Reduced Collagen Accumulation Research Articles

P0040 Measurement of collagen concentration throughout the intestinal layers using Deep Learning: implications in Inflammatory Bowel Disease (IBD)

Abstract Background IBD, particularly Crohn’s disease (CD), is marked by recurrent episodes of inflammation, leading to fibrosis and collagen deposition throughout the intestinal layers.1,2 In this study, we aim to quantify collagen levels across the distinct layers of the gut and examine their correlation with clinical characteristics and disease outcomes. Methods A total of 190 IBD patients were enrolled, including 98 with Ulcerative Colitis (UC) and 92 with CD (60% male; median age: 42 years, IQR: 29–48; follow-up: 110 months, IQR: 39-181), plus 73 controls. Biopsies were collected, stained with Sirius Red, and digitized. A total of 612 biopsies were included. Intestinal layers in biopsy images from the derivation group were manually annotated to train a mask Region-based Convolution Neural Network (mask R-CNN), which uses ResNet-101 backbone. After the detection of intestinal layers by the trained model, K-means was employed at the pixel level, to segment and quantify Collagen Proportionate Area (CPA) regions in each layer. Follow-up biopsies (>6 months) were available for 85 patients. Multivariate Cox regression was used to assess the risk of adverse outcomes, including variables with p<0.05 from the univariate analysis. Results Submucosal collagen was significantly correlated with mucosal CPA (r=0.36; p<0.001) and CPA in the muscular layer (r=0.54; p<0.001), as well as with C-reactive protein (r=0.38; p=0.001) and erythrocyte sedimentation rate (r=0.53; p=0.005). In CD, submucosal CPA showed a positive association with disease activity as measured by the Crohn’s Disease Activity Index (CDAI) (r=0.31; p=0.005), endoscopic activity assessed by the Simple Endoscopic Score for CD (SES-CD) (r=0.32; p=0.004), and Nancy histologic activity score (r=0.25; p=0.046). In UC, the Full Mayo score showed a trend with submucosal CPA, but it did not reach significance (r=0.13; p=0.09). Patients requiring treatment with biologics had higher baseline submucosal CPA levels (mean difference: 12; 95%CI: -0.6 to 24.6; p=0.06). Patients treated with biologics at follow-up biopsies (19/85) experienced a significantly greater reduction in submucosal CPA (-21.1; 95% CI: -38.3 to -3.9) compared to those not receiving biologics (-1.7; 95% CI: -9.4 to 6.1; p = 0.039). Baseline submucosal CPA was independently associated with an increased risk of surgery in CD patients during follow-up (aHR=1.03; 95%CI: 1.002–1.06; p=0.048), along with B2/B3 disease behavior (p=0.003) and elevated CDAI (p=0.004) (Table I). Conclusion Submucosal collagen levels correlate with clinical, endoscopic, and histologic activity in CD, potentially serving as a prognostic marker for long-term outcomes. Emerging therapies may help reduce collagen accumulation and its associated complications.

Qizhu Rougan Granules suppress liver fibrosis by inhibiting the expression of the P2Y14 receptor on hepatic stellate cells.

Liver fibrosis is a globally prevalent chronic liver disease, often representing the advanced stage of various chronic liver conditions. Despite its widespread occurrence, there is currently no widely accepted or effective treatment for liver fibrosis. However, increasing evidence supports the efficacy of Traditional Chinese Medicine (TCM) in inhibiting the progression of fibrosis. In this study, we explored the effects and potential mechanisms of Qizhu-Ruogan-Granules (QZRG), a formulation from the Affiliated Hospital of the Chengdu University of TCM, on carbon tetrachloride (CCl4)-induced liver fibrosis in mice. A total of 40 male C57BL/6J mice were randomly divided into five groups (n = 8 per group), with liver fibrosis induced by injecting 10% CCl4 for 15weeks. From the 7th week onward, QZRG granules were administered orally to the treatment groups at low, medium, and high doses. To assess liver function, serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) were measured. Liver morphology and fibrosis were evaluated using hematoxylin-eosin (H&E) and Masson's trichrome staining, while gene and protein expression levels were analyzed through quantitative reverse transcription polymerase chain reaction (RT-PCR) and western blot techniques. The results showed that QZRG granules significantly reduced serum levels of AST, ALT, and ALP in CCl4-treated mice, alleviated liver damage, and reduced collagen accumulation. Furthermore, QZRG granules inhibited the expression of apoptosis-related proteins BAX, Caspase9, Caspase8, and Caspase3, while reducing P2Y14 expression in fibrotic liver tissues. Additionally, QZRG granules suppressed the proliferation of activated hepatic stellate cells. Our findings suggest that QZRG granules may exert anti-fibrotic effects by downregulating P2Y14 expression and effectively slowing the progression of liver fibrosis.

