Decreased Collagen Deposition Research Articles

Nifuroxazide-loaded cubosomes exhibit an advancement in pulmonary delivery and attenuate bleomycin-induced lung fibrosis by regulating the STAT3 and NF-κB signaling: A new challenge for unmet therapeutic needs

Pulmonary fibrosis (PF) is a chronic progressive disease that portends a very poor prognosis. It has been suggested that STAT3 is a potential target in PF. This study highlights the importance of cubosomes as a drug delivery system in enhancing the bioavailability of nifuroxazide (NXZD), a poorly soluble STAT3 inhibitor. NXZD-loaded cubosomes (NXZD-LC) were in vitro and in vivo evaluated. In vitro, cubosomes presented a poly-angular nanosized particles with a mean size and zeta potential of 223.73 ± 4.73 nm and − 20.93 ± 2.38 mV, respectively. The entrapment efficiency of nifuroxazide was 90.56 ± 4.25%. The in vivo pharmacokinetic study and the lung tissue accumulation of NXZD were performed by liquid chromatography-tandem mass spectrometry after oral administration to rats. The nanoparticles exhibited a two-fold increase and 1.33 times of bioavailability and lung tissue concentration of NXZD compared to NXZD dispersion, respectively. In view of this, NXZD-LC effectively attenuated PF by targeting STAT3 and NF-κB signals. As a result, NXZD-LC showed a potential anti-inflammatory effect as revealed by the significant decrease in MCP-1, ICAM-1, IL-6, and TNF-α and suppressed fibrogenic mediators as indicated by the significant reduction in TGF-β, TIMP-1, and PDGF-BB in lung tissues. Besides, NXZD-LC improved antioxidant defense mechanisms and decreased LDH and BALF total protein. These effects contributed to decreased collagen deposition. To conclude, cubosomes represent an advantageous pharmaceutical delivery system for enhancing pulmonary delivery of poorly soluble drugs. Additionally, repurposing NXZD as an antifibrotic agent is a promising challenge and new therapeutic approach for unmet therapeutic needs.

Improved bladder contractility after transplantation of human mesenchymal stem cells overexpressing hepatocyte growth factor into underactive bladder from bladder outlet obstruction models of rats.

IntroductionAn underactive bladder can lead to difficulty in voiding that causes incomplete emptying of the bladder, suggesting the need for a new strategy to increase bladder contractility in such patients. This study was performed to investigate whether human mesenchymal stem cells (hMSCs) were capable of restoring bladder contractility in rats with underactive bladder due to bladder outlet obstruction (BOO) and enhancing their effects by overexpressing hepatocyte growth factor (HGF) in hMSCs.Materials and methodsThe hMSCs were transplanted into the bladder wall of rats. Fifty female Sprague-Dawley rats at six weeks of age were divided into five groups: group 1: control; group 2: sham intervention; group 3: eight-week BOO; group 4: BOO rats transplanted with hMSCs; and group 5: BOO rats transplanted with hMSCs overexpressing HGF. Two weeks after the onset of BOO in groups 4 and 5, hMSCs were injected into the bladder wall. Cystometry evaluation was followed by Masson’s trichrome staining of bladder tissues. Realtime PCR and immunohistochemical staining were performed to determine for hypoxia, apoptosis, and angiogenesis.ResultsCollagen deposition of bladder increased in BOO but decreased after transplantation of hMSCs. The increased inter-contraction interval and residual urine volume after BOO was reversed after hMSCs transplantation. The decreased maximal voiding pressure after BOO was restored by hMSCs treatment. The mRNA expression of bladder collagen1 and TGF-β1 increased in BOO but decreased after hMSCs transplantation. The decrease in vWF-positive cells in the bladder following BOO was increased after hMSCs transplantation. Caspase 3 and TUNEL-positive apoptosis of bladder cells increased in BOO but decreased after transplantation of hMSCs. These effects were enhanced by overexpressing HGF in hMSCs.ConclusionTransplantation of hMSCs into bladder wall increased the number of micro-vessels, decreased collagen deposition and apoptosis of detrusor muscle, and improved bladder underactivity. The effects were enhanced by overexpressing HGF in hMSCs. Our findings suggest that the restoration of underactive bladder using hMSCs may be used to rectify micturition disorders in patients following resolution of BOO. Further studies are needed before hMSCs can be used in clinical applications.

