Process Of Fibrogenesis Research Articles

The role of cholelithiasis and cholecystectomy in the prognosis of non-alcoholic fatty liver disease: Recommendations for patient management

There is a close connection between cholelithiasis (GSD) and non-alcoholic fatty liver disease (NAFLD). It is based on common risk factors, insulin resistance, disorders of carbohydrate and lipid metabolism, hepato-enteric circulation (HEC) of bile acids and the state of intestinal microflora. Chole-cystectomy (CE) is currently considered as an independent risk factor for the development and progres-sion of NAFLD and metabolic disorders in patients with cholelithiasis.That is why a patient suffering from cholelithiasis and NAFLD needs an individual approach before cholecystectomy. Patients with cholelithiasis and NAFLD are recommended to undergo a dynamic monitoring after cholecystectomy. The monitoring includes a control of general condition, biochemical parameters of the liver, lipid and carbohydrate metabolism, liver fibroelastography parameters. The treatment for this category of patients is aimed at eliminating risk factors, strict adherence to diet, phys-ical activity regimen, in order to correct obesity, dyslipidemia, hyperglycemia, the use of drugs that im-prove the condition of the hepatocytes, lipid and carbohydrate metabolism, the metabolic function of hepatocytes and inhibiting the processes of fibrogenesis in liver. At the same time, the common medi-cines of this category of patients are ursodeoxycholic acid, glycyrrhizic acid, phospholipids, antioxi-dants, and drugs to cure bacterial overgrowth syndrome in the intestines.

P0409 Clinical and serological characteristic of Crohn’s Disease patients with gastrointestinal strictures

Abstract Background Processes of inflammation and fibrogenesis in Crohn’s Disease (CD) contribute to development of strictures and subsequent complications, constituting a significant medical issue. The primary objective of the study was to identify clinical parameters associated with the occurrence of gastrointestinal stenosis, as well as to determine the clinical utility of promising biomarkers of fibrosis in a cohort of patients with CD. Methods A single-center cohort study was performed on 78 adults diagnosed with different phenotypes of CD. The study cohort included 36 subjects (46.2%) with gastrointestinal strictures identified by endoscopic or radiological methods, while the control group comprised 42 individuals (53.8%) without stenosis. The assessment of clinical factors, like gender, age of disease onset, disease duration, comorbidities, disease location, extra-intestinal symptoms was undertaken to explore potential associations with the presence of gastrointestinal strictures. Serum levels of candidate biomarkers of fibrosis, including granulocyte-macrophage colony stimulating factor antibodies (GM-CSF Ab), hepatocyte growth factor activator (HGFA), fibronectin isoform ED-A and cartilage oligomeric matrix protein (COMP) were measured using ELISA test. Results The overall cohort consisted of 47.44% females (n=37) and 52.56% males (n=41). The median age of disease onset was similar between the stenosis group (25.00 years, IQR: 19.00–37.00) and the non-stenosis group (26.00 years, IQR: 21.00–33.00). A proportion of 44.44% of the stenosis group required surgical or endoscopic treatment due to strictures. The most common sites of stenosis comprised the large intestine (52.78%) and the ileocecal valve region (50%), ranking ileocecal valve involvement as a significant predictor for stricture formation, with OR=3.65 (95% CI: 1.11–13.43, p=0.039). Fibronectin ED-A, with a cut-off of ≤14.52 ng/mL, was significantly associated with strictures occurrence, with OR=0.96 (95% CI: 0.92–1.00, p=0.036). Some trends in GM-CSF Ab levels were observed in patients with ileocecal involvement (median 1.75 ng/mL) compared to those without (median 1.37 ng/mL), nearing statistical significance (p=0.061). Conclusion The performed study provided some significant data about stricturing course of CD, with ileocecal valve involvement as a marker of stricture formation among CD patients. Evaluated serum biomarkers, especially fibronectin isoform ED-A, seem to be a promising diagnostic tool, however obtained data are inconsistent and demand further trials.

Serum biomarkers in the metabolic dysfunction-associated steatotic liver fibrosis diagnosis in children