Engineered lipid nanoparticles enable therapeutic gene silencing of GTSE1 for the treatment of liver fibrosis

Liver fibrosis is characterized by abnormal accumulation of extracellular matrix proteins, disrupting normal liver function. Despite its significant health impact, effective treatments remain limited. Here, we present the development of engineered lipid nanoparticles (LNPs) for targeted RNA therapeutic delivery in the liver. We investigated the therapeutic potential of modulating the G2 and S-phase expressed 1 (GTSE1) protein for treating liver fibrosis. Through screening, we identified P138Y LNP as a potent candidate with superior delivery efficiency and lower toxicity. Using these engineered LNPs, we successfully delivered siGTSE1 to hepatocytes, significantly reducing collagen accumulation and restoring liver function in a fibrosis animal model. Additionally, GTSE1 downregulation altered miRNA expression and upregulated hepatocyte nuclear factor 4 alpha (HNF4α). These findings suggest that therapeutic gene silencing of GTSE1 is a promising strategy for treating liver fibrosis by regenerating liver phenotypes and functions.

Amino acid-crosslinked 4arm-PLGA Janus patch with anti-adhesive and anti-bacterial properties for hernia repair

Presently, the non-biodegradable polypropylene (PP) patches frequently used for hernia repair can cause fibrous tissue growth and adhesions. This study created a Janus Patch with anti-adhesion and antimicrobial properties to improve hernia repair while promoting tissue repair. The biologically active 4arm-PLGA-BLPD was initially synthesized through the modification of 4arm-PLGA with lysine, followed by the fabrication of a Janus patch using a layer-by-layer electrostatic spinning technique. This patch consisted of three layers: a repair layer composed of 4arm-PLGA-BLPD/PCL fiber membrane, a mechanical layer of 4arm-PLGA/PCL fiber membrane, and an antimicrobial layer of EMO-4arm-PLGA/PCL fiber membrane loaded with Emodin (EMO). The results showed that Janus patch exhibited notable tensile strength and elongation at break, enabling it to offer enhanced mechanical reinforcement for abdominal wall defects. In addition, it slowly releases lysine for repair and inhibits bacterial growth with EMO. In vivo experiments demonstrated that the patch effectively induced neovascularization, reduced collagen ac-cumulation, and stabilized the expression of relevant proteins through the up-regulation of MMP1 and MMP9. This facilitated successful repair of the abdominal wall defect model and prevented adhesions. In summary, the Janus patch offers both practical application and theoretical insight for hernia repair.

Antagonizing Activin A/p15INK4b Signaling as Therapeutic Strategy for Liver Disease.

Activin A is involved in the pathogenesis of human liver diseases, but its therapeutic targeting is not fully explored. Here, we tested the effect of novel, highly specific small-molecule-based activin A antagonists (NUCC-474/555) in improving liver regeneration following partial hepatectomy and halting fibrosis progression in models of chronic liver diseases (CLDs). Cell toxicity of antagonists was determined in rat hepatocytes and Huh-7 cells using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay. Hepatocytes and hepatic stellate cells (HSCs) were treated with activin A and NUCC-555 and analyzed by reverse transcription-polymerase chain reaction and immunohistochemistry. Partial hepatectomized Fisher (F)344 rats were treated with NUCC-555, and bromodeoxyuridine (BrdU) incorporation was determined at 18/24/36/120/240 h. NUCC-555 was administered into thioacetamide- or carbon tetrachloride-treated F344 rats or C57BL/6 mice, and the fibrosis progression was studied. NUCC-474 showed higher cytotoxicity in cultured hepatic cells; therefore, NUCC-555 was used in subsequent studies. Activin A-stimulated overexpression of cell cycle-/senescence-related genes (e.g., p15INK4b, DEC1, Glb1) was near-completely reversed by NUCC-555 in hepatocytes. Activin A-mediated HSC activation was blocked by NUCC-555. In partial hepatectomized rats, antagonizing activin A signaling resulted in a 1.9-fold and 2.3-fold increase in BrdU+ cells at 18 and 24 h, respectively. Administration of NUCC-555 in rats and mice with progressing fibrosis significantly reduced collagen accumulation (7.9-fold), HSC activation indicated by reduced alpha smooth muscle actin+ and vimentin+ cells, and serum aminotransferase activity. Our studies demonstrate that activin A antagonist NUCC-555 promotes liver regeneration and halts fibrosis progression in CLD models, suggesting that blocking activin A signaling may represent a new approach to treating people with CLD.

