CCl4-induced Liver Cirrhosis Research Articles

Combination of omics, bioinformatics, molecular docking, and experimental validation to elucidate the hepatoprotective effects, mechanisms, and active compounds of Shandougen.

Omics, bioinformatics, molecular docking, and experimental validation were used to elucidate the hepatoprotective effects, mechanisms, and active compounds of Shandougen (SDG) based on the biolabel-led research pattern. Integrated omics were used to explore the biolabels of SDG intervention in liver tissue. Subsequently, bioinformatics and molecular docking were applied to topologically analyze its therapeutic effects, mechanisms, and active compounds based on biolabels. Finally, an animal model was used to verify the biolabel analysis results. Omics, bioinformatics, and molecular docking revealed that SDG may exert therapeutic effects on liver diseases in the multicompound and multitarget synergistic modes, especially liver cirrhosis. In the validation experiment, SDG and its active compounds (betulinic acid and gallic acid) significantly improved the liver histopathological damage in the CCl4-induced liver cirrhosis model. Meanwhile, they also produced significant inhibitory effects on the focal adhesion pathway (integrin alpha-1, myosin regulatory light chain 2, laminin subunit gamma-1, etc.) and alleviated the associated pathological processes: focal adhesion (focal adhesion kinase 1)-extracellular matrix (collagen alpha-1(IV) chain, collagen alpha-1(VI) chain, and collagen alpha-2(VI) chain) dysfunction, carcinogenesis (alpha-fetoprotein, NH3, and acetylcholinesterase), inflammation (tumor necrosis factor alpha, interleukin-1 [IL-1], IL-6, and IL-10), and oxidative stress (reactive oxygen species, malonaldehyde, and superoxide dismutase). This study provides new evidence and insights for the hepatoprotective effects, mechanisms, and active compounds of SDG.

Activated Natural Killer Cell Inoculation Alleviates Fibrotic Liver Pathology in a Carbon Tetrachloride-Induced Liver Cirrhosis Mouse Model

Activated hepatic stellate cells (HSCs) play a detrimental role in liver fibrosis progression. Natural killer (NK) cells are known to selectively recognize abnormal or transformed cells via their receptor activation and induce target cell apoptosis and, therefore, can be used as a potential therapeutic strategy for liver cirrhosis. In this study, we examined the therapeutic effects of NK cells in the carbon tetrachloride (CCl4)-induced liver cirrhosis mouse model. NK cells were isolated from the mouse spleen and expanded in the cytokine-stimulated culture medium. Natural killer group 2, member D (NKG2D)-positive NK cells were significantly increased after a week of expansion in culture. The intravenous injection of NK cells significantly alleviated liver cirrhosis by reducing collagen deposition, HSC marker activation, and macrophage infiltration. For in vivo imaging, NK cells were isolated from codon-optimized luciferase-expressing transgenic mice. Luciferase-expressing NK cells were expanded, activated and administrated to the mouse model to track them. Bioluminescence images showed increased accumulation of the intravenously inoculated NK cells in the cirrhotic liver of the recipient mouse. In addition, we conducted QuantSeq 3' mRNA sequencing-based transcriptomic analysis. From the transcriptomic analysis, 33 downregulated genes in the extracellular matrix (ECM) and 41 downregulated genes involved in the inflammatory response were observed in the NK cell-treated cirrhotic liver tissues from the 1532 differentially expressed genes (DEGs). This result indicated that the repetitive administration of NK cells alleviated the pathology of liver fibrosis in the CCl4-induced liver cirrhosis mouse model via anti-fibrotic and anti-inflammatory mechanisms. Taken together, our research demonstrated that NK cells could have therapeutic effects in a CCl4-induced liver cirrhosis mouse model. In particular, it was elucidated that extracellular matrix genes and inflammatory response genes, which were mainly affected after NK cell treatment, could be potential targets.

Pharmacokinetics and Therapeutic Potential of Teucrium polium against Liver Damage Associated Hepatotoxicity and Oxidative Injury in Rats: Computational, Biochemical and Histological Studies.

