tissue and its disorders leading to the development of fibrosis are considered. The data on the presence of stem (clonogenic) cell pool in the corneal tissues (epithelium, endothelium, stroma) are given; these cells can serve as a source for regeneration of the tissues at injury or various diseases. The main steps of regeneration of corneal tissues and their disorders that lead to outstripping proliferation of myofibroblasts and secretion of extracellular matrix in the wound area and eventually cause the formation of connective tissue scar and corneal opacity are considered. Particular attention is given to the successes of translational medicine in the treatment of corneal tissue fibrosis. The methods of cell therapy aimed at the restoration of stem cell pool of corneal tissues are the most promising. Gene therapy provides more opportunities; one of its main objectives is the suppression of the myofibroblast proliferation responsible for the development of fibrosis.

  13. Omega-3 Polyunsaturated Fatty Acids Attenuate Fibroblast Activation and Kidney Fibrosis Involving MTORC2 Signaling Suppression.

    PubMed

    Zeng, Zhifeng; Yang, Haiyuan; Wang, Ying; Ren, Jiafa; Dai, Yifan; Dai, Chunsun

    2017-04-10

    Epidemiologic studies showed the correlation between the deficiency of omega-3 polyunsaturated fatty acids (n-3 PUFAs) and the progression of chronic kidney diseases (CKD), however, the role and mechanisms for n-3 PUFAs in protecting against kidney fibrosis remain obscure. In this study, NRK-49F cells, a rat kidney interstitial fibroblast cell line, were stimulated with TGFβ1. A Caenorhabditis elegans fat-1 transgenic mouse model in which n-3 PUFAs are endogenously produced from n-6 PUFAs owing to the expression of n-3 fatty acid desaturase were deployed. Docosahexaenoic acid (DHA), one member of n-3 PUFAs family, could suppress TGFβ1-induced fibroblast activation at a dose and time dependent manner. Additionally, DHA could largely inhibit TGFβ1-stimulated Akt but not S6 or Smad3 phosphorylation at a time dependent manner. To decipher the role for n-3 PUFAs in protecting against kidney fibrosis, fat-1 transgenic mice were operated with unilateral ureter obstruction (UUO). Compared to the wild types, fat-1 transgenics developed much less kidney fibrosis and inflammatory cell accumulation accompanied by less p-Akt (Ser473), p-Akt (Thr308), p-S6 and p-Smad3 in kidney tissues at day 7 after UUO. Thus, n-3 PUFAs can attenuate fibroblast activation and kidney fibrosis, which may be associated with the inhibition of mTORC2 signaling.

  14. Omega-3 Polyunsaturated Fatty Acids Attenuate Fibroblast Activation and Kidney Fibrosis Involving MTORC2 Signaling Suppression

    PubMed Central

    Zeng, Zhifeng; Yang, Haiyuan; Wang, Ying; Ren, Jiafa; Dai, Yifan; Dai, Chunsun

    2017-01-01

    Epidemiologic studies showed the correlation between the deficiency of omega-3 polyunsaturated fatty acids (n-3 PUFAs) and the progression of chronic kidney diseases (CKD), however, the role and mechanisms for n-3 PUFAs in protecting against kidney fibrosis remain obscure. In this study, NRK-49F cells, a rat kidney interstitial fibroblast cell line, were stimulated with TGFβ1. A Caenorhabditis elegans fat-1 transgenic mouse model in which n-3 PUFAs are endogenously produced from n-6 PUFAs owing to the expression of n-3 fatty acid desaturase were deployed. Docosahexaenoic acid (DHA), one member of n-3 PUFAs family, could suppress TGFβ1-induced fibroblast activation at a dose and time dependent manner. Additionally, DHA could largely inhibit TGFβ1-stimulated Akt but not S6 or Smad3 phosphorylation at a time dependent manner. To decipher the role for n-3 PUFAs in protecting against kidney fibrosis, fat-1 transgenic mice were operated with unilateral ureter obstruction (UUO). Compared to the wild types, fat-1 transgenics developed much less kidney fibrosis and inflammatory cell accumulation accompanied by less p-Akt (Ser473), p-Akt (Thr308), p-S6 and p-Smad3 in kidney tissues at day 7 after UUO. Thus, n-3 PUFAs can attenuate fibroblast activation and kidney fibrosis, which may be associated with the inhibition of mTORC2 signaling. PMID:28393852