Hepatoprotective effects of norgalanthamine on carbon tetrachloride induced-hepatotoxicity in mice

Norgalanthamine is a major component of Crinum asiaticum var. japonicum that exhibits several biological activities. This study evaluated the anti-inflammatory and anti-oxidative properties of norgalanthamine in mice with carbon tetrachloride (CCl4)-induced liver injury. Norgalanthamine (1 and 10 mg/kg) was orally administered to mice for 7 or 14 days, after which liver injury was induced by CCl4 (1.5 ml/kg, i.p.). The vehicle and positive controls consisted of phosphate-buffered saline and silymarin (100 mg/kg), respectively. In CCl4-injured mice, norgalanthamine pretreatment significantly reversed the increases in serum alanine aminotransferase, aspartate aminotransferase, and total bilirubin levels, and the decrease in the serum glucose level. In the liver, norgalanthamine restored the activities of the antioxidant enzymes superoxide dismutase and catalase, while reducing lipid accumulation and, concurrently, the expression of genes involved in lipid synthesis, including peroxisome proliferator-activated receptor γ and adipocyte protein-2. Norgalanthamine also ameliorated inflammation by down-regulating the expression of the pro-inflammatory mediators, TNF-α, IL-1β, and MCP-1, and up-regulating the Nrf2/HO-1 pathway. In addition, norgalanthamine decreased collagen deposition in liver tissue as shown on picrosirius red staining by down-regulating expression of the fibrosis-related genes αSMA and fibronectin. Collectively, these findings imply that norgalanthamine mitigates CCl4-induced hepatic injury by increasing anti-oxidative activity, down-regulating pro-inflammatory mediators and fibrosis-related genes in the liver. Highlights Norgalanthamine ameliorated the hepatotoxicity after CCl4 injury. Norgalanthamine suppressed the activation of Kupffer cells and macrophages. Norgalanthamine down-regulated pro-inflammatory mediators. Norgalanthamine increased anti-oxidative activity via the Nrf2/HO-1 pathway. Norgalanthamine downregulated fibrosis-related genes in the liver.

Syzygium aqueum (Burm.f.) Alston Prevents Streptozotocin-Induced Pancreatic Beta Cells Damage via the TLR-4 Signaling Pathway.

Although several treatments are available for the treatment of type 2 diabetes mellitus, adverse effects and cost burden impose the search for safe, efficient, and cost-effective alternative herbal remedies. Syzygium aqueum (Burm.f.) Alston, a natural anti-inflammatory, antioxidant herb, may suppress diabetes-associated inflammation and pancreatic beta-cell death. Here, we tested the ability of the bioactive leaf extract (SA) to prevent streptozotocin (STZ)-induced oxidative stress and inflammation in pancreatic beta cells in rats and the involvement of the TLR-4 signaling pathway. Non-fasted rats pretreated with 100 or 200 mg kg−1 SA 2 days prior to the STZ challenge and for 14 days later had up to 52 and 39% reduction in the glucose levels, respectively, while glibenclamide, the reference standard drug (0.5 mg kg−1), results in 70% reduction. Treatment with SA extract was accompanied by increased insulin secretion, restoration of Langerhans islets morphology, and decreased collagen deposition as demonstrated from ELISA measurement, H and E, and Mallory staining. Both glibenclamide and SA extract significantly decreased levels of TLR-4, MYD88, pro-inflammatory cytokines TNF-α, and TRAF-6 in pancreatic tissue homogenates, which correlated well with minimal pancreatic inflammatory cell infiltration. Pre-treatment with SA or glibenclamide decreased malondialdehyde, a sensitive biomarker of ROS-induced lipid peroxidation, and restored depleted reduced glutathione in the pancreas. Altogether, these data indicate that S. aqueum is effective in improving STZ-induced pancreatic damage, which could be beneficial in treating type 2 diabetes mellitus.

Protection of CAPE-pNO2 Against Chronic Myocardial Ischemia by the TGF-Β1/Galectin-3 Pathway In Vivo and In Vitro.