Background. The COVID-19 epidemic and the war in Ukraine have led to a significant increase in the number of children suffering from metabolic dysfunction-associated steatotic liver disease (MASLD). One of the unresolved problems associated with MASLD is the identification of individuals at risk of rapid disease progression and development of irreversible liver changes. The search for alternative noninvasive markers suitable for the early detection of liver fibrosis in children remains extremely relevant. The aim of the study was to determine the diagnostic value of serum fibrosis markers and their relationship with sonographic and body composition parameters in children with MASLD. Materials and methods. The case-control study included 80 children aged 6 to 17 years (mean of (12.15 ± 2.51) years). The presence of steatosis and liver fibrosis was determined by transient elastography (Fibro­Scan® 502 touch F60156, Echosens, France). All subjects underwent anthropometric studies to determine body mass index. If it was within one-two Z-score, overweight was diagnosed. If the body mass index exceeded two Z-score, obesity was diagnosed. According to transient elastography and body mass index, all children were divided into four groups: group I — 27 children with MASLD and fibrosis ≥ F1, group II — 35 children with MASLD without fibrosis, group III — 18 obese or overweight children without MASLD and without fibrosis. The control group IV consisted of 14 children with normal weight without MASLD and without fibrosis. The groups had no significant differences in age and gender distribution. The study of body composition was performed by bioimpedance analysis using a TANITA MC-780MA analyzer (manufactured by Maeno-cho, Itabashi-ku, Tokyo, Japan). Quantitative determination of the serum concentration of vascular endothelial growth factor (VEGF) was performed by enzyme-linked immunosorbent assay (ELISA) using test systems from Wuhan Fine Biotech Co., Ltd (China) according to the manufacturer’s recommendations. The level of serum cytokeratin 18 (CK-18) was evaluated with IDL Biotech AB kits (Sweden) for ELISA. Serum content of transforming growth factor beta 1 (TGF-β1) was studied using an ELISA test system from IBL International (Germany). Fibrogenesis processes were evaluated by the serum content of free hydroxyproline (HPf), protein-bound hydroxyproline (HPp/b) and glycosaminoglycans (GAG). Results. The study revealed a significant increase in the level of CK-18 and TGF-β1 in children with MASLD-associated liver fibrosis. In children with liver fibrosis, an increase in the ratio of HPf/HPp/b and the level of GAG in the blood serum was observed compared to patients with MASLD without fibrosis and with overweight and obese children. The threshold value of CK-18 for liver fibrosis diagnosis was 90.3 U/l (sensitivity 81.3 %, specificity 76.9 %, AUC 0.843, p < 0.001). The sensitivity of the threshold value of serum TGF-β1 (96.8 pg/mL) in children with MASLD was 80.0 %, specificity 65.7 %, AUC 0.787 (p < 0.001). Threshold value of serum GAG (4.24 mmol/L) demonstrated a sensitivity of 70.6 % and a specificity of 69.6 %, AUC 0.743 (p < 0.01). CK-18, TGF-β1, GAG shown a positive correlation with liver stiffness and elasticity, body composition of MASLD children and had high levels of diagnostic accuracy, which allows them to be used in children when screening for MASLD-associated liver fibrosis. Conclusions. Children with liver fibrosis are characterized by elevated serum levels of CK-18, VEGF, TGF-β1, HPp/b and GAG. The threshold values of CK-18 (more than 90.3 U/l), TGF-β1 (above 96.8 pg/mL) and GAG (more than 4.24 mmol/l) have high sensitivity and specificity, which allows them to be used for the diagnosis of liver fibrosis in children with MASLD.

An L-type calcium channel blocker nimodipine exerts anti-fibrotic effects by attenuating TGF-β1 induced calcium response in an in vitro model of thyroid eye disease

BackgroundThyroid eye disease (TED) is a vision-threatening autoimmune disorder. Orbital tissue fibrosis leading to intractable complications remains a troublesome issue in TED management. Exploration of novel therapeutic targets and agents to ameliorate tissue fibrosis is crucial for TED. Recent work suggests that Ca2+ signaling participates in tissue fibrosis. However, whether an alteration of Ca2+ signaling has a role in fibrogenesis during TED remains unclear. In this study, we aimed to investigate the role of Ca2+ signaling in the fibrogenesis process during TED and the potential therapeutic effects of a highly selective inhibitor of the L-type calcium channel (LTCC), nimodipine, through a TGF-β1 induced in vitro TED model.MethodsPrimary culture of orbital fibroblasts (OFs) were established from orbital adipose connective tissues of patients with TED and healthy control donors. Real-time quantitative polymerase chain reaction (RT-qPCR) and RNA sequencing were used to assess the genes expression associated with LTCC in OFs. Flow cytometry, RT-qPCR, 5-ethynyl-2′-deoxyuridine (EdU) proliferation assay, wound healing assay and Western blot (WB) were used to assess the intracellular Ca2+ response on TGF-β1 stimulation, and to evaluate the potential therapeutic effects of nimodipine in the TGF-β1 induced in vitro TED model. The roles of Ca2+/calmodulin-dependent protein kinase II (CaMKII) and signal transducer and activator of transcription 1 (STAT1) in fibrogenesis during TED were determined by immunohistochemistry, WB, flow cytometry and co-immunoprecipitation assay. Selective inhibitors were used to explore the downstream signaling pathways.ResultsLTCC inhibitor nimodipine blocked the TGF-β1 induced intracellular Ca2+ response and further reduced the expression of alpha-smooth muscle actin (α-SMA), collagen type I alpha 1 (Col1A1) and collagen type I alpha 2 (Col1A2) in OFs. Besides, nimodipine inhibited cell proliferation and migration of OFs. Moreover, our results provided evidence that activation of the CaMKII/STAT1 signaling pathway was involved in fibrogenesis during TED, and nimodipine inhibited the pro-fibrotic functions of OFs by down-regulating the CaMKII/STAT1 signaling pathway.ConclusionsTGF-β1 induces an LTCC-mediated Ca2+ response, followed by activation of CaMKII/STAT1 signaling pathway, which promotes the pro-fibrotic functions of OFs and participates in fibrogenesis during TED. Nimodipine exerts potent anti-fibrotic benefits in vitro by suppressing the CaMKII/STAT1 signaling pathway. Our work deepens our understanding of the fibrogenesis process during TED and provides potential therapeutic targets and alternative candidate for TED.