  15. Quantitative Proteomic analysis on Activated Hepatic Stellate Cells reversion Reveal STAT1 as a key regulator between Liver Fibrosis and recovery

    PubMed Central

    Zhang, Hongyu; Chen, Fangyan; Fan, Xu; Lin, Cong; Hao, Yunwei; Wei, Handong; Lin, Weiran; Jiang, Ying; He, Fuchu

    2017-01-01

    Understanding the changes of activated HSCs reversion is an essential step toward clarifying the potential roles of HSCs in the treatment of liver fibrosis. In this study, we chose adipocyte differentiation mixture to induce LX-2 cells for 2 days in vitro as reversion phase, comparing with normal cultured LX-2 cells as activation phase. Mass spectrometric-based SILAC technology was adopted to study differentially expressed proteome of LX-2 cells between reversion and activation. Compared with activated HSCs, 273 proteins showed significant differences in reverted HSCs. The main pathway of up-regulated proteins associated with reversion of HSCs mainly related to oxidation-reduction and lipid metabolism, while the top pathway of down-regulated proteins was found in regulated cytoskeleton formation. Changes in the expression levels of selected proteins were verified by Western blotting analysis, especially STAT1, FLNA, LASP1, and NAMPT proteins. The distinct roles of STAT1 were further analyzed between activated and reverted of HSCs, it was found that STAT1 could affect cell proliferation of HSCs and could be viewed as a key regulator in the reversion of HSCs. Thus, the proteomic analysis could accelerate our understanding of the mechanisms of HSC reversion on cessation of fibrogenic stimuli and provide new targets for antifibrotic liver therapy. PMID:28322315

  16. Amine oxidase activity regulates the development of pulmonary fibrosis.

    PubMed

    Marttila-Ichihara, Fumiko; Elima, Kati; Auvinen, Kaisa; Veres, Tibor Z; Rantakari, Pia; Weston, Christopher; Miyasaka, Masayuki; Adams, David; Jalkanen, Sirpa; Salmi, Marko

    2017-03-01

    In pulmonary fibrosis, an inflammatory reaction and differentiation of myofibroblasts culminate in pathologic deposition of collagen. Amine oxidase copper containing-3 (AOC3) is a cell-surface expressed oxidase that regulates leukocyte extravasation. Here we analyzed the potential role of AOC3 using gene-modified and inhibitor-treated mice in a bleomycin-induced pulmonary fibrosis model. Inflammation and fibrosis of lungs were assessed by histologic, flow cytometric, and quantitative PCR analysis. AOC3-deficient mice showed a 30-50% reduction in fibrosis, collagen synthesis, numbers of myofibroblasts, and accumulation of CD4(+) lymphocytes, NK T cells, macrophages, and type 2 innate lymphoid cells compared with wild-type control mice. AOC3 knock-in mice, which express a catalytically inactive form of AOC3, were also protected from lung fibrosis. In wild-type mice, a small-molecule AOC3 inhibitor treatment reduced leukocyte infiltration, myofibroblast differentiation, and fibrotic injury both in prophylactic and early therapeutic settings by about 50% but was unable to reverse the established fibrosis. AOC3 was also induced in myofibroblasts in human idiopathic pulmonary fibrosis. Thus, the oxidase activity of AOC3 contributes to the development of lung fibrosis mainly by regulating the accumulation of pathogenic leukocyte subtypes, which drive the fibrotic response.-Marttila-Ichihara, F., Elima, K., Auvinen, K., Veres, T. Z., Rantakari, P., Weston, C., Miyasaka, M., Adams, D., Jalkanen, S., Salmi, M. Amine oxidase activity regulates the development of pulmonary fibrosis.