Abstract 5737: Enhancing cancer treatment Via a nanostructured chemoimmunotherapy gel facilitating interleukin-2 delivery and immunogenic cell death induction

Abstract Background: The landscape of immunotherapy, employing recombinant cytokines like interleukin-2 (IL-2), confronts challenges marked by its short half-life, systemic toxicity, and constrained tumor accumulation. These hurdles are particularly evident in the context of the immunosuppressive milieu characterizing pancreatic ductal adenocarcinoma (PDAC). Chemo-immuno-therapeutic synergies, notably those provoking immunogenic cell death (ICD), have demonstrated superior efficacy. This investigation delves into Pt-NHC as a type II ICD inducer to surmount immunosuppression and amplify antitumor immunity within a highly tumor-accumulated liposomal drug delivery system. Additionally, the co-administration of the angiotensin receptor blocker, losartan, mitigates desmoplasia in PDAC, further augmenting the efficacy of immunotherapy. Methods: To address the constraints of IL-2, a lipid-coated nanogel, IL2-Pt@Nanogel, was innovated, amalgamating silk fibroin-loaded IL-2 with Pt-NHC. The nanogel aimed to enhance IL-2 pharmacokinetics and optimize tumor targeting. Concurrently, losartan was employed to alleviate desmoplasia within the tumor microenvironment. Result: The chemoimmunotherapy nanogel, when coupled with losartan, showcased therapeutic potential in murine models of desmoplastic PDAC. Pt-NHC induced ER-localized reactive oxygen species (ROS) and DAMP release, reshaping the immunosuppressive microenvironment by repolarizing M2-type macrophages to M1 and diminishing the regulatory T-cell population. Co-delivery of IL-2 and Pt-NHC in the nanogel enhanced T-cell infiltration and activation, curbing tumor progression in primary and liver metastasized PDAC models. Importantly, the addition of losartan reduced collagen accumulation within tumors, resulting in heightened infiltration of effector T cells and more pronounced suppression of tumor growth in PDAC and metastasized liver. Conclusion: This study introduces an innovative nanogel delivery system (IL2-Pt@Nanogel) for chemoimmunotherapy, effectively addressing the challenges associated with IL-2 therapy. Leveraging the enhanced permeability and retention (EPR) effect, the nanogel improves IL-2 stability and tumor delivery. Pt-NHC incorporation induces ICD, reshaping the tumor microenvironment, and in synergy with losartan, elicits robust anticancer immunity. This strategy holds promise for clinical translation as a secure and efficient treatment for immunosuppressive and desmoplastic cancers, such as PDAC, presenting a distinctive therapeutic avenue. Citation Format: Hsuan Yu Mu, Yen-Nhi Ngoc Ta, Jing Rui Max Tham, Fu-Fei Hsu, Yu-Chieh Lin, Hsi-Chien Huang, Yun-Chieh Sung, Chih-I Huang, Ching-Ling Wu, Chao-Hung Chang, Sheng Yang, Tsung-Ying Lee, Jane Wang, Dan G. Duda, Yves Boucher, Jen-Huang Huang, Wee Han Ang, Yunching Chen. Enhancing cancer treatment Via a nanostructured chemoimmunotherapy gel facilitating interleukin-2 delivery and immunogenic cell death induction [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5737.

The role of sinusoidal endothelial cells and TIMP1 in the regulation of fibrosis in a novel human liver 3D NASH model.

The prevalence of NAFLD is rapidly increasing. NAFLD can progress to NASH, fibrosis, cirrhosis, and HCC, which will soon become the main causes of liver transplantation. To date, no effective drug for NASH has been approved by the Food and Drug Administration. This is partly due to the lack of reliable human in vitro models. Here, we present a novel human liver spheroid model that can be used to study the mechanisms underlying liver fibrosis formation and degradation. Such spheroids, which contain hepatocytes, stellate cells, KC, and LSECs, spontaneously develop fibrosis that is exacerbated by treatment with free fatty acids. Conditioned medium from activated LSECs caused similar activation of fibrosis in spheroids containing primary human hepatocyte and NPCs, indicating the action of soluble mediators from the LSECs. Spheroids containing LSECs treated with free fatty acids produced tissue inhibitor of metalloproteinases inhibitor 1, a matrix metalloproteinases inhibitor important for fibrosis progression. Tissue inhibitor of metalloproteinases inhibitor 1 knockdown using siRNA led to a reduction in collagen and procollagen accumulation, which could be partially rescued using a potent matrix metalloproteinases inhibitor. Interestingly, tissue inhibitor of metalloproteinases inhibitor 1 was found to be expressed at higher levels, specifically in a subtype of endothelial cells in the pericentral region of human fibrotic livers, than in control livers. Potential anti-NASH drugs and compounds were evaluated for their efficacy in reducing collagen accumulation, and we found differences in specificity between spheroids with and without LSECs. This new human NASH model may reveal novel mechanisms for the regulation of liver fibrosis and provide a more appropriate model for screening drugs against NASH.