Although it is known that caffeic acid phenethyl ester (CAPE) and its derivatives could ameliorate acute myocardial injury, their effects on chronic myocardial ischemia (CMI) were not reported. This study aimed to investigate the potential effect of caffeic acid p-nitro phenethyl ester (CAPE-pNO2, a derivative of CAPE) on CMI and underlying mechanisms. SD rats were subjected to high-fat-cholesterol-diet (HFCD) and vitamin D3, and the H9c2 cells were treated with LPS to establish CMI model, followed by the respective treatment with saline, CAPE, or CAPE-pNO2. In vivo, CAPE-pNO2 could reduce serum lipid levels and improve impaired cardiac function and morphological changes. Data of related assays indicated that CAPE-pNO2 downregulated the expression of transforming growth factor-β1 (TGF-β1) and galectin-3 (Gal-3). Besides, CAPE-pNO2 decreased collagen deposition, the number of apoptotic cardiomyocytes, and some related downstream proteins of Gal-3 in the CMI rats. Interestingly, the effects of CAPE-pNO2 on TGF-β1, Gal-3, and other proteins expressed in the lung were consistent with that in the heart. In vitro, CAPE-pNO2 could attenuate the fibrosis, apoptosis, and inflammation by activating TGF-β1/Gal-3 pathway in LPS-induced H9c2 cell. However, CAPE-pNO2-mediated cardioprotection can be eliminated when treated with modified citrus pectin (MCP, an inhibitor of Gal-3). And in comparison, CAPE-pNO2 presented stronger effects than CAPE. This study indicates that CAPE-pNO2 may ameliorate CMI by suppressing fibrosis, inflammation, and apoptosis via the TGF-β1/Gal-3 pathway in vivo and in vitro.

Comparison of the Therapeutic Effects of hUC-MSC Intravenous Delivery and Intraperitoneal Administration of MSCs Encapsulated in Alginate Capsules for the Treatment of Rat Liver Cirrhosis

Mesenchymal stem cells are the most promising regenerative medicine tool for treating various diseases, including liver disease, although the exact mechanism of their therapeutic action remains unclear. It was found that MSCs are captured by the lungs after systemic transplantation, quickly disappear, and are not detected at the site of injury but at the same time exhibit an obvious therapeutic effect. Comparison of the MSC efficiency depending on the route of their administration may shed light on the mechanisms involved in implementing MSC therapeutic potential. In this work, we compared the therapeutic effects of human umbilical cord MSCs (hUC-MSCs) administered systemically and intraperitoneally in the form of MSCs encapsulated in alginate capsules in a CCl4-induced model of liver cirrhosis in rats. Our study showed that both treatments resulted in liver recovery. MSC transplantation by two different routes led to a decrease in collagen deposition, the disappearance of the fibrous area by the 13th week, and the normalization of the morphometric parameters of liver parenchyma cells. In addition, the expression of some genes (EGF, alpha SMA, GFAP) which is activated in liver injury, decreased to the level observed in negative control animals. However, a detailed study of liver recovery in dynamics showed that encapsulated MSCs led to faster normalization in several parameters of the liver tissue. Our results showed that human umbilical cord MSCs effectively exhibit their therapeutic properties when using both transplantation methods. However, intraperitoneal administration of encapsulated MSCs accelerated the process of liver regeneration.

Vascular Stiffening Mediated by Rho-Associated Coiled-Coil Containing Kinase Isoforms.

BackgroundThe pathogenesis of vascular stiffening and hypertension is marked by non‐compliance of vessel wall because of deposition of collagen fibers, loss of elastin fibers, and increased vascular thickening. Rho/Rho‐associated coiled‐coil containing kinases 1 and 2 (ROCK1 and ROCK2) have been shown to regulate cellular contraction and vascular remodeling. However, the role of ROCK isoforms in mediating pathogenesis of vascular stiffening and hypertension is not known.Methods and ResultsHemizygous Rock mice (Rock1 +/− and Rock2 +/−) were used to determine the role of ROCK1 and ROCK2 in age‐related vascular dysfunction. Both ROCK activity and aortic stiffness increased to a greater extent with age in wild‐type mice compared with that of Rock1 +/− and Rock2 +/− mice. As a model for age‐related vascular stiffening, we administered angiotensin II (500 ng/kg per minute) combined with nitric oxide synthase inhibitor, L‐Nω‐nitroarginine methyl ester (0.5 g/L) for 4 weeks to 12‐week‐old male Rock1 +/− and Rock2 +/− mice. Similar to advancing age, angiotensin II/L‐Nω‐nitroarginine methyl ester caused increased blood pressure, aortic stiffening, and vascular remodeling, which were attenuated in Rock2 +/−, and to a lesser extent, Rock1 +/− mice. The reduction of aortic stiffening in Rock2 +/− mice was accompanied by decreased collagen deposition, relatively preserved elastin content, and less aortic wall hypertrophy. Indeed, the upregulation of collagen I by transforming growth factor‐β1 or angiotensin II was greatly attenuated in Rock2 −/− mouse embryonic fibroblasts.ConclusionsThese findings indicate that ROCK1 and ROCK2 mediate both age‐related and pharmacologically induced aortic stiffening, and suggest that inhibition of ROCK2, and to a lesser extent ROCK1, may have therapeutic benefits in preventing age‐related vascular stiffening.

