Deposition Of Fibrotic Tissue Research Articles

Autologous Minced Muscle Grafts Improve Muscle Strength in a Porcine Model of Volumetric Muscle Loss Injury.

The traumatic loss of muscle tissue, defined as volumetric muscle loss (VML) injury, has no definitive therapy. The purposes of this study were: (1) to develop a porcine model of VML and (2) to investigate autologous minced muscle grafts (1-mm pieces of muscle) as a potential therapeutic. Minced grafts were evaluated because they have promoted fiber regeneration and functional recovery in rat VML models and do not require US Food and Drug Administration approval for clinical use. In 5 female Yorkshire-cross pigs, ≈5 g (≈20%) of tissue was excised from the peroneous tertius muscle (≈3 × 3 × 1.5-cm defect) of each leg. The defect in one leg was treated with autologous minced grafts derived from the contralateral leg. Maximal isometric tetanic strength assessments of the dorsiflexor muscles (ie, the peroneous tertius muscle) were performed before and biweekly up to 12 weeks postinjury. VML injury resulted in a -43.5% ± 7.2% strength deficit 12 weeks postinjury in nonrepaired legs. Autologous minced muscle graft repair significantly improved strength over 12 weeks (32% strength increase 12 weeks postinjury vs. nonrepaired muscles with a remaining -27.8% ± 7.0% strength deficit; P < 0.001). Nonrepaired muscles developed extensive fibrosis and presented no evidence of muscle fiber regeneration within the defect area. Minced graft-treated muscles presented areas of putative de novo muscle fiber regeneration within the defect area, although extensive fibrotic tissue deposition was also present. Autologous minced muscle grafts partially restored neuromuscular strength in a novel porcine model of VML.

The Development of Macrophage-Mediated Cell Therapy to Improve Skeletal Muscle Function after Injury.

Skeletal muscle regeneration following acute injury is a multi-step process involving complex changes in tissue microenvironment. Macrophages (MPs) are one of the key cell types involved in orchestration and modulation of the repair process. Multiple studies highlight the essential role of MPs in the control of the myogenic program and inflammatory response during skeletal muscle regeneration. A variety of MP phenotypes have been identified and characterized in vitro as well as in vivo. As such, MPs hold great promise for cell-based therapies in the field of regenerative medicine. In this study we used bone-marrow derived in vitro LPS/IFN-y-induced M1 MPs to enhance functional muscle recovery after tourniquet-induced ischemia/reperfusion injury (TK-I/R). We detected a 15% improvement in specific tension and force normalized to mass after M1 (LPS/IFN-γ) MP transplantation 24 hours post-reperfusion. Interestingly, we found that M0 bone marrow-derived unpolarized MPs significantly impaired muscle function highlighting the complexity of temporally coordinated skeletal muscle regenerative program. Furthermore, we show that delivery of M1 (LPS/IFN-γ) MPs early in regeneration accelerates myofiber repair, decreases fibrotic tissue deposition and increases whole muscle IGF-I expression.

Identifying the inflammatory and fibrotic bowel stricture: MRI diffusion-weighted imaging in Crohn's disease

PurposeTo identify inflammatory and fibrotic mural stricture in Crohn's disease (CD) using MR diffusion-weighted imaging (DWI) and to compare DWI findings with those of enteroscope and histological markers. Method and materialsThirty-one patients with CD (18 males, 13 females; mean age, 38.9 years) were recruited in this approved retrospective study and an informed consent was obtained from each subject. All subjects underwent bowel MRI examination with conventional and DWI sequences at 3.0 T. Colonoscopy results were distributed within 24 h after examination. According to the endoscopic manifestations and pathological results, the patients were divided into two groups: inflammatory (21/31) and fibrotic (10/31). ResultsIn the group of inflammatory stricture, the mean ADC value of stricture bowel was 1.4 ± 0.23 × 10−3 mm2/s, whereas 0.8 ± 0.16 × 10−3 mm2/s in the group of fibrotic stricture. Inter-group independent sample t-test was performed. A statistically significant difference was observed (t = 7.403, P < 0.05). The area under receiver-operating characteristic curve was 0.981 (95% confidence interval, 0.943–1.000), with 1.11 × 10−3 mm2/s as the cutoff point. The sensitivity of low ADC values in detecting inflammatory bowels was 90.5%, and the specificity of high ADC values in excluding inflammatory bowels was 100%. ConclusionDecreased ADC values in inflammatory stricture bowel may be resulted from multiple factors, including an increase in cellularity, presence of edema, micro-abscesses and increased perfusion. Meanwhile, fibrotic tissue deposition was indicated to lead to restrictions in diffusion. Distinguishable ADCs were observed between inflammatory and fibrotic bowel stricture, where DWI sequence could contribute to the identification.