The Upregulation of Leucine-Rich Repeat Containing 1 Expression Activates Hepatic Stellate Cells and Promotes Liver Fibrosis by Stabilizing Phosphorylated Smad2/3.

Liver fibrosis poses a significant global health risk due to its association with hepatocellular carcinoma (HCC) and the lack of effective treatments. Thus, the need to discover additional novel therapeutic targets to attenuate liver diseases is urgent. Leucine-rich repeat containing 1 (LRRC1) reportedly promotes HCC development. Previously, we found that LRRC1 was significantly upregulated in rat fibrotic liver according to the transcriptome sequencing data. Herein, in the current work, we aimed to explore the role of LRRC1 in liver fibrosis and the underlying mechanisms involved. LRRC1 expression was positively correlated with liver fibrosis severity and significantly elevated in both human and murine fibrotic liver tissues. LRRC1 knockdown or overexpression inhibited or enhanced the proliferation, migration, and expression of fibrogenic genes in the human hepatic stellate cell line LX-2. More importantly, LRRC1 inhibition in vivo significantly alleviated CCl4-induced liver fibrosis by reducing collagen accumulation and hepatic stellate cells' (HSCs) activation in mice. Mechanistically, LRRC1 promoted HSC activation and liver fibrogenesis by preventing the ubiquitin-mediated degradation of phosphorylated mothers against decapentaplegic homolog (Smad) 2/3 (p-Smad2/3), thereby activating the TGF-β1/Smad pathway. Collectively, these results clarify a novel role for LRRC1 as a regulator of liver fibrosis and indicate that LRRC1 is a promising target for antifibrotic therapies.

TC14012 enhances the anti-fibrosis effects of UC-MSCs on the liver by reducing collagen accumulation and ameliorating inflammation

BackgroundMesenchymal stem cells (MSCs) are attracting attention as a promising cell-based therapy for the treatment of liver fibrosis or cirrhosis. However, the strategies and potential mechanisms of MSCs therapy need further investigation. The CXCL12/CXCR4/CXCR7 chemokine axis is well known to regulate cell migration and is involved in the regulation of liver fibrosis. This study aims to treat MSCs with a CXCR7-specific agonist to evaluate its therapeutic effects on hepatic fibrosis and potential mechanisms.MethodsTC14012, a potent agonist of CXCR7, has been used to pretreat human umbilical cord-derived MSCs (UC-MSCs) and assess its effect on proliferation, apoptosis, migration, immunoregulation, and gene regulatory network. Then, CCl4-induced liver fibrosis mice models were used to evaluate the therapeutic effect and mechanism of TC14012-treated UC-MSCs for treating hepatic fibrosis.ResultsTC14012 increased CXCR7 expression in UC-MSCs. Notably, co-culture of liver sinusoidal endothelial cells (LSEC) with TC14012-pretreated UC-MSCs increased CXCR7 expression in LSEC. Additionally, TC14012 promoted cell migration and mediated the immunoregulation of UC-MSCs. Compared to UC-MSCs without TC14012 pretreatment, UC-MSCs treated with TC14012 ameliorated live fibrosis by restoring CXCR7 expression, reducing collagen fibril accumulation, inhibiting hepatic stellate cells activation, and attenuating the inflammatory response.ConclusionThis study suggests that TC14012 pretreatment can enhance the therapeutic effects of UC-MSCs on liver fibrosis, mainly by promoting the migration and immunoregulation of MSCs.

Preclinical Investigations on Anti-fibrotic Potential of Long-Term Oral Therapy of Sodium Astragalosidate in Animal Models of Cardiac and Renal Fibrosis.

In traditional Chinese medicine, Radix Astragali has played a vital role in treating progressive fibrotic diseases. One of its main active components, astragaloside IV, is a promising anti-fibrotic treatment despite its extremely low bioavailability. Our study aimed to optimize sodium astragalosidate (SA) by salt formation to improve solubility and oral absorption for anti-fibrotic therapy in vivo. Isoproterenol-induced myocardial fibrosis rat models and obese BKS-db mice presenting diabetic kidney fibrosis were used in this study. Daily oral administration of SA (20 mg/kg) for 14 days ameliorated cardiac fibrosis by reducing collagen accumulation and fibrosis-related inflammatory signals, including TNF-α, IL-1β, and IL-6. In db/db mice, SA (5,10, and 20 mg/kg per day for 8 weeks) dose-dependently alleviated lipid metabolism impairment and renal dysfunction when administered orally. Furthermore, Western blot and immunohistochemistry analyses demonstrated that SA treatment inhibited renal fibrosis by suppressing TGF-β1/Smads signaling. Taken together, our findings provide the oral-route medication availability of SA, which thus might offer a novel lead compound in preclinical trial-enabling studies for developing a long-term therapy to treat and prevent fibrosis.