Unraveling the Exosome-miR-133a Axis: Targeting TGF-β Signaling via WJ-MSC-Derived Exosomes for Anti-Fibrotic Therapy in Liver Fibrosis.

One of the primary drivers of liver fibrosis is the excessive accumulation of ECM, primarily caused by the over-proliferation of HSCs. The activation of HSCs by TGF-β has a critical role in initiating fibrosis. Recent studies have suggested that miRNA-133a significantly regulates the fibrogenesis process, which its downregulation is associated with the fibrosis progression. Understanding the role of miRNA-133a provides potential therapeutic insights for targeting TGF-β signaling and mitigating liver fibrosis. We investigated whether exosomes could attenuate liver fibrosis by enhancing the antifibrotic effects of miR-133a. The LX-2 cell line was treated with TGF-β for 24 hours, followed by an additional 24 hours of treatment with exosomes. After this treatment period, we assessed the mRNA expression levels of α-SMA, collagen 1, and miR-133a, as well as the protein levels of p-Smad3. TGF-β exposure significantly increased the expression level of α-SMA and collagen 1 genes and elevated the levels of p-Smad3 protein. Additionally, it resulted in a significant downregulation of miR-133a compared to the control group. Exosome administration effectively reduced the TGF-β-induced upregulation of p-Smad3, α-SMA, and collagen 1 genes, but increased miR-133a expression levels. Our findings indicate that by partially mitigating the downregulation of miR-133a, exosomes can effectively inhibit the persistent activation of HSCs. Furthermore, in the context of in vitro liver fibrosis, exosomes can suppress the TGF-β/Smad3 pathway, reducing the accumulation of ECM.

Could Renin-angiotensin System Inhibitors Reduce Fibrosis in Rheumatic Heart Disease by Inhibiting Soluble Suppression of Tumorigenicity 2 (sST2)?: Systematic Review

Introduction: Fibrosis and valvular thickening are clinical signs of rheumatic heart disease (RHD), which can cause significant hemodynamic disturbances. Fibrogenesis is associated with immune processes, there is a sensitive biomarker of cardiac fibrosis called soluble suppression of tumorigenicity 2 (sST2). Soluble ST2 is affected by angiotensin II and cardiomyocyte stretch. It is hypothesised that inhibiting the angiotensin II pathway can reduce cardiac fibrosis through sST2 inhibition by renin-angiotensin system inhibitors, but clinical trials in the RHD population are limited. Thus, this study will systematically review other heart disease populations with a fibrogenesis process similar to RHD. Materials and methods: We conducted a data search on online databases: PubMed, ScienceDirect, and Google Scholar. Data screening and selection process using PRISMA flowchart. We assessed the quality of articles using the GRADE method. Results: 770 articles were obtained and 8 of them were relevant. The use of sacubitril/ valsartan compared to valsartan or enalapril was shown to significantly reduce sST2 levels at the end of the study (p< 0.05) and improved the risk of morbidity, mortality, hospitalisation, and echocardiographic outcomes. Objective parameters that showed sST2 reduction indirectly reduced cardiac fibrosis were decreased left ventricular end-diastolic volume index (p= 0.02), left ventricular end-systolic volume index (p= 0.045), left atrial volume index (p< 0.001), and mitral E/e′ ratio (p= 0.001). Conclusion: Although this study did not directly utilize the RHD patient population, therapy using renin-angiotensin system inhibitors may reduce the incidence of cardiac fibrosis through the sST2 inhibition pathway in conditions with similar pathogenesis.