This study investigated the druggability, pharmacokinetics and ethyl acetate extract of Teucrium polium (EA T. polium) and the protective effect against carbon tetrachloride (CCl4) induced liver cirrhosis in rats. The total antioxidant capacity (TAC) and scavenging activity of the extract were examined. The in vivo protective study was based on the use of an animal model of CCl4-induced liver cirrhosis. Four groups of rats have been used: Group I: control rats; Group II: received CCl4 in olive oil (0.5 mL/kg); Group III: received the EA T. polium (25 mg/kg) of pretreatment for seven days by gavage then CCl4 in olive oil by gavage for 15 days. Group IV: received the EA of T. polium for seven days (25 mg/kg). EA T. polium was found to possess significant antioxidant capacity. CCl4 caused a hepatotoxicity associated increase in both levels of AST and ALT, which were reduced back to normal values following EA T. polium pretreatment. Hepatotoxicity associated structural modifications of liver tissues and increase in thiobarbituric acid reactive substances (TBARS), conjugated dienes (CD) and carbonyl proteins (CP), associated decreases in several assessed antioxidant enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT). The in vivo findings on the protective effect of T. polium were supported by its druggability, its pharmacokinetic properties and molecular docking assays. These results confirm the modulatory antioxidant and hepatoprotective potential of T. polium in this experimental liver cirrhosis model. T. polium phytochemicals are good candidates for further pharmaceutical explorations and drug design.

Differences of core genes in liver fibrosis and hepatocellular carcinoma: Evidence from integrated bioinformatics and immunohistochemical analysis.

BACKGROUNDLiver fibrosis and hepatocellular carcinoma (HCC) are common adverse consequences of chronic liver injury. The interaction of various risk factors may cause them to happen. Identification of specific biomarkers is of great significance for understanding the occurrence, development mechanisms, and determining the novel tools for diagnosis and treatment of both liver fibrosis and HCC.AIMTo identify liver fibrosis-related core genes, we analyzed the differential expression pattern of core genes in liver fibrosis and HCC.METHODSGene expression profiles of three datasets, GSE14323, GSE36411, and GSE89377, obtained from the Gene Expression Omnibus (GEO) database, were analyzed, and differentially expressed genes (DEGs) between patients with liver cirrhosis and healthy controls were identified by screening via R software packages and online tool for Venn diagrams. The WebGestalt online tool was used to identify DEGs enriched in biological processes, molecular functions, cellular components, and Kyoto Encyclopedia of Genes and Genomes pathways. The protein–protein interactions of DEGs were visualized using Cytoscape with STRING. Next, the expression pattern of core genes was analyzed using Western blot and immunohistochemistry in a carbon tetrachloride (CCl4)-induced liver cirrhosis mouse model and in patient liver samples. Finally, Kaplan-Meier curves were constructed using the Kaplan-Meier plotter online server. RESULTSForty-five DEGs (43 upregulated and 2 downregulated genes) associated with liver cirrhosis were identified from three GEO datasets. Ten hub genes were identified, which were upregulated in liver cirrhosis. Western blot and immunohistochemical analyses of the three core genes, decorin (DCN), dermatopontin (DPT), and SRY-box transcription factor 9 (SOX9), revealed that they were highly expressed in the CCl4-induced liver cirrhosis mouse model. The expression levels of DCN and SOX 9 were positively correlated with the degree of fibrosis, and SOX 9 level in HCC patients was significantly higher than that in fibrosis patients. However, high expression of DPT was observed only in patients with liver fibrosis, and its expression in HCC was low. The gene expression profiling interactive analysis server (GEPIA) showed that SOX9 was significantly upregulated whereas DCN and DPT were significantly downregulated in patients with HCC. In addition, the Kaplan-Meier curves showed that HCC patients with higher SOX9 expression had significantly lower 5-year survival rate, while patients with higher expression of DCN or DPT had significantly higher 5-year survival rates. CONCLUSIONThe expression levels of DCN, DPT, and SOX9 were positively correlated with the degree of liver fibrosis but showed different correlations with the 5-year survival rates of HCC patients.