Tetrathiomolybdate (TM)-associated copper depletion influences collagen remodeling and immune response in the pre-metastatic niche of breast cancer

Tetrathiomolybdate (TM) is a novel, copper-depleting compound associated with promising survival in a phase II study of patients with high-risk and triple-negative breast cancer. We sought to elucidate the mechanism of TM by exploring its effects on collagen processing and immune function in the tumor microenvironment (TME). Using an exploratory cohort, we identified markers of collagen processing (LOXL2, PRO-C3, C6M, and C1M) that differed between those with breast cancer versus controls. We measured these collagen biomarkers in TM-treated patients on the phase II study and detected evidence of decreased collagen cross-linking and increased degradation over formation in those without disease compared to those who experienced disease progression. Preclinical studies revealed decreased collagen deposition, lower levels of myeloid-derived suppressor cells, and higher CD4+ T-cell infiltration in TM-treated mice compared with controls. This study reveals novel mechanisms of TM targeting the TME and immune response with potential applications across cancer types.

New Therapeutic Approach for Intestinal Fibrosis Through Inhibition of pH-Sensing Receptor GPR4.

Patients suffering from inflammatory bowel diseases (IBDs) express increased mucosal levels of pH-sensing receptors compared with non-IBD controls. Acidification leads to angiogenesis and extracellular matrix remodeling. We aimed to determine the expression of pH-sensing G protein-coupled receptor 4 (GPR4) in fibrotic lesions in Crohn's disease (CD) patients. We further evaluated the effect of deficiency in Gpr4 or its pharmacologic inhibition. Paired samples from fibrotic and nonfibrotic terminal ileum were obtained from CD patients undergoing ileocaecal resection. The effects of Gpr4 deficiency were assessed in the spontaneous Il-10-/- and the chronic dextran sodium sulfate (DSS) murine colitis model. The effects of Gpr4 deficiency and a GPR4 antagonist (39c) were assessed in the heterotopic intestinal transplantation model. In human terminal ileum, increased expression of fibrosis markers was accompanied by an increase in GPR4 expression. A positive correlation between the expression of procollagens and GPR4 was observed. In murine disease models, Gpr4 deficiency was associated with a decrease in angiogenesis and fibrogenesis evidenced by decreased vessel length and expression of Edn, Vegfα, and procollagens. The heterotopic animal model for intestinal fibrosis, transplanted with terminal ileum from Gpr4-/- mice, revealed a decrease in mRNA expression of fibrosis markers and a decrease in collagen content and layer thickness compared with grafts from wild type mice. The GPR4 antagonist decreased collagen deposition. The GPR4 expression was also observed in human and murine intestinal fibroblasts. The GPR4 inhibition reduced markers of fibroblast activation stimulated by low pH, notably Acta2 and cTgf. Expression of GPR4 positively correlates with the expression of profibrotic genes and collagen. Deficiency of Gpr4 is associated with a decrease in angiogenesis and fibrogenesis. The GPR4 antagonist decreases collagen deposition. Targeting GPR4 with specific inhibitors may constitute a new treatment option for IBD-associated fibrosis.

1: Fat Grafting Improves Dorsal Skin Fibrosis after Radiation in an Engrailed-1 Mouse Model