Modern approaches to the creation of 2D and 3D cellular models for the study of lung fibrosis

Lung fibrosis is a consequence of many influences leading to damage to lung tissue and the development of subsequent inflammation. Fibrosis is an overgrowth of connective tissue, which can lead to a violation of the architectonics of the lungs and reduce their functionality up to a fatal outcome. At the same time, the mechanisms underlying fibrogenesis are currently insufficiently studied. In this regard, the task of studying them does not lose its relevance, and its solution requires the development of models of lung fibrosis that can reflect all the key processes of fibrogenesis.The in vivo model using animals has multiple undeniable advantages, but at the same time it has strict ethical limitations and does not reflect all the mechanisms of lung fibrosis inherent in the human body. At the same time, in vitro research, scientists can afford to use biomaterials not only of animals, but also of humans, and build cellular systems based on them — from 2D to 3D models. Modeling of pulmonary fibrosis is mainly based on the use of the main types of cells involved in the development of pulmonary fibrosis, such as myofibroblasts, fibroblasts, alveolocytes and others. Some models are also based on a specific fibrosis-associated extracellular matrix and further study of the interaction of cells with each other and with the matrix. It should be borne in mind that different models display individual nuances of the native processes of lung fibrogenesis, which requires the research community to use a wide range of models. Taking into account the multifactorial pathogenesis of pulmonary fibrosis, it is important to understand the totality of the processes taking place in order to obtain the completeness of the real picture, close to the picture in vivo, and therefore the multicomponence of models is important. This review focuses on the analysis of various models of lung fibrosis in vitro in two-dimensional and three-dimensional systems, shows approaches to their creation, key differences, main advantages and disadvantages of models, both particular and general.

Milk fat globule-epidermal growth factor 8 (MFGE8) prevents intestinal fibrosis

ObjectiveIntestinal fibrosis is considered an inevitable consequence of chronic IBD, leading to stricture formation and need for surgery. During the process of fibrogenesis, extracellular matrix (ECM) components critically regulate the...

Possible contributions of fibrogenesis to recurrent miscarriages – A transcriptome analysis

Recurrent miscarriages (RM) generally refer to two or more consecutive pregnancy losses. The risk of miscarriages grows with its frequency of occurrences, so as the future obstetric complications or longer-term health problems for patients. Most previous researches sought to discover the etiology of RM by making comparisons between patients with RM and fertile women. Our study collected decidua tissues from patients with RM and single miscarriage (SM) for transcriptome sequencing analysis and aimed at identifying vital factors contributing to additional miscarriages after previous miscarriage. Between the RM and SM group, a total of 122 differentially expressed genes (DEGs) were detected and pathways associated with cell adhesion and ECM remodeling were particularly enriched in the RM group, which indicated abnormally activated fibrogenesis process. Particularly, the enhancement of ITGB6, EGFLAM and COL3A1 in the RM group were validated by RT-qPCR. Our study discovered that fibrogenesis, which might be caused by intrauterine manipulation, could lead to recurrent miscarriages after a previous miscarriage. Therefore, we encourage higher attention to thorough prevention and prompt remedies towards fibrotic disorders related diseases.

Plasma levels of IL-6, IL-10, IL-18, TNFα under the conditions of tobacco intoxication and after treatment with aminophthalhydrazide

In response to damaging factors, a cascade of cytokines is released, triggering the processes of inflammation and fibrogenesis. The purpose of our study was to evaluate the levels of cytokines (IL-6, IL-10, IL-18, TNFα) in blood plasma under conditions of tobacco intoxication and with intramuscular administration of aminophthalhydrazide. The experiment was performed with 40 outbred white male rats. Intact group (n = 10) were in a priming chamber in absence of tobacco smoke. Experimental groups (1, 2, and 3) – n = 30, 10 animals in each group. The first and second groups of animals were exposed to tobacco smoke for one and two months. The third experimental group underwent intramuscular injections with sodium aminodihydrophthalazindione for 3, 7, 14 days. The content of IL-6, IL-10, IL-18, TNFα in blood plasma was determined by enzyme immunoassay technique. In 1st group, the TNFα level in blood plasma increased to 452.14±176.18 pg/mL (in intact animals, 15.23±3.13 pg/mL); in the 2nd group, this index decreased to 9.8; in group 3, these values showed a sharp increase (to 167.44±5.93 pg/mL). The level of IL-6 showed maximal values after 4 months of tobacco intoxication (group 3). The following IL-10 changes were observed: in the 1st group, its amounts increased by 1.2-fold; in the 2nd group a sharp decrease in the indindex was noted (13.7 times); in the 3rd group a decreased content of the cytokine was found (28.8 times) as compared with intact animals. After administration of aminodihydrophthalazinedione for 3 and 7 days, the content of TNFα showed sharp increas (14.8 times and 7.6 times, respectively); after 14 days of treatment, the level of TNFα decreased by 1.3 times compared to intact animals. We noted fluctuations in the levels of pro-inflammatory cytokines (IL-6 and IL-18). The level of IL- 10 in blood plasma had a statistically significant increase (5.3-fold at 3 days after drug administration); 8.2-fold 7 days after administration of the drug, and 23-fold after 14 days of treatment, as compared with the 3rd group. The plasma concentrations of TNFα, IL-6, IL-10, IL-18 showed sufficient changes under the conditions of tobacco intoxication. The immune response is characterized by increased levels of pro-inflammatory cytokines in blood plasma. Upon corrective therapy with sodium aminodihydrophthalazindione, an increase in the antiinflammatory IL-10 cytokine occurs, along with systemic decrease in the TNFα, IL-6, and IL-18 contents.