Comprehensive Analysis of the Differentially Expressed Transcriptome with ceRNA Networks in a Mouse Model of Liver Cirrhosis

Background: Hepatic cirrhosis is the consequence of various chronic liver disease for which there is no curative treatment. In this study, based on RNA sequencing (RNA-seq) and subsequent bioinformatic analysis, we aim to explore the biological function of non-coding RNAs (ncRNAs) in hepatic cirrhosis. Methods: The hepatic cirrhosis models were induced by the intraperitoneal injection of carbon tetrachloride (CCl4). The transcriptome profile was aquired by RNA-seq, of which result was verified by quantitative real-time PCR (qRT-PCR). The competing endogenous RNA (ceRNA) networks were visualized by Cytoscape software. The enrichment analyses of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were conducted. Results: The differentially expressed transcript of liver cirrhosis is consist of 2369 mRNAs, 374 lncRNAs, 91 circRNAs and 242 miRNAs (|log2(fold change)|≥1 and P<0.05). The RNA-seq results were highly consistent with qRT-PCR validation of DEGs (four upregulated and four down-regulated, including ENSMUSG00000047517, ENSMUST00000217449, novel-circ-001366, miR-383-5p, ENSMUSG00000078683, ENSMUST00000148206, novel-circ-002669 and miR-216a-5p). Based on ceRNA theory, a circRNA-lncRNA co-regulated ceRNA network was established. Enrichment analysis revealed the potential key regulatory process during the liver cirrhosis progression. Conclusion: In conclusion, the present study comprehensively analyzed differentially expressed transcripts in CCl4-induced liver cirrhosis. Our findings explored the gene signatures for liver cirrhosis’ diagnosis and precise treatment.

Stem Cells From Human Exfoliated Deciduous Teeth Alleviate Liver Cirrhosis via Inhibition of Gasdermin D-Executed Hepatocyte Pyroptosis.

Liver cirrhosis represents a type of end-stage liver disease with few effective therapies, which was characterized by damaged functional liver tissue due to long-term inflammation. Gasdermin D (GSDMD)-executed programmed necrosis is reported to be involved in inflammation. However, the role of GSDMD in liver cirrhosis remains unclear. In this study, we used a CCl4-induced cirrhosis model and found stem cells from human exfoliated deciduous teeth (SHED) infusion showed profound therapeutic effects for liver cirrhosis. Mechanistically, NLRP3 inflammasome-activated GSDMD and its pyroptosis were upregulated in liver cirrhosis, while SHED infusion could suppress the expression of GSDMD and Caspase-1, resulting in reduced hepatocyte pyroptosis and inflammatory cytokine IL-1β release. Consistently, SHED could inhibit the elevated expression of NLRP3, GSDMD and Caspase-1 induced by CCl4 treatment in vitro co-culture system, which was mediated by decreasing reactive oxygen species (ROS) generation. Moreover, the pyroptosis inhibitor disulfiram showed similar therapeutic effects for liver cirrhosis as SHED. In conclusion, SHED alleviates CCl4-induced liver cirrhosis via inhibition of hepatocytes pyroptosis. Our findings could provide a potential treatment strategy and novel target for liver cirrhosis.

Postprandial Glycogen Content Is Increased in the Hepatocytes of Human and Rat Cirrhotic Liver

Chronic hepatitises of various etiologies are widespread liver diseases in humans. Their final stage, liver cirrhosis (LC), is considered to be one of the main causes of hepatocellular carcinoma (HCC). About 80–90% of all HCC cases develop in LC patients, which suggests that cirrhotic conditions play a crucial role in the process of hepatocarcinogenesis. Carbohydrate metabolism in LC undergoes profound disturbances characterized by altered glycogen metabolism. Unfortunately, data on the glycogen content in LC are few and contradictory. In this study, the material was obtained from liver biopsies of patients with LC of viral and alcohol etiology and from the liver tissue of rats with CCl4-induced LC. The activity of glycogen phosphorylase (GP), glycogen synthase (GS), and glucose-6-phosphatase (G6Pase) was investigated in human and rat liver tissue by biochemical methods. Total glycogen and its labile and stable fractions were measured in isolated individual hepatocytes, using the cytofluorometry technique of PAS reaction in situ. The development of LC in human and rat liver was accompanied by an increase in fibrous tissue (20- and 8.8-fold), an increase in the dry mass of hepatocytes (by 25.6% and 23.7%), and a decrease in the number of hepatocytes (by 50% and 28%), respectively. The rearrangement of the liver parenchyma was combined with changes in glycogen metabolism. The present study showed a significant increase in the glycogen content in the hepatocytes of the human and the rat cirrhotic liver, by 255% and 210%, respectively. An increased glycogen content in cells of the cirrhotic liver can be explained by a decrease in glycogenolysis due to a decreased activity of G6Pase and GP.