Simultaneous isolation of enriched myoblasts and fibroblasts for migration analysis within a novel co-culture assay.

Skeletal muscle injury elicits the activation of satellite cells and their migration to the wound area for subsequent terminal differentiation and tissue integration. However, interstitial fibroblasts recruited to the site of injury promote deposition of fibrotic tissue, which hampers myoblast-mediated muscle regeneration. Currently, analysis of myoblast migration in vitro can be accomplished using chemotactic, cell-exclusion, or wound healing assays. Yet, to investigate cell motility following skeletal muscle damage more accurately, migration assays need to better simulate the repair process. Here we present a protocol for the simultaneous isolation of myoblasts and fibroblasts from the same muscle tissue, ensuring the consistent generation of enriched, purified, and matched cell populations at a low passage number. We then describe a wound assay that uses a novel approach to the co-culture of myoblasts and fibroblasts to mimic the injured environment more closely than other established methods. Using this assay, we demonstrate that fibroblasts are able to increase myoblast migration significantly, validating our new in vitro method. As the observed effect on migration is most likely mediated by secreted factors, our assay could easily be extended to include antibody-based protein analysis of secreted factors in animal or human systems.

Macrophages promote osteoblastic differentiation in-vivo: implications in fracture repair and bone homeostasis.

Macrophages are activated in inflammation and during early phases of repair processes. Interestingly, they are also present in bone during development, but their function during this process is unclear. Here, we explore the function of macrophages in bone development, growth, and repair using transgenic mice to constitutively or conditionally deplete macrophages. Depletion of macrophages led to early skeletal growth retardation and progressive osteoporosis. By 3 months of age, macrophage-deficient mice displayed a 25% reduction in bone mineral density and a 70% reduction in the number of trabecular bone compared to control littermates. Despite depletion of macrophages, functional osteoclasts were still present in bones, lining trabecular bone and the endosteal surface of the cortical bone. Furthermore, ablation of macrophages led to a 60% reduction in the number of bone marrow mesenchymal progenitor cells and a decrease in the ability of these cells to differentiate to osteoblasts. When macrophages were depleted during fracture repair, bone union was impaired. Calluses from macrophage-deficient animals were smaller, and contained less bone and more fibrotic tissue deposition. Taken together, this shows that macrophages are crucial for maintaining bone homeostasis and promoting fracture repair by enhancing the differentiation of mesenchymal progenitors.

Erdheim-Chester disease: from palliative care to targeted treatment.

Erdheim–Chester disease (ECD) is a life-threatening multi-systemic non-Langerhans histiocytosis with cardiovascular complications as the leading cause of death. ECD affects the kidneys in up to 30% of cases, with fibrotic tissue deposition in the perirenal fat and renal hilum. Diagnosis is usually based on histological analysis of the pathologic tissue, which typically shows xanthogranulomatous infiltrates of foamy CD68+/CD1a- histiocytes surrounded by fibrosis. A consistent percentage of patients affected by ECD develop renal failure and hypertension as a consequence of renal artery stenosis and hydronephrosis. These conditions have been generally treated with the placement of stents and nephrostomies that frequently led to disappointing outcomes. Before the introduction of interferon-alpha (IFNα) treatment, the mortality rate was as high as 57% in the long term. Recent studies have granted new insights into the pathogenesis of ECD, which seems to bear a dual component of clonal and inflammatory disease. These advances led to use specific therapies targeting either the oncogenes (BRAFV600E) or the effectors of the immune response implicated in ECD (IL-1, TNFα). Drugs such as anakinra (recombinant human IL-1 receptor antagonist), infliximab (monoclonal antibody against TNFα) and vemurafenib (inhibitor of mutant BRAF) showed promising results in small single-centre series. Although larger trials will be needed to address the impact of these drugs on ECD prognosis and to select the most effective treatment, targeted therapies hold the premises to drastically change the outcome of this condition.

Notch signaling and new therapeutic options in liver disease

Notch signaling is a crucial determinant of cell fate decision during development and disease in several organs. Notch effects are strictly dependent on the cellular context in which it is activated. In the liver, Notch signaling is involved in biliary tree development and tubulogenesis. Recent advances have shed light on Notch as a critical player in liver regeneration and repair, as well as in liver metabolism and inflammation and cancer. Notch signaling is finely regulated at several levels. The complexity of the pathway provides several possible targets for development of therapeutic agents able to inhibit Notch. Recent reports have shown that persistent activation of Notch signaling is associated with liver malignancies, particularly hepatocellular with stem cell features and cholangiocarcinoma. These novel findings suggest that interfering with the aberrant activation of the Notch pathway may have therapeutic relevance. However, further studies are needed to clarify the mechanisms regulating physiologic and pathologic Notch activation in the adult liver, to better understand the mechanistic role(s) of Notch in liver diseases and to develop safe and specific therapeutic agents.