TGF-β1 stimulation and VDR-dependent activation modulate calcitriol action on skeletal muscle fibroblasts and Smad signalling-associated fibrogenesis

Fibroblasts play a pivotal role in fibrogenesis after skeletal muscle injury. Excess fibrous formation can disrupt contractile functions and delay functional recovery. Although vitamin D receptor (VDR) is expressed explicitly in regenerating muscle compared with uninjured muscle, how calcitriol [1α,25(OH)2D3] directly regulates skeletal muscle primary fibroblast proliferation, the transition to myofibroblasts, and Smad signalling-associated fibrogenesis is currently unknown. Herein, the effects of calcitriol on cultured skeletal muscle primary fibroblasts of male C57BL/6 mice (aged 1 month old) were investigated. The percentage of BrdU+ nuclei in primary fibroblasts was significantly decreased after calcitriol treatment; however, the antiproliferative effect of calcitriol was diminished after TGF-β1 stimulation to induce fibroblast to myofibroblast transition. This suppressive effect was associated with significantly decreased VDR expression in TGF-β1-treated cells. In addition, Vdr siRNA transfection abolished the effects of calcitriol on the suppression of α-SMA expression and Smad2/3 signalling in myofibroblasts, supporting that its antifibrogenic effect requires VDR activation. Compared with calcitriol, the antifibrotic agent suramin could inhibit fibroblast/myofibroblast proliferation and suppress the expression of TCF-4, which regulates fibrogenic determination. Collectively, these findings suggest that profibrotic stimulation and VDR-dependent activation could modulate the effects of calcitriol on skeletal muscle fibroblast proliferation and fibrogenesis processes. Therefore, TGF-β1 and VDR expression levels are crucial determinants for the antifibrogenic effect of calcitriol on skeletal muscle after injury.

RESULTS OF ANTIFIBROTIC THERAPY IN PATIENTS WITH CHRONIC HEPATITIS B+C

Modern antiviral therapy regimens for patients with chronic viral hepatitis aim to achieve either long-term suppression of pathogen replication (e.g., nucleoside analogs in chronic hepatitis B) or complete elimination of the virus (such as direct-acting antiviral drugs in chronic hepatitis C). However, antiviral agents do not have a significant impact on the complete restoration of biochemical processes or the prevention of further progression of morphological changes in the liver. These limitations emphasize the ongoing need for new therapeutic strategies that target the processes of fibrogenesis.
 The aim of the work is to assess the possibility of the effect of the drug "Bicyclol" on fibrotic changes in patients with chronic hepatitis B+C using a non-invasive scale of the rate of fibrosis.
 Materials and methods
 An analysis of the dynamics of 62 patients with chronic hepatitis B and C (HCV+C) was conducted. All patients received long-term antiviral therapy consisting of pegylated interferon for 48 weeks. In the main group (Group I), patients with chronic hepatitis B and C were prescribed the drug "Bicyclol" after completing antiviral treatment with interferon. The control group (Group II) followed the principles of proper nutrition and took traditional hepatoprotectors. Based on the identified correlations, a non-invasive scale was proposed to assess the individual risk of liver fibrosis progression.
 Research results
 Against the background of the use of an antiviral treatment regimen, the normalization of cytolysis indicators was observed in most patients with CHB+C, but in some patients it was short-lived. Treatment with the drug "Bicyclol" contributed to a further decrease in cytolysis indicators, in the vast majority of patients with CHB+C, the activity of ALT and AST reached the upper limit of the norm. In patients with CHB+C who received only basic treatment without an antifibrotic component, a tendency to maintain an elevated level of transaminases was observed.
 Conclusions.
 Thus, the use of the hepatoprotector "Bicyclol" for 12 months ensures a decrease in cytolysis in the liver, is accompanied by a decrease in the severity of fibrosis, and in some patients - its reverse development. The use of the proposed prognostic scale makes it possible to assess the need for early appointment of antifibrotic therapy.