Hepatoprotective effect of bone marrow-derived mesenchymal stromal cells in CCl4-induced liver cirrhosis

Bone marrow mesenchymal stromal cells (BM-MSCs) are multipotent stem cells which are ideal candidates for use in regenerative medicine. The objectives of this study were to evaluate the hepatoprotective effect of BM-MSC and its combination treatment with silymarin in carbon tetrachloride (CCl4)-induced liver cirrhosis animal model and to investigate whether tail vein or portal vein infusion was the ideal route for BM-MSC transplantation. 36 female Wistar rats were randomly divided into six groups (n = 6): Group 1 (normal control), Group 2 (received only CCl4, disease model), Group 3 (CCl4 + BM-MSCs through tail vein), Group 4 (CCl4 + BM-MSCs through portal vein), Group 5 (CCl4 + silymarin), Group 6 (CCl4 + BM-MSCs + silymarin). On the 21st day after treatment, blood samples were collected for biochemical estimations. After the experiment, the rats were sacrificed. Liver was dissected out and processed for histopathology and scanning electron microscopy studies. Liver enzyme and marker analysis, histopathological studies indicated that the combination of BM-MSCs and silymarin was effective in treating liver cirrhosis. Transplanted BM-MSCs in combination with silymarin ameliorated the liver tissue damage through their immunoregulatory activities. Among the two routes, the intravenous administration of cells through the tail vein was found to be more effective and safe.

Regenerative effect of mesenteric fat stem cells on ccl4-induced liver cirrhosis, an experimental study

BackgroundLiver cirrhosis is a chronic disease in which normal liver tissue is replaced by fibrous tissue, leads to liver malfunction. Although transplantation is the most certain cure, stem cell therapies are shedding light on efforts to regenerate cirrhotic liver. The purpose of this study was to evaluate the regenerative potential of mesenteric fat stem cells in CCL4-induced liver cirrhosis in an animal model. MethodsThirty rats were treated with the mixture of CCL4 and olive oil intraperitoneally by a dose of 0.2 ml (0.1 ml CCL4 and 0.1 ml olive oil) every other day for 16 weeks till cirrhosis signs appeared. Fifteen rats were randomly selected as control group. Others treated by mesenteric fat derived mesenchymal stem cells transferred into the liver parenchyma. ResultsAfter 5 weeks, rats received stem cells had improved clinically by increased movements, appetite, improvement in overall behavior and decreased abdomen size. Histopathologically, liver cells showed state of regeneration and forming new colonies. ConclusionLiver cirrhosis was induced. The mesenchymal stem cells derived from mesenteric adipose tissue could improve hepatic status of the rats, as cirrhotic livers were regenerated back into normal appearing parenchyma. Rats’ clinical behavior also reached healthy status.