Comparative Analysis of the Therapeutic Effects of MSCs From Umbilical Cord, Bone Marrow, and Adipose Tissue and Investigating the Impact of Oxidized RNA on Radiation-Induced Lung Injury.

Radiation-induced lung injury (RILI) is frequently observed in patients undergoing radiotherapy for thoracic malignancies, constituting a significant complication that hampers the effectiveness and utilization of tumor treatments. Ionizing radiation exerts both direct and indirect detrimental effects on cellular macromolecules, including DNA, RNA and proteins, but the impact of oxidized RNA in RILI remains inadequately explored. Mesenchymal stem cells (MSCs) can repair injured tissues, and the reparative potential and molecular mechanism of MSCs in treating RILI remains incompletely understood. This study aimed to investigate the therapeutic effects and mechanisms of action of three distinct sources of MSCs, including human umbilical cord mesenchymal stem cells (UCMSCs), bone marrow mesenchymal stem cells (BMSCs), and adipose-derived stem cells (ADSCs), in thoracically irradiated mice. Comparative analysis revealed that all three types of MSCs exhibited the ability to mitigate radiation-induced inflammatory infiltration, alveolar hemorrhage, and alveolar wall thickening in the lung tissue of the mice. MSCs also attenuated RILI by decreasing inflammatory factors, upregulating anti-inflammatory factor expression, and reducing collagen accumulation. Immunohistochemical results showed that all three MSCs reduced radiation-induced cell apoptosis and promoted the regeneration of lung tissue cells. The analysis of malondialdehyde (MDA) and 8-hydroyguanosine (8-OHG) content indicated that MSCs possess reparative properties against radiation-induced oxidative damage in lung tissue. The study provides evidence that UCMSCs are a more appropriate therapeutic option for RILI compared to BMSCs and ADSCs. Additionally, MSCs effectively reduce the accumulation of oxidized RNA in RILI, thereby, presenting a unique avenue for investigating the underlying mechanism of MSC-based treatment for RILI.

Acne Scar Management: Minoxidil as a Promising Approach or a Mirage?

Atrophic and hypertrophic scars can result from various conditions, such as acne, trauma, and surgery. Minoxidil, a medication used for the treatment of severe hypertension and hair loss, has been explored as a potential treatment for scars. This review aims to evaluate the current evidence regarding the role of minoxidil in the treatment of scars. Previously published reviews have primarily focused on the use of minoxidil in hair loss and have only briefly mentioned its potential use for scars. However, minoxidil may have a beneficial effect as an antifibrotic agent. Several studies have reported reduced collagen accumulation and fibrosis after treatment with minoxidil. The proposed mechanism of action is inhibition of the production of lysyl hydroxylases (LHs), which modify and cross-link proteins by converting lysine to hydroxylysine, making collagen more resistant to degradation. Minoxidil, as an LH inhibitor, has been shown to potentially benefit wound healing and regeneration in vitro by inhibiting the proliferation and migration of fibroblasts. To date, direct studies of the efficacy of minoxidil in treating acne scars have not been conducted; however, its inhibitory effects on fibroblast function and antifibrotic outcomes in some in vivo studies suggest that such use may be considered.

Discovery of a chalcone derivative as an anti-fibrotic agent targeting transforming growth factor-β1 signaling: Potential therapy of renal fibrosis

As a final common pathway of renal injuries, renal fibrosis leads to chronic kidney disease (CKD). Currently, there is no safe and effective therapy to prevent the progression of renal fibrosis to CKD. Inhibition of transforming growth factor-β1 (TGF-β1) pathway is proposed as one of the most promising approaches for anti-renal fibrosis therapies. This study aimed to identify novel anti-fibrotic agents using the TGF-β1-induced fibrosis in renal proximal tubule epithelial cells (RPTEC) and characterize their mechanism of action as well as in vivo efficacy. By screening 362 natural product-based compounds for their ability to reduce collagen accumulation assessed by picro-sirius red (PSR) staining in RPTEC cells, a chalcone derivative AD-021 was identified as an anti-fibrotic agent with IC50 of 14.93 μM. AD-021 suppressed TGF-β1-induced collagen production, expression of pro-fibrotic proteins (fibronectin and α-smooth muscle actin (αSMA)), and Smad-dependent and Smad-independent signaling pathways via suppression of TGF-β receptor II (TGFβRII) phosphorylation in RPTEC cells. Furthermore, TGF-β1-induced mitochondrial fission in RPTEC cells was ameliorated by AD-021 via mechanisms involving inhibition of Drp1 phosphorylation. In a mouse model of unilateral ureteral obstruction (UUO)-induced renal fibrosis, AD-021 reduced plasma TGF-β1, ameliorated renal fibrosis and improved renal function. Collectively, AD-021 represents a novel class of natural product-based anti-fibrotic agent that has therapeutic potential in the prevention of fibrosis-associated renal disorders including CKD.