CLINICAL AND LABORATORY FEATURES OF THE COURSE OF CHRONIC VIRAL HEPATITIS B AND C IN CHILDREN OF PODILSKY REGION: POSSIBILITY FOR MODERN MONITORING

Introduction. In modern hepatology, a special place is occupied by chronic viral hepatitis, including B and C, because today they occupy the leading positions due to their high prevalence, the possibility of severe complications, and also they could lead to death in both adults and children. Specialists should have a comprehensive understanding of the clinical course and laboratory diagnosis of viral hepatitis to facilitate timely diagnosis and effective management of this condition.
 The aim: to evaluate the clinical and laboratory features of the course of chronic viral hepatitis B and C in children by determining patterns between parametric indexes characterizing the processes of liver fibrosis.
 Materials and methods. In the process of scientific research, 49 children were examined, including 29 children with diagnosed chronic viral hepatitis B and C (group I) and 20 almost healthy children (group II). The diagnoses of CHB and CHC were confirmed by the PCR method and specific markers detected by ELISA. All examined patients underwent anamnesis collection, general clinical examination, laboratory examination, calculation of CDS, Lok, GUCI, APRI, FIB-4 and "Fibrotest" diagnostic indexes. Statistical data analysis was carried out with "R-Studio" and "Statistica 10.0" software, using the methods of descriptive statistics, correlation analysis. Significance was considered significant at p<0.05. The study was conducted in compliance with the principles of the Declaration of Helsinki.
 Results. It was established that the levels of ALT, AST, GGT, ALP, indirect bilirubin, international normalized ratio (INR) were significantly higher in the examined patients of the I group. The level of the CDS index was significantly higher in the examined patients of the I group (5,97±0,30), compared with the patients of the II group (4,35±0,28) (p<0,001). A similar trend was noted with the Lok index and FIB-4 (p<0,001). The GUCI and APRI indices were significantly higher in patients of the I group (1,02±0,22; 0,73±0,04), compared to the examined patients of the II group (0,27±0,02; 0,23±0,02) (p<0,01).
 Conclusions. Implementation of non-invasive indices into clinical practice for the management of children with chronic hepatitis B and C will allow for dynamic monitoring of liver fibrogenesis processes.

Gypenosides Synergistically Reduce the Extracellular Matrix of Hepatic Stellate Cells and Ameliorate Hepatic Fibrosis in Mice.

Liver fibrosis resulting from chronic liver damage is becoming one of the major threats to health worldwide. Active saponin constituents isolated from Gynostemma pentaphyllum were found to possess a protective effect in liver diseases. Here, we obtained a naturally abundant gypenoside, XLVI, and evaluated its liver protection activity in both animal and cellular models. The results showed that it ameliorated acute and chronic liver injuries and lightened the process of fibrogenesis in vivo. XLVI can inhibit TGF-β-induced activation of hepatic stellate cells and ECM deposition in vitro. The underlying mechanism study verified that it upregulated the protein expression of protein phosphatase 2C alpha and strengthened the vitality of the phosphatase together with a PP2Cα agonist gypenoside NPLC0393. These results shed new light on the molecular mechanisms and the potential therapeutic function of the traditional herb Gynostemma pentaphyllum in the treatment of liver fibrosis.

SPECT Imaging of Lysyl Oxidase-like 2 in a Model of Idiopathic Pulmonary Fibrosis.

Noninvasive imaging of idiopathic pulmonary fibrosis (IPF) remains a challenge. The aim of this study was to develop an antibody-based radiotracer targeting Lysyl Oxidase-like 2 (LOXL2), an enzyme involved in the fibrogenesis process, for SPECT/CT imaging of pulmonary fibrosis. The bifunctional chelator DOTAGA-PEG4-NH2 was chemoenzymatically conjugated to the murine antibody AB0023 using microbial transglutaminase, resulting in a degree of labeling (number of chelators per antibody) of 2.3. Biolayer interferometry confirmed that the binding affinity of DOTAGA-AB0023 to LOXL2 was preserved with a dissociation constant of 2.45 ± 0.04 nM. DOTAGA-AB0023 was then labeled with 111In and in vivo experiments were carried out in a mice model of progressive pulmonary fibrosis induced by intratracheal administration of bleomycin. [111In]In-DOTAGA-AB0023 was injected in three groups of mice (control, fibrotic, and treated with nintedanib). SPECT/CT images were recorded over 4 days p.i. and an ex vivo biodistribution study was performed by gamma counting. A significant accumulation of the tracer in the lungs of the fibrotic mice was observed at D18 post-bleomycin. Interestingly, the tracer uptake was found selectively upregulated in fibrotic lesions observed on CT scans. Images of mice that received the antifibrotic drug nintedanib from D8 up to D18 showed a decrease in [111In]In-DOTAGA-AB0023 lung uptake associated with a decrease in pulmonary fibrosis measured by CT scan. In conclusion, we report the first radioimmunotracer targeting the protein LOXL2 for nuclear imaging of IPF. The tracer showed promising results in a preclinical model of bleomycin-induced pulmonary fibrosis, with high lung uptake in fibrotic areas, and accounted for the antifibrotic activity of nintedanib.

PPAR-Gamma Orchestrates EMT, AGE, and Cellular Senescence Pathways in Colonic Epithelium and Restrains the Progression of IBDs.