Mesenchymal stem cell-based Smad7 gene therapy for experimental liver cirrhosis

BackgroundBone mesenchymal stem cells (MSCs) can promote liver regeneration and inhibit inflammation and hepatic fibrosis. MSCs also can serve as a vehicle for gene therapy. Smad7 is an essential negative regulatory gene in the TGF-β1/Smad signalling pathway. Activation of TGF-β1/Smad signalling accelerates liver inflammation and fibrosis; we therefore hypothesized that MSCs overexpressing the Smad7 gene might be a new cell therapy approach for treating liver fibrosis via the inhibition of TGF-β1/Smad signalling.MethodsMSCs were isolated from 6-week-old Wistar rats and transduced with the Smad7 gene using a lentivirus vector. Liver cirrhosis was induced by subcutaneous injection of carbon tetrachloride (CCl4) for 8 weeks. The rats with established liver cirrhosis were treated with Smad7-MSCs by direct injection of cells into the main lobes of the liver. The expression of Smad7, Smad2/3 and fibrosis biomarkers or extracellular matrix proteins and histopathological change were assessed by quantitative PCR, ELISA and Western blotting and staining.ResultsThe mRNA and protein level of Smad7 in the recipient liver and serum were increased after treating with Smad-MSCs for 7 and 21 days (P < 0.001). The serum levels of collagen I and III and collagenase I and III were significantly (P < 0.001) reduced after the treatment with Smad7-MSCs. The mRNA levels of TGF-β1, TGFBR1, α-SMA, TIMP-1, laminin and hyaluronic acid were decreased (P < 0.001), while MMP-1 increased (P < 0.001). The liver fibrosis score and liver function were significantly alleviated after the cell therapy.ConclusionsThe findings suggest that the MSC therapy with Smad7-MSCs is effective in the treatment of liver fibrosis in the CCl4-induced liver cirrhosis model. Inhibition of TGF-β1 signalling pathway by enhancement of Smad-7 expression could be a feasible cell therapy approach to mitigate liver cirrhosis.

P0520EFFECT OF HUMAN UMBILICAL CORD MESENCHYMAL STEM CELLS ON SECONDARY ACUTE KIDNEY INJURY IN RATS WITH LIVER CIRRHOSIS

Abstract Background and Aims Acute kidney injury( AKI) is one of the most common complications of decompensated cirrhosis, and it primarily presents as a sharp decrease in glomerular filtration rate, rapid increase in serum creatinine( SCr) and urea nitrogen. And the search for specific and safe treatment has been a research hot spot in recent years. In this article, the effect of human umbilical cord mesenchymal stem cells on carbon tetrachloride (CCl4)-induced liver fibrosis (HF) in rats with acute kidney injury and the possible mechanism are investigated. Method Human umbilical cord blood mesenchymal stem cells were sub-cultured by adherent method, and the cells were identified by morphological observation, cell phenotypic analysis and multi-directional differentiation potential analysis methods. WASTA rats were randomly divided into control group, cirrhosis model group and treatment group, with 10 rats in each group. Model group and treatment group were injected with CCl4-olive oil (1:1) solution 3 mL·kg -1, and the control group was given the same amount of olive oil for intervention, twice a week for 8 weeks. Rats in treatment group were administrated wth Human umbilical cord mesenchymal stem cells (2 × 109 /L) via the tail vein at the 5th week after injection of CCl4-olive oil solution, but the other rats were injected with 0.9% normal saline, once a week for 6 weeks. After the intervention, Serum, kidneys and 24 hours urine of rats in each group were collected, which were applied for a detection of serum creatinine and urea nitrogen, malondialdehyde (MDA), NO content and superoxide dismutase (SOD), as well as renal pathological examination. Results 1.In vitro, umbilical cord blood mesenchymal stem cells was passaged to the third generation, and the morphology was uniform and spiraled. Phenotypic analysis showed that the positive rates of stem cell markers CD29, CD44 and CD105 were all greater than 95%, the positive rate of HLA-DR (graft-versus-host disease-associated factor) less than 10%, and the positive rate of CD34 and CD45 lower than 20% (Figure 1). 2. Compared with the cirrhotic model group, MDA content of serum and kidney in model group significantly decreased under the effect of mesenchymal stem cell (p &amp;lt;0.01) (Table 1). 3. The normal group had normal liver tissue structure, ordered liver cells, no hepatic edema, and no lesions. In the model group, large-area lesions, including edema of liver cells, rupture of cell membranes, and infiltration of inflammatory cells, had appeared. Compared with the model group, Hepatocellular necrosis, edema, and inflammatory cell infiltration were significantly improved after transplanting Human umbilical cord mesenchymal stem cells (Figure 2). 4.In the model group, the rat renal tubules disappeared and the lumen was disordered. After injection of Human umbilical cord mesenchymal stem cells, renal tubular and renal interstitial damage is improved and the thickening of glomerular basement membrane is reduced (Figure 3). Conclusion In CCl4-induced liver cirrhosis model rats, human umbilical cord mesenchymal stem cells can protect the kidney by reducing free radicals and cellular lipid peroxidation in vivo.