  17. Matrilysin (MMP-7) is a major matrix metalloproteinase upregulated in biliary atresia-associated liver fibrosis.

    PubMed

    Huang, Chao-Cheng; Chuang, Jiin-Haur; Chou, Ming-Huei; Wu, Chia-Lin; Chen, Ching-Mei; Wang, Chih-Chi; Chen, Yaw-Sen; Chen, Chao-Long; Tai, Ming-Hong

    2005-07-01

    Matrix metalloproteinases (MMPs) are the proteases responsible for tissue remodeling during liver fibrosis caused by various disorders including biliary atresia. However, information regarding the relative contribution of these proteases to liver fibrosis is still limited. We studied matrix metalloproteinase-2 (MMP-2), -7, -9 and -13 mRNA expressions in the liver tissue of early-stage biliary atresia at the time of Kasai's procedure, late-stage biliary atresia at the time of liver transplantation with advanced fibrosis and nondiseased control without liver fibrosis. The results of real-time quantitative reverse transcriptase-PCR analysis revealed that only MMP-2 and -7 expressions were significantly different between groups. MMP-2 was significantly higher in Liver Transplantation group than both in Control (P=0.010) and in Kasai's Procedure (P=0.001) groups, whereas the difference of MMP-2 expression between Control and Kasai's Procedure was not significant. However, the relative expression level of MMP-7 was sequentially elevated when comparing Control, Kasai's Procedure and Liver Transplantation groups, and there was significant (P=0.019) difference when comparing Control and Liver Transplantation groups. Moreover, the fold difference in MMP-7 mRNA was much higher than that in MMP-2 mRNA between groups. The expressions of MMP-7 were further confirmed by agarose gel electrophoresis and Western blotting. Immunohistochemical analysis revealed a significant positive correlation of the scores of MMP-7 immunostaining with the stages of liver fibrosis. In situ hybridization demonstrated that the bile ductular epithelial cells, Kupffer cells and hepatocytes were the major producers of matrix metalloproteinase-7 in the liver. Our results imply that MMP-7 is a major MMP associated with the tissue remodeling during the progression of liver fibrosis in biliary atresia.

  18. Adherence of Candida albicans and Candida parapsilosis to epithelial cells correlates with fungal cell surface carbohydrates.

    PubMed

    Lima-Neto, Reginaldo G; Beltrão, Eduardo I C; Oliveira, Patrícia C; Neves, Rejane P

    2011-01-01

    Many studies have described the adherence of Candida albicans to epithelial cells but little is known about Candida parapsilosis adhesion and its role in host cell surface recognition. This study was designed to evaluate the correlation between the adherence of 20 C. albicans and 12 C. parapsilosis strains to human buccal epithelial cells and the expression of fungal cell surface carbohydrates using lectin histochemistry. Adherence assays were carried out by incubating epithelial cells in yeast suspensions (10(7) cells ml(-1) ) and peroxidase conjugated lectins (Con A, WGA, UEA I and PNA at 25 μg ml(-1) ) were used for lectin histochemistry. The results showed that adherence was overall greater for C. albicans than for C. parapsilosis (P < 0.01) and that the individual strain differences correlated with a high content of cell surface α-l-fucose residues as indicated by the UEA I staining pattern. Based on the saccharide specificity of the lectins used, these results suggest that l-fucose residues on cell surface glycoconjugates may represent recognition molecules for interactions between the yeast strain studied and the host (r = 0.6985, P = 0.0045). In addition, our results indicated the presence of α-d-glucose/α-d-mannose, N-acetyl-D-glucosamine/N-acetylneuraminic acid and D-galactose/N-acetyl-D-galactosamine in fungal cell wall.

  19. Disruption of Interleukin-1β Autocrine Signaling Rescues Complex I Activity and Improves ROS Levels in Immortalized Epithelial Cells with Impaired Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Function

    PubMed Central

    Clauzure, Mariángeles; Valdivieso, Angel G.; Massip Copiz, María M.; Schulman, Gustavo; Teiber, María Luz; Santa-Coloma, Tomás A.