Intestinal fibrosis, the most common complication of inflammatory bowel disease (IBD), is characterized by an uncontrolled deposition of extracellular matrix proteins leading to complications resolvable only with surgery. Transforming growth factor is the key player in the epithelial-mesenchymal transition (EMT) and fibrogenesis process, and some molecules modulating its activity, including peroxisome proliferator-activated receptor (PPAR)-γ and its agonists, exert a promising antifibrotic action. The purpose of this study is to evaluate the contribution of signaling other than EMT, such as the AGE/RAGE (advanced glycation end products/receptor of AGEs) and the senescence pathways, in the etiopathogenesis of IBD. We used human biopsies from control and IBD patients, and we used a mouse model of colitis induced by dextran-sodium-sulfate (DSS), without/with treatments with GED (PPAR-gamma-agonist), or 5-aminosalicylic acid (5-ASA), a reference drug for IBD treatment. In patients, we found an increase in EMT markers, AGE/RAGE, and senescence signaling activation compared to controls. Consistently, we found the overexpression of the same pathways in DSS-treated mice. Surprisingly, the GED reduced all the pro-fibrotic pathways, in some circumstances more efficiently than 5-ASA. Results suggest that IBD patients could benefit from a combined pharmacological treatment targeting simultaneously different pathways involved in pro-fibrotic signals. In this scenario, PPAR-gamma activation could be a suitable strategy to alleviate the signs and symptoms of IBD and also its progression.

Combination of Niclosamide and Pirfenidone Alleviates Pulmonary Fibrosis by Inhibiting Oxidative Stress and MAPK/Nf-κB and STATs Regulated Genes.

The pathogenesis of pulmonary fibrosis (PF) is extremely complex and involves numerous intersecting pathways. The successful management of PF may require combining multiple agents. There is a growing body of evidence that suggests the potential benefits of niclosamide (NCL), an FDA-approved anthelminthic drug, in targeting different fibrogenesis molecules. This study aimed at investigating the anti-fibrotic potential of NCL alone and in combination with pirfenidone (PRF), an approved drug for PF, in a bleomycin (BLM) induced PF experimental model. PF was induced in rats by intratracheal BLM administration. The effect of NCL and PRF individually and in combination on different histological and biochemical parameters of fibrosis was investigated. Results revealed that NCL and PRF individually and in combination alleviated the histopathological changes, extracellular matrix deposition and myofibroblastic activation induced by BLM. NCL and PRF either individually or in combination inhibited the oxidative stress and subsequent pathways. They modulated the process of fibrogenesis by inhibiting MAPK/NF-κB and downstream cytokines. They inhibited STATs and downstream survival-related genes including BCL-2, VEGF, HIF-α and IL-6. Combining both drugs showed significant improvement in the tested markers in comparison to the monotherapy. NCL, therefore, has a potential synergistic effect with PRF in reducing the severity of PF.

Characteristics of <i>m. Psoas minor</i> and <i>m. Sacrocaudalis (coccygeus)</i> dorsalis lateralis in simultaneous modeling of lateral interbodial spinnylodesis and posterior sacro-iliac joint arthodesis

BACKGROUND: Simultaneous surgical interventions on the spine with the use of high-tech instruments and minimally invasive access techniques allow to eliminate several problems all at once, to activate patients at an early date and to reduce the number of complications. AIM: To evaluate morphological changes to evaluate morphological changes in the m. Psoas minor and m. Sacrocaudalis dorsalis lateralis during simultaneous modeling of lateral interbody fusion and posterior sacroiliac joint arthrodesis MATERIALS AND METHODS: Experiments were carried out on 14 outbred dogs; 3 animals formed a control group. The animals underwent consecutive lateral interbody fusion of the lumbar spine and posterior arthrodesis of the sacroiliac joint. The lumbar spine and sacroiliac joint were stabilized with external fixation device. Paraffin sections of muscles were stained with hematoxylin-eosin, according to Van Gieson, and Masson. Biochemical analysis of blood serum was performed during the experiment. RESULTS: The morphological study of the muscles revealed pathohistological features such as an increase in the variety of myosymplast diameters, loss of their profiles polygonality, massive fibers fatty degeneration, endo- and perimysial fibrosis, sclerotization of vessel membranes, obliteration of their lumens. At the end of the experiment, the degree of the small lumbar muscle fibrosis was 161% and of the sacrocaudal dorsal lateral muscle fibrosis was 240% of the control parameters (p 0.05); the rate of the muscle fatty infiltration was 339 and 310% of the normal value, respectively. The sacroiliac-caudal dorsal lateral muscle underwent more marked changes, especially in the early stages of the experiment. A significant increase in the enzymes activity, skeletal muscle damage markers was detected on the 14th day after surgery. CONCLUSION: Simultaneous surgical interventions on the spine should minimize mechanical effects on the paravertebral muscles and use techniques to stimulate their function in the postoperative period, which will reduce the processes of fibrogenesis and fat involution as well as provide an overall shorter rehabilitation period for the target patients.