Obeticholic acid prevents carbon tetrachloride-induced liver fibrosis through interaction between farnesoid X receptor and Smad3

Liver fibrosis results from sustained liver injury and is characterized by inflammation, hepatic stellate cell (HSC) activation, extracellular matrix (ECM) accumulation and liver structure destruction. The Farnesoid-X receptor (FXR) antagonizes toxic liver injury and fibrosis, yet the mechanism in liver fibrosis remains unclear. We investigated the effects of FXR agonist obeticholic acid (OCA) on liver fibrosis in mice. Mice were injected with carbon tetrachloride (CCl4) for 3 weeks or 6 weeks to induce liver fibrosis. OCA (5 mg/kg) or PBS is administered daily during CCl4-treatment. At sacrifice, biochemical parameters and fibrosis were assessed. Pretreatment with OCA alleviated hepatic injury in 6 weeks group but not in 3 weeks group of CCl4 liver cirrhosis. At same time, pretreatment with OCA exhibit a dramatic protection of liver fibrosis in both 3 weeks group and 6 weeks group. Further experiments found that OCA pretreatment inhibited α-SMA expression and the activation of hepatic pSmad3 in 3 weeks group and 6 weeks group of CCl4-induced liver cirrhosis. Moreover, OCA activated FXR nuclear translocation and increased the interaction between liver FXR and pSmad3. This led to the discovery of a novel role for FXR in regulating fibrosis through interaction with pSmad3. Our data suggest that CCl4-induced liver fibrosis is protected by OCA through interaction between farnesoid X receptor and Smad3.

Nutrient mixture from germinated legumes: Enhanced medicinal value with herbs-attenuated liver cirrhosis.

Among various food processing strategies, germination and dehulling enhance the nutritional content of the food, and the addition of herbs to this could improve the medicinal value. The milled powders of germinatedMacrotyloma uniflorum(horse gram) andVigna radiata(green gram) were used to make the nutrient mixture. Further,Curcuma longa(turmeric) andTrigonella foenum graecum(fenugreek) were used to improve its medicinal value. The prepared nutrient mixture has high nutritional value, antioxidant potential, and reduced antinutrient factors. Supplementation of nutrient mixture reduced oxidative stress-mediated hepatocyte injury on the CCl4 -induced liver cirrhosis model. Further, histological examination (H&E and Sirius red), matrix metalloproteinase gelatin zymography, and Western blot revealed the management of hepatic stellate cells in an inactive stage thereby reduced cirrhosis. These findings conclude that the supplementation of nutrient mixture formulation protected and effectively prevented liver cirrhosis. PRACTICAL APPLICATIONS: This study has a good impact on nutritional therapy for liver diseases. Many of the chronic liver diseases are associated with severe malnutrition and hypoalbuminemia, which further worsens the condition. This study would emphasize the nutritional therapy to treat such imbalance and enriching the medicinal value of nutrition mixture with herbs could target different pathophysiological changes and provide better defense in liver disease patients. Since this nutrient mixture is from common edible natural resources, it could reach the pharmaceutical industry's attention to the highest production and marketing.

Effects of Hematopoietic Stem Cells on Liver Cirrhosis of Albino Rats

AbstractBackground: Liver cirrhosis is a common serious disease in Egypt due to high incidence rate of infectious liver diseases; hepatitis. Application of Hematopoietic Stem Cells (HSCs) may provide a hope for patients with liver cirrhosis as it is not only safe but also to avoid hazardous complications and immunosuppressive drug following liver transplantation.Aim of Study: Evaluation of the effects of CD34+ hemat-opoietic stem cells in regeneration of hepatic tissue as a new therapy for cirrhotic liver.Material and Methods: Forty five male albino rats were divided into four groups: Control group: Include 18 rats injected with olive oil subcutaneously, (carbon tetrachloride treated group): 9 rats received carbon tetrachloride subcuta-neously, (carbon tetrachloride/hematopiotic stem cells treated group): 9 rats injected with hematopiotic stem cells after induction of liver cirrhosis by carbon tetrachloride and (carbon tetrachloride/fibroblast treated group): 9 rats injected with fibroblast cells after carbon tetrachloride induced liver cirrho-sis. For histological and biochemical study of liver and blood samples were taken from all rats.Results: Rats of carbon tetrachloride treated group & carbon tetrachloride/fibroblast treated group indicated significant changes in liver histological structures accompanied with significant decline of liver function tests. Injection of HSC showed marked improvement in both histological picture and biochemical profile.Conclusion: This study indicates the effectiveness of human CD34+ hematopoietic stem cells in amelioration of CCl4 induced liver cirrhosis in rats. So, treatment with he-matopoietic stem cells therapy can be considered a new hope for cirrhotic patients, especially in Egypt.