Human Wharton's jelly mesenchymal stem cells derived-exosomes enriched by miR-124 promote an anti-fibrotic response in an experimental model of liver fibrosis.

Liver fibrosis is a significant challenge to global health that results in organ failure through inflammation and the release of fibrotic biomarkers. Due to the lack of effective treatments for liver fibrosis, anti-fibrotic and anti-inflammatory therapies are being developed. Since there has been an association between aberrant expression of miR-124 and liver disease progression, we investigated whether delivery of miR-124 through human Wharton's jelly mesenchymal stem cells derived-exosomes (hWJMSC-Exo) can improve liver fibrosis. We established a 6-week carbon tetrachloride (CCl4)-induced mouse model of liver fibrosis, then we administered hWJMSC-Exo and miR-124-3p-enriched exosomes (ExomiR-124) for three weeks. The extent of fibrosis and inflammation was assessed by histology, biochemistry, Real-time PCR, immunohistochemistry, and Enzyme-linked immunoassays (ELISA). The inflammatory status of the spleen was also investigated using flow cytometry. Based on the gene and protein expression measurement of IL-6, IL-17, TGF-β, STAT3, α-SMA, and COL1, In vivo administration of Exo and ExomiR-124 effectively reduce collagen accumulation and inhibition of inflammation. Regarding histopathology findings, the therapeutic effect of ExomiR-124 against liver fibrosis was significantly greater than hWJMSC-Exo. In addition, we found that Exo and ExomiR-124 was capable of phenotype switching of splenic monocytes from inflammatory Ly6Chi to restorative Ly6Clo. MSC-derived exosomes demonstrated anti-inflammatory effect via different aspects. Aside from the therapeutic approach, enrichment of exosomes as a nanocarrier by miR-124 revealed the down-regulation of STAT3, which plays a crucial role in liver fibrosis. The anti-inflammatory and anti-fibrotic properties of ExomiR-124 could be a promising option in liver fibrosis combination therapies.

Inhibition of the extracellular enzyme A disintegrin and metalloprotease with thrombospondin motif 4 prevents cardiac fibrosis and dysfunction.

Heart failure is a condition with high mortality rates, and there is a lack of therapies that directly target maladaptive changes in the extracellular matrix (ECM), such as fibrosis. We investigated whether the ECM enzyme known as A disintegrin and metalloprotease with thrombospondin motif (ADAMTS) 4 might serve as a therapeutic target in treatment of heart failure and cardiac fibrosis. The effects of pharmacological ADAMTS4 inhibition on cardiac function and fibrosis were examined in rats exposed to cardiac pressure overload. Disease mechanisms affected by the treatment were identified based on changes in the myocardial transcriptome. Following aortic banding, rats receiving an ADAMTS inhibitor, with high inhibitory capacity for ADAMTS4, showed substantially better cardiac function than vehicle-treated rats, including ∼30% reduction in E/e' and left atrial diameter, indicating an improvement in diastolic function. ADAMTS inhibition also resulted in a marked reduction in myocardial collagen content and a down-regulation of transforming growth factor (TGF)-β target genes. The mechanism for the beneficial effects of ADAMTS inhibition was further studied in cultured human cardiac fibroblasts producing mature ECM. ADAMTS4 caused a 50% increase in the TGF-β levels in the medium. Simultaneously, ADAMTS4 elicited a not previously known cleavage of TGF-β-binding proteins, i.e. latent-binding protein of TGF-β and extra domain A-fibronectin. These effects were abolished by the ADAMTS inhibitor. In failing human hearts, we observed a marked increase in ADAMTS4 expression and cleavage activity. Inhibition of ADAMTS4 improves cardiac function and reduces collagen accumulation in rats with cardiac pressure overload, possibly through a not previously known cleavage of molecules that control TGF-β availability. Targeting ADAMTS4 may serve as a novel strategy in heart failure treatment, in particular, in heart failure with fibrosis and diastolic dysfunction.