Mechanisims of asthma and allergic disease – 1092. Regulation of profibrotic cytokines release from human eosinophils by Th17 cytokines

Background Asthma is a chronic inflammatory disorder of the lung airways that is associated with airway remodeling and hyperresponsiveness. One of the most critical structural changes that affect airway functionality is fibrotic tissue deposition within the airway wall. Eosinophils have been proposed in different studies to contribute to the production of several mediators and cytokines, including the profibrotic cytokines, TGF-b and IL-11. In this study, we hypothesize that cytokines prevailing in asthmatic tissue such as Th1, Th2, and Th17 cytokines, may induce eosinophils to produce pro-fibrotic cytokines. Methods Eosinophils were isolated from peripheral blood of 6 mild asthmatics and 6 normal control subjects. Eosinophils were stimulated with Th1, Th2 and Th17 cytokines and production of pro-fibrotic cytokines, TGF-b and IL-11, were determined using Intra-cellular cytokine detection and FACS analysis, immunohistochemistry, as well as real time PCR. Results The level of basal expression of eosinophil TGF-b and IL-11 was significantly upregulated in asthmatic patients compared to healthy individuals. Stimulating eosinophils with Th1 and Th2 cytokines did not induce expression of eosinophils derived pro-fibrotic cytokines. However, stimulating eosinophils with IL-17 resulted in the enhancement of the expression TGF-b and IL-11 in asthmatic individuals. Conclusions

Abstract 19398: Diastolic Dysfunction and Deposition of Fibrosis in Pressure Overloaded Hearts is Dependent on Pentraxin3

On the basis of the underlying pathophysiology, it is possible to distinguish two major forms of heart failure (HF): HF owing to reduced left ventricular (LV) systolic function and HF owing to LV diastolic dysfunction. In both cases, chronic remodeling processes in the myocardium antedate overt HF by many years. Inflammation has a pivotal role in cardiac remodeling, and circulating markers of systemic inflammation are independently associated with risk of developing HF and with prognosis after onset. However, the pathophysiological role of many inflammatory biomarkers in HF is poorly understood. In this context, the relationship between circulating levels of pentraxin 3 (PTX3), a novel marker of inflammation, and the presence of diastolic dysfunction and/or overt HF has been emerging. PTX3 belongs to pentraxins, members of a superfamily of multimeric pattern-recognition proteins that can be short (such as CRP) or long (the prototype of this group being PTX3), with important roles at the interface of innate immune response, inflammation, and extracellular matrix remodeling. To dissect the pathophysiological role of PTX3 in the two forms of HF, we subjected mice with a genetic ablation for PTX3 to chronic pressure overload. Both echocardiographic analysis and PV loops revealed an early and sustained protection from LV diastolic dysfunction. As regard the process of cardiac remodeling to pressure overload, absence of PTX3 resulted in reduced recruitment of monocyte and deposition of fibrotic tissue. The systolic function was not affected by the absence of PTX3 and both WT and KO mice developed a comparable systolic dysfunction over time. In conclusion our data fully support the clinical association among PTX3 levels and the presence of diastolic dysfunction and candidate this molecular target as a novel therapeutic approach for HF owing to LV diastolic dysfunction.

Th17 cytokines regulate profibrotic cytokines release by human eosinophils

Rationale Asthma is a chronic inflammatory disorder of the lung airways that is associated with airway remodeling and hyperresponsiveness. One of the most critical structural changes that affect airway functionality is fibrotic tissue deposition within the airway wall. Eosinophils have been proposed in different studies to contribute to the production of several mediators and cytokines, including the profibrotic cytokines, TGF-b and IL-11. In this study, we hypothesize that cytokines prevailing in asthmatic tissue such as Th1, Th2, and Th17 cytokines, may induce eosinophils to produce pro-fibrotic cytokines. Methods Eosinophils were isolated from peripheral blood of 6 mild asthmatics and 6 normal control subjects. Eosinophils were stimulated with Th1, Th2 and Th17 cytokines and production of pro-fibrotic cytokines, TGF-b and IL-11, were determined using Intra-cellular cytokine detection and FACS analysis, immunohistochemistry, as well as real time PCR. Results The level of basal expression of eosinophil TGF-b and IL-11 was significantly upregulated in asthmatic patients compared to healthy individuals. Stimulating eosinophils with Th1 and Th2 cytokines did not induce expression of eosinophils derived pro-fibrotic cytokines. However, stimulating eosinophils with IL-17 resulted in the enhancement of the expression TGF-b and IL-11 in asthmatic individuals. Conclusions

Contribution of circulating biomarkers to unravel the role of extracellular matrix in hypertensive cardiac remodelling

Contribution of circulating biomarkers to unravel the role of extracellular matrix in hypertensive cardiac remodelling

Non-invasive biomarkers of liver fibrosis in liver transplant patients with hepatitis C: Can we avoid liver biopsies?