Purpose:Fat grafting is known to rejuvenate and improve fibrosis in irradiated skin, but the underlying mechanisms behind these effects are poorly understood. We have previously identified the Engrailed-1 fibroblast sub-population as having a role in post-natal dorsal skin fibrosis in mice. Using a radiation model to the scalp of Engrailed-1 mice, we sought to better understand the regenerative effects of fat grafting in irradiated tissue.Methods:Adult (60-day old) En1Cre;R26mTmG transgenic mice (n=5) underwent total body irradiation with 9 Gy for immunodepletion. Mice were then immediately reconstituted with 2 million nucleated bone marrow cells from donor NSG(NOD.CB17-Prkdcsscid/J) mice via retro-orbital injections. Reconstitution was confirmed with fluorescence-activated cell sorting (FACS) 2 weeks after whole body irradiation. Mice scalps were then irradiated with 5 Gy every other day for 12 days (30 Gy total) and allowed to recover for 4 weeks to facilitate fibrotic conversion. Irradiated scalps were grafted with 200μL of fresh human lipoaspirate. Graph retention was measured in-vivo for 8 weeks using Micro-Computed Tomography Scanner (Bruker SkyScan 1726™) and scalp skin was harvest for histology.Results:Two weeks post bone-marrow transplant, >90% of circulating hematopoietic cells (CD45+) cells in En1Cre;R26mTmG reporter mice were non-fluorescent, signifying successful reconstitution. Volumetric analysis of fat grafting in-vivo was performed using 3-dimensional reconstruction. At 8 weeks post-grafting, 4 of the 5 grafts demonstrated >50% graft retention, confirming successful grafting. Histological sections of scalp skin 8 weeks post-grafting demonstrated significantly less En1+GFP cells compared to non-grafted scalp skin, signifying less presence of En1+ fibroblasts. Furthermore, histology demonstrated significantly less dermal thickening, epidermal thinning, and collagen deposition and disorganization in fat grafted irradiated scalp skin compared to non-grafted scalp skin.Conclusion:Fat grafting mitigates radiation-induced fibrosis in scalp skin by decreasing collagen deposition, remodeling collagen formation, decreasing epidermal thinning, and reducing presence of pro-fibrotic fibroblast sub-populations.

Deregulated glutamate to pro-collagen conversion is associated with adverse outcome in lung cancer and may be targeted by renin-angiotensin-aldosterone system (RAS) inhibition

BackgroundThe tumor-microenvironment (TME) represents an attractive therapeutic target in NSCLC and plays an important role for efficacy of cancer therapeutics. We hypothesized that upregulation of collagen synthesis might be associated with adverse outcome in NSCLC. Literature evidence suggests that renin-angiotensin system inhibitors (RASi) decrease collagen deposition. Therefore, we aimed to explore the prognostic role of RASi intake and their influence on the TME in NSCLC. MethodsFour publicly available datasets were used to evaluate the impact of key enzymes involved in collagen biosynthesis. To investigate the influence of RASi intake on the TME and prognosis we evaluated a cohort of metastatic NSCLC patients and performed histopathological characterization of the TME. A three-dimensional microtissue in vitro model was developed to define the impact of RASi on collagen synthesis. ResultsExpression of three genes of the collagen synthesis pathway, ALDH18A1, PLOD2 and P4HA1, was upregulated in NSCLC compared to normal lung tissue and linked to shortened overall survival in all investigated cohorts. Together, these genes formed a ‘Collagen Signature’ which represents an independent unfavourable prognostic factor in two NSCLC cohorts and was linked to alterations of the extracellular matrix deposition and cell cycle pathways. In the cohort of metastatic NSCLC, RASi intake was linked to improved overall response rate and survival. Exploratory in vitro experiments revealed that RASi led to a dose dependent reduction of collagen deposition and degradation of three-dimensional lung cancer cell spheroids. ConclusionWe demonstrate that collagen synthesis is a key upregulated process in the NSCLC TME and its transcriptional readout, the three gene Collagen Signature is independently associated with poor outcome. Pharmacological targeting of this pathways e.g. by RASi bears potential of improving outcome in NSCLC.

PH-Triggered Aggregation of Gold Nanoparticles for Enhanced Labeling and Long-Term CT Imaging Tracking of Stem Cells in Pulmonary Fibrosis Treatment.

Gold nanoparticles (AuNPs) pose a great challenge in the development of nanotracers that can self-adaptively alter their properties in response to certain cellular environments for long-term stem cell tracking. Herein, pH-sensitive Au nanotracers (CPP-PSD@Au) are fabricated by sequential coupling of AuNPs with sulfonamide-based polymer (PSD) and cell-penetrating peptide (CPP), which can be efficiently internalized by mesenchymal stem cells (MSCs) and undergo pH-induced self-assembly in endosomes, facilitating long-term computed tomography (CT) imaging tracking MSCs in a murine model of idiopathic pulmonary fibrosis (IPF). Using the CPP-PSD@Au, the transplanted MSCs for the first time can be monitored with CT imaging for up to 35 days after transplantation into the lung of IPF mice, clearly elucidating the migration process of MSCs in vivo. Moreover, we preliminarily explored the mechanism of the CPP-PSD@Au labeled MSCs in the alleviation of IPF, including recovery of alveolar integrity, decrease of collagen deposition, as well as down-regulation of relevant cytokine level. This work facilitates our understanding of the behavior and effect of MSCs in the therapy of IPF, thereby providing an important insight into the stem cell-based treatment of lung diseases.