Nintedanib Ameliorates Bleomycin-Induced Pulmonary Fibrosis, Inflammation, Apoptosis, and Oxidative Stress by Modulating PI3K/Akt/mTOR Pathway in Mice

Idiopathic pulmonary fibrosis (IPF) seriously threatens human life and health, and no curative therapy is available at present. Nintedanib is the first agent approved by the US Food and Drug Administration (FDA) in order to treat IPF; however, its mechanism of inhibition of IPF is still elusive. According to recent studies, nintedanib is a potent inhibitor. It can antagonize platelet-derived growth factor (PDGF), basic fibroblast growth factor (b-FGF), vascular endothelial growth factor (VEGF), etc., to inhibit pulmonary fibrosis. Whether there are other signaling pathways involved in IPF remains unknown. This study focused on investigating the therapeutic efficacy of nintedanib in bleomycin-mediated pulmonary fibrosis (PF) mice through PI3K/Akt/mTOR pathway. Following the induction of pulmonary fibrosis in C57 mice through bleomycin (BLM) administration, the mice were randomized into five groups: (1) the normal control group, (2) the BLM model control group, (3) the low-dose Nintedanib administration model group, (4) the medium-dose nintedanib administration model group, and (5) the high-dose nintedanib administration model group. For lung tissues, morphological changes were found by HE staining and Masson staining, ELISA method was used to detect inflammatory factors, alkaline water method to estimate collagen content, and western blotting for protein levels. TUNEL staining and immunofluorescence methods were used to analyze the effect of nintedanib on lung tissue and the impacts and underlying mechanisms of bleomycin-induced pulmonary fibrosis. After 28 days, bleomycin-treated mice developed significant pulmonary fibrosis. Relative to bleomycin-treated mice, nintedanib-treated mice had markedly reduced degrees of PF. In addition, nintedanib showed lung-protective effects by up-regulating antioxidant levels, down-regulating inflammatory protein expression, and reducing collagen accumulation. We demonstrated that nintedanib ameliorated bleomycin-induced lung injury by inhibiting the P13K/Akt/mTOR pathway as well as apoptosis. In addition, significant improvement in pulmonary fibrosis was seen after nintedanib (30/60/120 mg/kg body weight/day) treatment through a dose-dependent way. Histopathological results further corroborated the effect of nintedanib treatment on remarkably attenuating bleomycin-mediated mouse lung injury. According to our findings, nintedanib restores the antioxidant system, suppresses pro-inflammatory factors, and inhibits apoptosis. Nintedanib can reduce bleomycin-induced inflammation by downregulating PI3K/Akt/mTOR pathway, PF, and oxidative stress (OS).

Sclareol attenuates liver fibrosis through SENP1-mediated VEGFR2 SUMOylation and inhibition of downstream STAT3 signaling.

Liver fibrosis is a key global health care burden. Sclareol, isolated from Salvia sclarea, possesses various biological activities. Its effect on liver fibrosis remains unknown. This study was proposed to evaluate the antifibrotic activity of sclareol (SCL) and explore its underlying mechanisms. Stimulated hepatic stellate cells served as an in vitro liver fibrosis model. The expression of fibrotic markers was assessed by western blot and real-time PCR. Two classical animal models, bile duct-ligated rats and carbon tetrachloride-treated mice, were utilized for the in vivo experiments. The liver function and fibrosis degree were determined by serum biochemical and histopathological analyses. VEGFR2 SUMOylation was analyzed using coimmunoprecipitation assay. Our results indicated that SCL treatment restricted the profibrotic propensity of activated HSCs. In fibrotic rodents, SCL administration alleviated hepatic injury and reduced collagen accumulation. Mechanistic studies indicated that SCL downregulated the protein level of SENP1 and enhanced VEGFR2 SUMOylation in LX-2 cells, which affected its intracellular trafficking. Blockade of the interaction between VEGFR2 and STAT3 was observed, resulting in the suppression of downstream STAT3 phosphorylation. Our findings demonstrated that SCL has therapeutic efficacy against liver fibrosis through mediating VEGFR2 SUMOylation, suggesting that SCL may be a potential candidate compound for its treatment.

HKLK alleviates myocardial fibrosis in mice with viral myocarditis.