Bone Morphogenetic Protein 9 Is a Mechanistic Biomarker of Portopulmonary Hypertension.

BMP9 (bone morphogenetic protein 9) is a circulating endothelial quiescence factor with protective effects in pulmonary arterial hypertension (PAH). Loss-of-function mutations in BMP9, its receptors, and downstream effectors have been reported in heritable PAH. To determine how an acquired deficiency of BMP9 signaling might contribute to PAH. Plasma levels of BMP9 and antagonist soluble endoglin were measured in group 1 PAH, group 2 and 3 pulmonary hypertension (PH), and in patients with severe liver disease without PAH. BMP9 levels were markedly lower in portopulmonary hypertension (PoPH) versus healthy control subjects, or other etiologies of PAH or PH; distinguished PoPH from patients with liver disease without PAH; and were an independent predictor of transplant-free survival. BMP9 levels were decreased in mice with PH associated with CCl4-induced portal hypertension and liver cirrhosis, but were normal in other rodent models of PH. Administration of ALK1-Fc, a BMP9 ligand trap consisting of the activin receptor-like kinase-1 extracellular domain, exacerbated PH and pulmonary vascular remodeling in mice treated with hypoxia versus hypoxia alone. BMP9 is a sensitive and specific biomarker of PoPH, predicting transplant-free survival and the presence of PAH in liver disease. In rodent models, acquired deficiency of BMP9 signaling can predispose to or exacerbate PH, providing a possible mechanistic link between PoPH and heritable PAH. These findings describe a novel experimental model of severe PH that provides insight into the synergy between pulmonary vascular injury and diminished BMP9 signaling in the pathogenesis of PAH.

Engineered Relaxin as theranostic nanomedicine to diagnose and ameliorate liver cirrhosis

Hepatic cirrhosis is a growing health problem with increasing mortality worldwide. So far, there is a lack of early diagnosis and no clinical therapy is approved for the treatment. In this study, we developed a novel theranostic nanomedicine by targeting relaxin (RLX) that is known to possess potent anti-fibrotic properties but simultaneously has poor pharmacokinetics and detrimental off-target effects. We conjugated RLX to PEGylated superparamagnetic iron-oxide nanoparticles (RLX-SPIONs) and examined hepatic stellate cells (HSCs) specific binding/uptake. Thereafter, we assessed the therapeutic efficacy of RLX-SPIONs on human HSCs in vitro and in vivo in CCl4-induced liver cirrhosis mouse model. RLX-SPIONs showed specific binding and uptake in TGFβ-activated HSCs, and inhibited TGFβ-induced HSCs differentiation, migration and contraction. In vivo, RLX-SPIONs strongly attenuated cirrhosis and showed enhanced contrast in MR imaging. Altogether, this study presents RLX-SPIONs as a novel theranostic nanomedicine that provides new opportunities for the diagnosis and treatment of liver cirrhosis.

Regulation of CCl4-induced liver cirrhosis by hepatically differentiated human dental pulp stem cells.