H2S alleviates aortic aneurysm and dissection: Crosstalk between transforming growth factor 1 signaling and NLRP3 inflammasome

BackgroundVascular remodeling and inflammation are involved in aortic aneurysm (AA) and aortic dissection (AD). TGF-β1 signaling is involved in tissue fibrosis, extracellular matrix remodeling and inflammation, which are linked with AA and AD. The inhibition of NLRP3 inflammasome suppresses AA and AD. Hydrogen sulfide (H2S) exerts anti-vascular remodeling and anti-inflammatory properties, but little is known about its action on AA and AD progression. MethodsThe effect of H2S on AA and AD formation was investigated in Sprague-Dawley (SD) rat fed a normal diet supplemented with 0.25% β-aminopropionitrile (BAPN). HE staining, Masson's trichrome staining, Picrosirius red staining and EVG staining were to evaluate vascular remodeling in the aortic wall. Western blotting and IHC were to detect the expression of TGF-β1 and matrix metalloproteinases (MMPs) and NLRP3 inflammasome-associated proteins. The interaction between TGF-β1 signaling and NLRP3 inflammasome was explored in Human aortic vascular smooth muscle cells (HA-VSMCs). ResultsH2S alleviated AA and AD progression. Specifically, it improved irregular tissue arrangement and vascular fibrosis, increased the expression of elastin fibers, decreased collagen deposition and the expression of TGF-β1 and matrix metalloproteinases (MMP-2/9). In addition, H2S inhibited the expression of proteins involved in NLRP3 inflammasome. Furthermore, H2S down-regulated TGF-β1 signaling and then ameliorated vascular fibrosis by preventing NLRP3 inflammasome activation. Finally, H2S inhibited NLRP3 inflammasome activation and decreased the level of IL-1β by disrupting TGF-β1 signaling. ConclusionsThese data support a crosstalk between TGF-β1 signaling and NLRP3 inflammasome. H2S inhibits AA and AD progression via blocking the crosstalk.

Chaihu Guizhi Ganjiang Decoction Ameliorates Pancreatic Fibrosis via JNK/mTOR Signaling Pathway.

Pancreatic fibrosis is a pathological characteristic of chronic pancreatitis (CP) and pancreatic cancer. Chaihu Guizhi Ganjiang Decoction (CGGD) is a traditional Chinese medicine, which is widely used in the clinical treatment of digestive diseases. However, the potential anti-fibrosis mechanism of CGGD in treating CP remains unclear. Here, we conducted a series of experiments to examine the effect of CGGD on the CP rat model and primary isolated pancreatic stellate cells (PSCs). The results revealed that CGGD attenuated pancreatic damage, decreased collagen deposition, and inhibited PSC activation in the pancreas of CP rats. However, compared with the CP group, CGGD had no effect on body weight and serum amylase and lipase. In addition, CGGD suppressed autophagy by downregulating Atg5, Beclin-1, and LC3B and facilitated phosphorylation of mTOR and JNK in pancreatic tissues and PSCs. Moreover, the CGGD-containing serum also decreased LC3B or collagen I expression after rapamycin (mTOR inhibitor) or SP600125 (JNK inhibitor) treatment in PSCs. In conclusion, CGGD attenuated pancreatic fibrosis and PSC activation, possibly by suppressing autophagy of PSCs through the JNK/mTOR signaling pathway.

RS4651 suppresses lung fibroblast activation via the TGF-β1/SMAD signalling pathway