Myocardial fibrosis is the most serious complication of viral myocarditis (VMC). This study aimed to investigate the therapeutic benefits and underlying mechanisms of lentivirus-mediated human tissue kallikrein gene transfer in myocardial fibrosis in VMC mice. We established VMC mouse model via intraperitoneal injection with Coxsackie B3 virus. The effect was then assessed after treatment with vehicle, the empty lentiviral vectors (EZ.null), and the vectors expressing hKLK1 (EZ.hKLK1) via tail vein injection for 30 days, respectively. The results showed that administering EZ.hKLK1 successfully induced hKLK1 overexpression in mouse heart. Compared with EZ.null treatment, EZ.hKLK1 administration significantly reduced the heart/weight ratio, improved cardiac function, and ameliorated myocardial inflammation in VMC mice, suggesting that hKLK1 overexpression alleviates VMC in mice. EZ.hKLK1 administration also significantly abrogated the increased myocardial collagen content, type I/III collagen ratio, TGF-β1 mRNA and protein expression in VMC mice, suggesting that hKLK1 overexpression reduces collagen accumulation and blunts TGF-β1 signaling in the hearts of VMC mice. In conclusion, our results suggest that hKLK1 alleviates myocardial fibrosis in VMC mice, possibly by downregulating TGF-β1 expression.

The protective effects of a novel AT2 receptor agonist, β-Pro7Ang III in ischemia-reperfusion kidney injury

Background and purposeThis study investigated the reno-protective effects of a highly selective AT2R agonist peptide, β-Pro7Ang III in a mouse model of acute kidney injury (AKI). MethodsC57BL/6 J mice underwent either sham surgery or unilateral kidney ischemia-reperfusion injury (IRI) for 40 min. IRI mice were treated with either β-Pro7Ang III or perindopril and at 7 days post-surgery the kidneys analysed for histopathology and the development of fibrosis and matrix metalloproteinase (MMP)-2 and -9 activity. The association of the therapeutic effects of β-Pro7Ang III with macrophage number and phenotype was determined in vivo and in vitro. Key resultsDecreased kidney tubular injury, interstitial matrix expansion and reduced interstitial immune cell infiltration in IRI mice receiving β-Pro7Ang III treatment was observed at day 7, compared to IRI mice without treatment. This correlated to reduced collagen accumulation and MMP-2 activity in IRI mice following β-Pro7Ang III treatment. FACS analysis showed a reduced number and proportion of CD45+CD11b+F4/80+ macrophages in IRI kidneys in response to β-Pro7Ang III, correlating with a significant increase in M2 macrophage markers and decreased M1 markers at day 3 and 7 post-IR injury, respectively. In vitro analysis of cultured THP-1 cells showed that β-Pro7Ang III attenuated lipopolysaccharide (LPS)-induced tumour necrosis factor-α (TNF-α) and interleukin (IL)− 6 production but increased IL-10 secretion, compared to LPS alone. ConclusionAdministration of β-Pro7Ang III via mini-pump improved kidney structure and reduced interstitial collagen accumulation, in parallel with an alteration of macrophage phenotype and anti-inflammatory cytokine release, therefore mitigating the downstream progression of ischemic AKI.

Mangiferin relieves CCl4-induced liver fibrosis in mice

Hepatic fibrosis is a late stage process of many chronic liver diseases. Blocking the fibrosis process will be beneficial to the treatment and recovery of the diseases. Mangiferin has many pharmacological activities. Recently, it has been reported that mangiferin may relieve tissue fibrosis, including renal, myocardial, pulmonary fibrosis via anti-inflammatory and anti-oxidative effects in animal models. Here, we investigate the effects of mangiferin on CCl4-induced liver fibrosis and the underlying mechanism in mice. Thirty-two male C57BL/6 mice were randomly divided into 4 groups (n = 8 in each group), injected with carbon tetrachloride (10% CCl4) for 8 weeks, and oral administrated with mangiferin (50 mg/kg or 100 mg/kg) from the fifth week. The serum levels of ALT, AST were analyzed to evaluate liver function. H&E, Masson’s trichrome and Sirius red staining were used to assess liver morphology and the degree of liver fibrosis. Quantitative RT-PCR and Western blot were used to assay the gene expression and protein levels. The results showed that mangiferin alleviated the serum levels of AST, ALT, ALP, TBA and TBIL, reduced liver lesions, prevented hepatic parenchymal necrosis, and ameliorated collagen accumulation in the liver of CCl4-treated mice. Meanwhile, mangiferin inhibited the expression of inflammatory genes IL-6 and IL-1β, fibrogenic genes α-SMA, TGF-β and MMP-2 and bile acid metabolism genes ABCB4, ABCB11, SULT2A1 in the liver of CCl4-treated mice. Furthermore, mangiferin reduced collagen accumulation and HSCs activation, inhibited the p-IκB and p-p65 protein levels. Our results suggest that mangiferin could alleviate liver fibrosis in CCl4-treated mice through inhibiting NF-κB signaling, and mango consuming may have beneficial effects to hepatic fibrosis.