    2014-01-01

    Patients with cystic fibrosis (CF) have elevated concentration of cytokines in sputum and a general inflammatory condition. In addition, CF cells in culture produce diverse cytokines in excess, including IL-1β. We have previously shown that IL-1β, at low doses (∼30 pM), can stimulate the expression of CFTR in T84 colon carcinoma cells, through NF-κB signaling. However, at higher doses (>2.5 ng/ml, ∼150 pM), IL-1β inhibit CFTR mRNA expression. On the other hand, by using differential display, we found two genes with reduced expression in CF cells, corresponding to the mitochondrial proteins CISD1 and MTND4. The last is a key subunit for the activity of mitochondrial Complex I (mCx-I); accordingly, we later found a reduced mCx-I activity in CF cells. Here we found that IB3-1 cells (CF cells), cultured in serum-free media, secrete 323±5 pg/ml of IL-1β in 24 h vs 127±3 pg/ml for S9 cells (CFTR-corrected IB3-1 cells). Externally added IL-1β (5 ng/ml) reduces the mCx-I activity and increases the mitochondrial (MitoSOX probe) and cellular (DCFH-DA probe) ROS levels of S9 (CFTR-corrected IB3-1 CF cells) or Caco-2/pRSctrl cells (shRNA control cells) to values comparable to those of IB3-1 or Caco-2/pRS26 cells (shRNA specific for CFTR). Treatments of IB3-1 or Caco-2/pRS26 cells with either IL-1β blocking antibody, IL-1 receptor antagonist, IKK inhibitor III (NF-κB pathway) or SB203580 (p38 MAPK pathway), restored the mCx-I activity. In addition, in IB3-1 or Caco-2/pRS26 cells, IL-1β blocking antibody, IKK inhibitor III or SB203580 reduced the mitochondrial ROS levels by ∼50% and the cellular ROS levels near to basal values. The AP-1 inhibitors U0126 (MEK1/2) or SP600125 (JNK1/2/3 inhibitor) had no effects. The results suggest that in these cells IL-1β, through an autocrine effect, acts as a bridge connecting the CFTR with the mCx-I activity and the ROS levels. PMID:24901709

  20. T-Cell surface CCR5 density is not correlated with hepatitis severity in hepatitis C virus/HIV-coinfected individuals: implications for the therapeutic use of CCR5 antagonists.

    PubMed

    Vincent, Thierry; Portales, Pierre; Baillat, Vincent; de Boever, Corinne Merle; Le Moing, Vincent; Vidal, Michèle; Ducos, Jacques; Clot, Jacques; Reynes, Jacques; Corbeau, Pierre

    2005-03-01

    CCR5 antagonists represent promising anti-HIV agents. Yet, if the CCR5 chemokine receptor plays a positive role in hepatitis C virus (HCV) infection, CCR5 antagonists might be contraindicated in HCV/HIV-coinfected subjects. Therefore, we tested the hypothesis that the level of T-cell surface CCR5 expression, which might determine the intensity of HCV-specific T-cell recruitment into the liver, and thereby the efficiency of the anti-HCV response, could determine HCV disease evolution. For this purpose, we compared CCR5 density, measured by quantitative flow cytometry at the surface of nonactivated (human leukocyte antigen-D-related [HLA-DR]-) T cells of 51 HCV/HIV patients, with HCV load, serum aminotransferase levels, and liver histology (inflammatory activity, fibrosis, and rate of fibrosis progression). DR-CD4+ T-cell surface CCR5 density, which correlated with DR-CD8+ T-cell surface CCR5 density and was stable over time in HCV/HIV-coinfected individuals, did not correlate with any of the biologic parameters of HCV infection analyzed and was not linked to the capacity to clear the virus. In conclusion, we failed to demonstrate any impact of interindividual variability in T-cell surface CCR5 density on HCV infection, which would have argued against the use of CCR5 antagonists in HIV/HCV-coinfected subjects.

  1. Toward surface quantification of liver fibrosis progression

    NASA Astrophysics Data System (ADS)

    He, Yuting; Kang, Chiang Huen; Xu, Shuoyu; Tuo, Xiaoye; Trasti, Scott; Tai, Dean C. S.; Raja, Anju Mythreyi; Peng, Qiwen; So, Peter T. C.; Rajapakse, Jagath C.; Welsch, Roy; Yu, Hanry