Liver transplantation is the most effective treatment for treating liver cirrhosis. However, a limited number of donors, graft rejection, and other complications can undermine transplant success. It is considered that cell transplantation is an alternative approach of liver transplantation. We previously developed a protocol for hepatic differentiation of cluster of differentiation 117+ stem cells isolated from human exfoliated deciduous tooth pulp (SHEDs) under hydrogen sulfide exposure. These cells showed excellent hepatic function. Here, we investigated whether hepatocyte-like cell transplantation is effective for treating carbon tetrachloride (CCl4)-induced liver cirrhosis. SHEDs were hepatically differentiated, which was confirmed via immunological analyses and albumin concentration determination in the medium. Rats were intraperitoneally injected with CCl4 for and the differentiated cells were injected into rat spleen. Histopathological and immunohistochemical analyses were performed. Liver functions were serologically and pathologically determined. Quantitative real-time-polymerase chain reaction was implemented to clarify the treatment procedure of liver cirrhosis. In vitro-differentiated hepatocyte-like cells were positive for all examined hepatic markers. SHED-derived hepatocyte transplantation eliminated liver fibrosis and restored liver structure in rats. Liver immunohistochemical analyses showed the presence of human-specific hepatic markers, i.e., a large amount of human hepatic cells were very active in the liver and spleen. Serological tests revealed significant liver function recovery in the transplantation group. Expression of genes promoting fibrosis increased after cirrhosis induction but was suppressed after transplantation. Our results suggest that xenotransplantation of hepatocyte-like cells of human origin can treat cirrhosis. Moreover, cell-based therapy of chronic liver conditions may be an effective option.

Haemostatic potential of human bone marrow-derived mesenchymal stromal cells in Wistar rats with carbon tetrachloride induced liver cirrhosis.

To evaluate the haemostatic potential of human bone marrow-derived mesenchymal stromal cells (BM-MSCs) in carbon tetrachloride (CCl4) induced liver cirrhosis in Wistar rats. This was an experimental study. Liver cirrhosis was induced in adult female Wistar rats using CCl4. Rats were randomly divided into 6 groups with ten rats in each group: group 1 (normal control group), group 2 (received only CCl4), group 3 (CCl4 + low dose BM-MSCs), group 4 (CCl4 + high dose BM-MSCs), group 5 (CCl4 + silymarin), group 6 (CCl4 + high dose BM-MSCs + silymarin). Thirty days after the treatment, blood samples were collected for liver enzyme level analysis, prothrombin time test and plasma fibrinogen estimations. The rats were then sacrificed, livers were excised and used for histopathological and scanning electron microscopy (SEM) study. BM-MSCs and the combination treatment of high dose BM-MSCs and silymarin effectively decreased the prothrombin time and increased plasma fibrinogen concentration in rats with CCl4 induced liver cirrhosis. BM-MSCs treatment produces significant anti-fibrotic effect which was supported by the liver enzyme level analysis, histopathology and SEM study. Results indicate that treatment of BM-MSCs in combination with silymarin had a better haemostatic effect when compared to the administration of BM-MSCs alone.

Impact of omega-3 fatty acids on evolution of carbon tetrachloride- (CCl4) induced liver cirrhosis in mice : A histological and immunohistochemical study

hepatic architecture. Omega-3 polyunsaturated fatty acids are common antioxidant, anti-inflammatory and anti-fibrotic agentsAim: This study was conducted to investigate the potential effects of omega-3 fatty acids on experimentally induced cirrhosis.Materials and Methods: 60 animals were divided into three groups: Group I (control): 10 animals, group II: 25 animals treated with carbon tetrachloride (CCl4), and Group III : 25 animals treated with CCl4 then with omega-3 concomitant with CCl4 for additional two weeks after induction of cirrhosis. Body weight and liver weight coefficient were measured. Liver samples were taken for histological and immunohistochemical studies.Results: Omega-3 fatty acids treatment attenuated the CCl4 induced cirrhosis. Histological studies revealed that it decreased the stage of fibrosis, and the grade of the necroinflammatory changes. Immunohistochemical studies demonstrated that apoptotic changes diminished in cirrhotic livers after treatment with omega-3 fatty acids with a significant decrease in the cleaved caspase-3 index. Furthermore, there was a significant decrease in the number of macrophages, and hepatic stellate cells activation.Conclusion: Our results suggest that omega-3 fatty acids have an anti-inflammatory and anti-fibrotic effects on CCl4 induced liver cirrhosis in mice.

Quantitative Analysis of Reticular Fibers in CCl4-Induced Liver Cirrhosis in Mice

Quantitative Analysis of Reticular Fibers in CCl4-Induced Liver Cirrhosis in Mice