BackgroundIdiopathic pulmonary fibrosis (IPF) is a progressive disease resulting in respiratory failure with no efficient treatment options. We investigated the protective effect of RS4651 on pulmonary fibrosis in mice and the mechanism. MethodsIntratracheal injection of bleomycin (BLM) was used to induce pulmonary fibrosis in mice. RS4561 was administered intraperitoneally at different doses. Histopathological changes were observed. The level of alpha-smooth muscle actin (α-SMA) were also tested. In vitro, the proliferation and migratory effects of RS4651 treatment on MRC-5 cells pre-treated with transforming growth factor (TGF-β1) were examined. RNA-sequencing was used to detect differentially expressed target genes. Then, the expression of α-SMA, pSMAD2 and SMAD7 were analysed during RS4651 treatment of MRC-5 cells with or without silencing by SMAD7 siRNA. ResultsHistopathological staining results showed decreased collagen deposition in RS4651 administered mice. Additionally, a lower level of α-SMA was also observed compared to the BLM group. The results of in vitro studies confirmed that RS4651 can inhibit the proliferation and migration, as well as α-SMA and pSMAD2 expression in MRC-5 cells treated with TGF-β1. RNA-sequencing data identified the target gene SMAD7. We found that RS4651 could upregulate SMAD7 expression and inhibit the proliferation and migration of MRC-5 cells via SMAD7, and RS4651 inhibition of α-SMA and pSMAD2 expression was blocked in SMAD7-siRNA MRC-5 cells. In vivo studies further confirmed that RS4651 could upregulate SMAD7 expression in BLM-induced lung fibrosis in mice. ConclusionsOur data suggest that RS4651 alleviates BLM-induced pulmonary fibrosis in mice by inhibiting the TGF-β1/SMAD signalling pathway.

Alpinetin exerts anti-inflammatory, anti-oxidative and anti-angiogenic effects through activating the Nrf2 pathway and inhibiting NLRP3 pathway in carbon tetrachloride-induced liver fibrosis

Alpinetin is the major active ingredient of Alpiniakatsumadai Hayata. As a kind of novel plant-derived flavonoid, alpinetin has shown potent hepatoprotective effect against many liver diseases such as non-alcoholic fatty liver and lipopolysaccharide/d-Galactosamine-induced liver injury. However, its roles in liver fibrosis remain to be determined. The aim of the current study was to investigate the effect of alpinetin in mice with carbon tetrachloride (CCl4)-induced liver fibrosis, and to elucidate the underlying mechanisms of action. Alpinetin ameliorated the CCl4-induced liver injury and fibrosis in mice, as shown by decreased collagen deposition and the decreased expression of liver fibrosis marker proteins. Alpinetin suppressed the inflammation and oxidative stress in fibrotic livers of mice, as evidenced by decreased levels of proinflammatory factors, the decreased reactive oxygen species (ROS) and malondialdehyde (MDA) levels, and the increased activities of antioxidant enzymes. In addition, alpinetin attenuated the angiogenesis in fibrotic livers of the test animals. Mechanistically, alpinetin inhibited the CCl4-induced expression of NLRP3, ASC, cleaved caspase-1, mature (cleaved-) IL-1β, and IL-18 in livers of mice. Furthermore, alpinetin resulted in an increased in the nuclear expression and a decrease in the cytoplasmic expression of Nrf2, as well as increased protein expression of downstream target enzymes, GCLC, HO-1, NQO1, and GCLM, thus exerting the antioxidant effect. Overall, these findings suggested that the anti-fibrotic effect of alpinetin can be attributed to the inhibition of NLRP3-mediated anti-inflammatory activities and Nrf2-mediated anti-oxidative activities, in addition to the decrement of hepatic angiogenesis.

Effect of Resistance Exercise and Liposomal Vitamin C on Some Factors of Mitochondrial Dynamics and Biogenesis

Objective: Aging is associated with some changes in the liver function including increased collagen deposition and decreased mitochondrial function. This study aims to evaluate the effect of resistance exercise and vitamin C intake on the expression of Peroxisome Proliferator-Activated Receptor-Γ Coactivator-1α (PGC-1α) and Mitofusin 1 (MFn1) genes and collagen deposition in older rats. Methods: This is an experimental study conducted on 25 male Wistar rats aged 24 weeks and weighted 280-320 g. They were randomly divided into five groups of young control, older exercise, older vitamin C, older exercise + vitamin C, and older control. In the supplementation groups, rats were given liposomal vitamin C (100 mg/kg per body weight) by gavage daily. One-way ANOVA was used to examine the difference between the groups and Tukey’s post hoc test was used to determine the location of group differences. For all analyses, the significance level was set at 0.05. Results: Aging significantly reduced the expression of PGC-1α and MFn1 and increased collagen deposition in the liver tissue of rats (P=0.001). In the older exercise + vitamin C group, a significant increase in PGC-1α expression was observed compared to the older control group (P=0.001), but there was no significant changes in MFn1 expression. A significant decrease in collagen deposition was reported in the older exercise, older vitamin C, and older exercise + vitamin C groups compared to the older control group (P=0.001). Conclusion: Resistance exercise combined with vitamin C intake reduces collagen deposition in liver tissue and increases PGC-1α expression in older rats.