HCV-related end-stage liver disease is the main indication for liver transplantation (LT) in most centers. Unfortunately, recurrent HCV infection is the rule with most patients developing post-transplantation HCV-graft disease. In fact, the deposition of fibrotic tissue in the new liver is a process that takes place more rapidly than it does in the native liver [ [1] Samuel D. Forns X. Berenguer M. Trautwein C. Burroughs A. Rizzetto M. et al. Report of the monothematic EASL conference on liver transplantation for viral hepatitis (Paris, France, January 12–14, 2006). J Hepatol. 2006; 45: 127-143 Abstract Full Text Full Text PDF PubMed Scopus (142) Google Scholar ]. The liver biopsy is the gold standard to assess the severity and progression of liver damage, including the degree of necro-inflammation and stage of fibrosis, and to exclude additional pathologies. Several histological scores are used, including the Knodell index, and its modified version by Ishak and METAVIR, with no evidence of superiority for any of these scores [ [2] Dienstag J. The role of liver biopsy in chronic hepatitis C. Hepatology. 2002; 36: S152-S160 Crossref PubMed Google Scholar ]. In the last few years, a search for non-invasive assessment of liver fibrosis has emerged, as a result of the increased awareness of the limitations of liver biopsies [ 2 Dienstag J. The role of liver biopsy in chronic hepatitis C. Hepatology. 2002; 36: S152-S160 Crossref PubMed Google Scholar , 3 Poynard T. Morra R. Ingiliz P. Imbert-Bismut F. Thabut D. Messous D. et al. Biomarkers of liver fibrosis. Adv Clin Chem. 2008; 46: 131-160 Crossref PubMed Scopus (57) Google Scholar ]. Indeed, major complications and liver-biopsy mortality have a prevalence of 0.15 and 0.01%, respectively. These data though come from early studies performed in immune competent patients, and similar information in LT is mostly missing. Additional limitations include the cost, inter- and intra-variability, sampling error, and particularly the fact that a biopsy provides a static picture of fibrosis while this is a very dynamic process [ [4] Friedman S.L. Rockey D.C. Bissell D.M. Hepatic fibrosis 2006: report of the Third AASLD Single Topic Conference. Hepatology. 2007; 45: 242-249 Crossref PubMed Scopus (98) Google Scholar ]. In addition, as the efficacy of antiviral drugs increase, the impact of histologic findings on therapeutic decisions is also questioned.

Transforming Growth Factor-β (TGF-β) Type I Receptor/ALK5-dependent Activation of the GADD45β Gene Mediates the Induction of Biglycan Expression by TGF-β

We have recently shown that induction of biglycan (BGN) expression by transforming growth factor-beta1 (TGF-beta1) required sequential activation of both Smad and p38 mitogen-activated protein kinase signaling (Ungefroren, H., Lenschow, W., Chen, W.-B., and Kalthoff, H. (2003) J. Biol. Chem. 278, 11041-11049). Here, we have analyzed the receptors through which TGF-beta1 controls expression of BGN and GADD45beta, the latter of which is postulated to link early Smad signaling to delayed activation of p38. Ectopic expression of a dominant-negative mutant of the TGF-beta type II receptor in PANC-1 cells abrogated TGF-beta-induced BGN up-regulation. Similarly, inhibition of the TGF-beta type I receptor/ALK5 with either SB431542 or by enforced stable expression of a kinase-dead mutant greatly attenuated the TGF-beta effect on both BGN and GADD45beta expression in PANC-1 and MG-63 cells. The enhancing effect of ALK5 on TGF-beta-mediated GADD45beta and BGN expression and on GADD45beta promoter activity was also dependent on its ability to activate Smad signaling, because an ALK5 mutant defective in Smad activation (TbetaRImL45) but with an otherwise functional kinase domain failed to mediate these responses. The TGF-beta/ALK5 effect on p38 activation and BGN expression was mimicked by overexpression of GADD45beta alone (in the absence of TGF-beta stimulation) and suppressed upon antisense inhibition of GADD45beta expression. These results show that TGF-beta induces BGN expression through (the Smad-activating function of) ALK5 and GADD45beta and suggest that the sensitivity of MyD118 to activation by TGF-beta, which varies between tissues, ultimately determines the strength of the TGF-beta effect on BGN.