    2010-09-01

    Monitoring liver fibrosis progression by liver biopsy is important for certain treatment decisions, but repeated biopsy is invasive. We envision redefinition or elimination of liver biopsy with surface scanning of the liver with minimally invasive optical methods. This would be possible only if the information contained on or near liver surfaces accurately reflects the liver fibrosis progression in the liver interior. In our study, we acquired the second-harmonic generation and two-photon excitation fluorescence microscopy images of liver tissues from bile duct-ligated rat model of liver fibrosis. We extracted morphology-based features, such as total collagen, collagen in bile duct areas, bile duct proliferation, and areas occupied by remnant hepatocytes, and defined the capsule and subcapsular regions on the liver surface based on image analysis of features. We discovered a strong correlation between the liver fibrosis progression on the anterior surface and interior in both liver lobes, where biopsy is typically obtained. The posterior surface exhibits less correlation with the rest of the liver. Therefore, scanning the anterior liver surface would obtain similar information to that obtained from biopsy for monitoring liver fibrosis progression.

  2. Inhibition of the SphK1/S1P signaling pathway by melatonin in mice with liver fibrosis and human hepatic stellate cells.

    PubMed

    González-Fernández, Bárbara; Sánchez, Diana I; Crespo, Irene; San-Miguel, Beatriz; Álvarez, Marcelino; Tuñón, María J; González-Gallego, Javier

    2016-11-01

    The sphingosine kinase 1/sphingosine 1-phosphate (SphK1/S1P) system is involved in different pathological processes, including fibrogenesis. Melatonin abrogates activation of hepatic stellate cells (HSCs) and attenuates different profibrogenic pathways in animal models of fibrosis, but it is unknown if protection associates with its inhibitory effect on the SphK1/S1P axis. Mice in treatment groups received carbon tetrachloride (CCl4 ) 5 μL g(-1) body wt i.p. twice a week for 4 or 6 weeks. Melatonin was given at 5 or 10 mg kg(-1)  day(-1) i.p, beginning 2 weeks after the start of CCl4 administration. At both 4 and 6 weeks following CCl4 treatment, liver mRNA levels, protein concentration and immunohistochemical labelling for SphK1 increased significantly. S1P production, and expression of S1P receptor (S1PR)1, S1PR3 and acid sphingomyelinase (ASMase) were significantly elevated. However, there was a decreased expression of S1PR2 and S1P lyase (S1PL). Melatonin attenuated liver fibrosis, as shown by a significant inhibition of the expression of α-smooth muscle actin (α-SMA), transforming growth factor (TGF)-β and collagen (Col) Ι. Furthermore, melatonin inhibited S1P production, lowered expression of SphK1, S1PR1, SP1R3, and ASMase, and increased expression of S1PL. Melatonin induced a reversal of activated human HSCs cell line LX2, as evidenced by a reduction in α-SMA, TGF-β, and Col I expression. Melatonin-treated cells also exhibited an inhibition of the SphK1/S1P axis. Antifibrogenic effect of SphK1 inhibition was confirmed by treatment of LX2 cells with PF543. Abrogation of the lipid signaling pathway by the indole reveals novel molecular pathways that may account for the protective effect of melatonin in liver fibrogenesis. © 2016 BioFactors, 2016.

  3. CXCR4 pos circulating progenitor cells coexpressing monocytic and endothelial markers correlating with fibrotic clinical features are present in the peripheral blood of patients affected by systemic sclerosis.

    PubMed

    Campioni, Diana; Lo Monaco, Andrea; Lanza, Francesco; Moretti, Sabrina; Ferrari, Luisa; Fotinidi, Maria; La Corte, Renato; Cuneo, Antonio; Trotta, Francesco

    2008-08-01

    There is still controversy regarding the role of circulating endothelial and progenitor cells (CECs/CEPs) in the pathogenesis of systemic sclerosis (SSc). Using a sequential Boolean gating strategy based on a 4-color flow cytometric protocol, an increased number of CD31(pos)/CD184(pos)(CXCR4)/CD34(pos)/CD45(pos) and CD31(pos)/CD117(pos) (c-kit-R) /CD34(pos)/ CD45(pos) hematopoietic circulating progenitor cells (HCPCs) was detected in SSc patients compared with healthy subjects. In SSc, no circulating mature and progenitor endothelial cells were observed, while an enhanced generation of erythroid progenitor cells was found to be correlated with the presence of CD117+ HCPCs. The presence of freshly detected CXCR4posHCPC was correlated either to the in vitro cultured spindle-shaped endothelial like cells (SELC) with an endo/myelomonocytic profile or to SDF-1 and VEGF serum level. These data are related to more fibrotic clinical features of the disease, thus supporting a possible role of these cells in fibrosis.