The Potential Safe Antifibrotic Effect of Stem Cell Conditioned Medium and Nilotinib Combined Therapy by Selective Elimination of Rat Activated HSCs.

Hepatic fibrosis is a progressive disease with serious clinical complications that arise from abnormal propagation and activation of multiple inflammatory pathways. Nilotinib is an oral tyrosine kinase inhibitor with antifibrotic activity. Mesenchymal stem cells (MSCs) are blank cells and can differentiate into specific cell types. They have the potential to repair and regenerate cells. MSCs have a special paracrine fashion where they produce special exosomes, microvesicles, and cytokines like IL-6, transforming growth factor-beta (TGF-β), and HGF as well as hepatic stellate cell suppressors. This paracrine fashion can decrease collagen deposition, enhance antifibrotic, anti-inflammatory, and angiogenic activity in vitro and in vivo. In our study, the rat's hepatic stellate cells (HSCs) in addition to different normal cell lines were treated with Nilotinib alone and in combination with liver mesenchymal stem cells conditioned medium (LMSCs-CM) for 24 h. Mono and combined therapy antifibrotic and cytotoxicity effects were evaluated using different parameters including α-SMA, cytochrome c, P53 expression, collagen deposition, DNA content, oxidative stress parameters, cell viability, and apoptosis by flow cytometry analysis. Our results showed that Nilotinib and LMSCs-CM in combination had a significantly potent antifibrotic and anti-inflammatory effect on activated hepatic stellate cells than Nilotinib alone; otherwise, this combination showed the best safety with minimal cytotoxicity on different normal cell lines.

Transplantation of human amniotic epithelial cells promotes morphological and functional regeneration in a rat uterine scar model

BackgroundCesarean scar defect (CSD) is characterized by the presence of fibrotic tissue and decreased muscular density which is induced by cesarean section. Serious CSD may eventually result in infertility or obstetrical complications. Human amniotic epithelial cells (hAECs) have shown great promise in tissue regeneration. This study aims to investigate whether hAEC transplantation has the therapeutic effects on the rat uterine scar following full-thickness injury.MethodsA rat uterine scar model was established by excising the full-thickness uterine wall of about 1.0 cm in length and 1/2–2/3 of the total circumference in width. At day 30 post-surgery, hAECs were transplanted into the uterine scar. At day 30 and 60 post-transplantation, hematoxylin and eosin (H&E) staining, Masson staining, and IHC staining for vWF, VEGFA, α-SMA, and MMP-8 were performed to evaluate the regeneration of the scarred uterus and the underlying mechanism. Pregnancy outcomes were assessed at day 60 after hAEC transplantation. Finally, hAECs were incubated with hydrogen peroxide to verify the paracrine effect of hAECs.ResultsCollagen deposition, thin myometrium, and injured endometrium were observed in the rat uterine scar model. After hAEC transplantation, collagen deposition in the uterine scar decreased, and myometrial and endometrial recovery was facilitated. hAEC transplantation also increased the fetus number implanted within the scarred area. Moreover, we found hAECs promoted angiogenesis via upregulation of VEGFA and decreased collagen deposition by upregulating MMP-8 in the uterine scar. The in vitro studies further demonstrated an increase in the expression level of MMP-8 in hAECs cultured with hydrogen peroxide.ConclusionsThese results suggested that hAEC transplantation may be efficacious in the functional repair and collagen degradation of uterine scars, which provides a new therapeutic strategy to CSD.

Cerium oxide nanoparticle delivery of microRNA-146a for local treatment of acute lung injury

Acute respiratory distress syndrome (ARDS) is a devastating pulmonary disease with significant in-hospital mortality and is the leading cause of death in COVID-19 patients. Excessive leukocyte recruitment, unregulated inflammation, and resultant fibrosis contribute to poor ARDS outcomes. Nanoparticle technology with cerium oxide nanoparticles (CNP) offers a mechanism by which unstable therapeutics such as the anti-inflammatory microRNA-146a can be locally delivered to the injured lung without systemic uptake. In this study, we evaluated the potential of the radical scavenging CNP conjugated to microRNA-146a (termed CNP-miR146a) in preventing acute lung injury (ALI) following exposure to bleomycin. We have found that intratracheal delivery of CNP-miR146a increases pulmonary levels of miR146a without systemic increases, and prevents ALI by altering leukocyte recruitment, reducing inflammation and oxidative stress, and decreasing collagen deposition, ultimately improving pulmonary biomechanics.