DOP32 Crohn’s disease stricture matrisome analysis reveals the anti-fibrotic activity of milk-fat globule-epidermal growth factor 8 (MFGE8)

Abstract Background Intestinal fibrosis is considered an inevitable consequence of chronic inflammatory bowel disease (IBD), leading to stricture formation and need for surgery. During the process of fibrogenesis, extracellular matrix (ECM) components critically regulate the function of mesenchymal cells. We characterized the composition and function of the ECM in fibrostenosing Crohn’s disease (CD) and control tissues. Methods Decellularized full thickness intestinal tissue platforms were tested using three different protocols and ECM composition in different tissue phenotypes was explored by proteomics and validated by quantitative polymerase chain reaction (qPCR) and immunohistochemistry. Primary human intestinal myofibroblasts (HIMF) treated with Milk Fat Globule-Epidermal growth factor 8 (MFGE8) were evaluated regarding mechanism of their anti-fibrotic response, and the action of MFGE8 was tested in experimental intestinal fibrosis. Results We established and validated an optimal decellularization protocol for intestinal IBD tissues. Matrisome analysis revealed elevated MFGE8 expression in CD strictured tissue (CDs), which was confirmed at the mRNA and protein levels. Treatment with MFGE8 inhibited ECM production in normal control (NL) HIMF but not CDs HIMF. Next generation sequencing (NGS) uncovered functionally relevant integrin-mediated signaling pathways, and blockade of integrin αvβ5 and focal adhesion kinase rendered HIMF non-responsive to MFGE8. MFGE8 prevented and reversed experimental intestinal fibrosis in vitro and in vivo. Conclusion MFGE8 displays anti-fibrotic effects, and its administration may represent a future therapeutic approach for prevention of IBD-induced intestinal strictures.

РОЛЬ ЭНДОТЕЛИЯ В ФОРМИРОВАНИИ ФИБРОЗА ПЕЧЕНИ

Chronic liver disease is a worldwide health problem. A significant proportion of the manifestations and complications of chronic liver diseases is due to liver fibrosis and its subsequent transition to liver cirrhosis. The initial and the key link in the formation of fibrotic changes in the liver are liver sinusoidal endothelial cells (LSEC). In view of the unique structure and central position of the liver endothelial tissue in the pathogenesis of fibrosis, it seems relevant to present current data in the form of this review. LSECs are the only endothelial cells in the body that lack a basement membrane and contain transcellular pores called fenestrae. At the initial stages of the fibrogenesis process, LSECs change their phenotype: they lose fenestrae and develop a basement membrane, turning into a continuous endothelium. LSECs are involved in fibrosis through the secretion of angiocrine signals that act as paracrine factors that balance the liver’s response to injury towards fibrosis or regeneration. LSEC cells are very sensitive to the slightest changes in the microenvironment, with prolonged exposure they quickly change their phenotype, their numerous functions are disrupted, including vasodilator, anti-inflammatory, antithrombotic and antifibrotic, as well as the regulation of angiogenesis and regeneration, and prevention of HSC activation. Such changes in the phenotype are called endothelial dysfunction. Loss of fenestres (capillaryization) by LSEC cells is the initial event in liver fibrosis. It precedes HSC activation and promotes fibrosis and progression to cirrhosis. Fenestra maintains liver homeostasis, promotes efficient transport of lipoproteins, regulates liver regeneration and immune tolerance. LSEC fenestra are approximately 50-200 nm in diameter, and most of them are clustered into several dozen ultrastructures called sieve plates. Chronic liver injury leads to deep dedifferentiation of LSECs, which lose their vasoprotective properties and become vasoconstrictive, proinflammatory, and prothrombotic. Major molecular dysregulations seen in LSEC in chronic liver disease include fenestra loss and basement membrane development that interfere with the exchange of molecules such as lipoproteins and oxygen with hepatocytes, promoting steatosis and parenchymal apoptosis; reduction of NO by suppression of KLF2 and endothelial NO synthase (eNOS) activity, together with an increase in ROS-mediated NO uptake, leading to activation of hepatic stellate cells and deposition of extracellular matrix; increased production of vasoconstrictors (such as endothelin 1 or thromboxane A2) and pro-inflammatory cytokines, further exacerbating sinusoidal constriction. These pathological changes lead to sinusoidal vasoconstriction, microvascular dysfunction, fibrosis, and ultimately portal hypertension. Thus, LSECs play complex interrelated roles in maintaining liver homeostasis and are involved as factors in inflammation and fibrogenesis in liver diseases. Their unique position, phenotype, and function make them attractive candidates for organ-specific therapies, and it is likely that more therapies targeting these cells will be tested in the future as novel therapies to reduce liver damage and inflammation, and to prevent or reversal of fibrogenesis.