  4. Caffeine inhibits TGFβ activation in epithelial cells, interrupts fibroblast responses to TGFβ, and reduces established fibrosis in ex vivo precision-cut lung slices.

    PubMed

    Tatler, Amanda L; Barnes, Josephine; Habgood, Anthony; Goodwin, Amanda; McAnulty, Robin J; Jenkins, Gisli

    2016-06-01

    Caffeine is a commonly used food additive found naturally in many products. In addition to potently stimulating the central nervous system caffeine is able to affect various systems within the body including the cardiovascular and respiratory systems. Importantly, caffeine is used clinically to treat apnoea and bronchopulmonary dysplasia in premature babies. Recently, caffeine has been shown to exhibit antifibrotic effects in the liver in part through reducing collagen expression and deposition, and reducing expression of the profibrotic cytokine TGFβ. The potential antifibrotic effects of caffeine in the lung have not previously been investigated. Using a combined in vitro and ex vivo approach we have demonstrated that caffeine can act as an antifibrotic agent in the lung by acting on two distinct cell types, namely epithelial cells and fibroblasts. Caffeine inhibited TGFβ activation by lung epithelial cells in a concentration-dependent manner but had no effect on TGFβ activation in fibroblasts. Importantly, however, caffeine abrogated profibrotic responses to TGFβ in lung fibroblasts. It inhibited basal expression of the α-smooth muscle actin gene and reduced TGFβ-induced increases in profibrotic genes. Finally, caffeine reduced established bleomycin-induced fibrosis after 5 days treatment in an ex vivo precision-cut lung slice model. Together, these findings suggest that there is merit in further investigating the potential use of caffeine, or its analogues, as antifibrotic agents in the lung.

  5. Modulation of the Expression of the Proinflammatory IL-8 Gene in Cystic Fibrosis Cells by Extracts Deriving from Olive Mill Waste Water

    PubMed Central

    Lampronti, Ilaria; Borgatti, Monica; Vertuani, Silvia; Manfredini, Stefano; Gambari, Roberto

    2013-01-01

    A persistent recruitment of neutrophils in the bronchi of cystic fibrosis (CF) patients contributes to aggravate the airway tissue damage, suggesting the importance of modulating the expression of chemokines, including IL-8 during the management of the CF patients. Polyphenols rich extracts derived from waste water from olive mill, obtained by a molecular imprinting approach, have been investigated in order to discover compounds able to reduce IL-8 expression in human bronchial epithelial cells (IB3-1 cells), derived from a CF patient with a ΔF508/W1282X mutant genotype and stimulated with TNF-alpha. Initially, electrophoretic mobility shift assays (EMSAs) were performed to determine whether the different active principles were able to inhibit the binding between transcription factor (TF) NF-kappaB and DNA consensus sequences. Among different representative active principles present in the extract, three compounds were selected, apigenin, oleuropein, and cyanidin chloride, which displayed remarkable activity in inhibiting NF-kappaB/DNA complexes. Utilizing TNF-alpha-treated IB3-1 cells as experimental model system, we demonstrated that apigenin and cyanidin chloride are able to modulate the expression of the NF-kappaB-regulated IL-8 gene, while oleuropein showed no effect in regulating the expression of the gene IL-8. PMID:23935691

  6. Overexpression of the cystic fibrosis transmembrane conductance regulator in NIH 3T3 cells lowers membrane potential and intracellular pH and confers a multidrug resistance phenotype.

    PubMed Central

    Wei, L Y; Stutts, M J; Hoffman, M M; Roepe, P D

    1995-01-01

    Because of the similarities between the cystic fibrosis transmembrane conductance regulator (CFTR) and multidrug resistance (MDR) proteins, recent observations of decreased plasma membrane electrical potential (delta psi) in cells overexpressing either MDR protein or the CFTR, and the effects of delta psi on pa