Xianwen Yuan

Targeted migration of mesenchymal stem cells modified with CXCR4 to acute failing liver improves liver regeneration

AIM To improve the colonization rate of transplanted mesenchymal stem cells (MSCs) in the liver and effect of MSC transplantation for acute liver failure (ALF). METHODS MSC was modified with the chemokine CXC receptor 4 (CXCR4) gene (CXCR4-MSC) or not (Null-MSC) through lentiviral transduction. The characteristics of CXCR4-MSCs and Null-MSCs were determined by real-time quantitative polymerase chain reaction, Western blotting and flow cytometry. CXCR4-MSCs and Null-MSCs were infused intravenously 24 h after administration of CCl4 in nude mice. The distribution of the MSCs, survival rates, liver function, hepatocyte regeneration and growth factors of the recipient mice were analyzed. RESULTS In vitro, CXCR4-MSCs showed better migration capability toward stromal cell-derived factor-1α and a protective effect against thioacetamide in hepatocytes. In vivo imaging showed that CXCR4-MSCs migrated to the liver in larger numbers than Null-MSCs 1 and 5 d after ALF. Higher colonization led to a longer lifetime and better liver function. Either CXCR4-MSCs or Null-MSCs exhibited a paracrine effect through secreting hepatocyte growth factor and vascular endothelial growth factor. Immunohistochemical analysis of Ki-67 showed increased cell proliferation in the damaged liver of CXCR4-MSC-treated animals. CONCLUSION Genetically modified MSCs expressing CXCR4 showed greater colonization and conferred better functional recovery in damaged liver.

The mobilization, recruitment and contribution of bone marrow-derived endothelial progenitor cells to the tumor neovascularization occur at an early stage and throughout the entire process of hepatocellular carcinoma growth.

Obvious neovascularization is a key feature of hepatocellular carcinoma (HCC) and the status of neovascularization in HCC is closely correlated with the tumor growth and patient prognosis. The actual effect of current antivascular treatment including embolization to HCC is not satisfactory. Compensatory angiogenesis is one of the primary causes responsible for failure of antiangiogenic therapy. Bone marrow-derived endothelial progenitor cells (BM-EPCs) are considered as important building blocks for adult neovascularization. However, the role of mobilized BM-EPCs in HCC remains unknown. In this study, GFP+-BM orthotropic HCC mice were established to investigate whether BM-EPCs are involved in HCC-induced neovascularization. We found that a large number of BM-EPCs were mobilized into the circulation with the development of HCC, recruited into the HCC region and incorporated into the vascular endothelium directly by differentiation into vascular endothelial cells, including sinus, capillary vessels and great vessels. Dynamic observation revealed that the mobilization and the incorporation of BM-EPCs into different types of vessels were present in early phases and throughout the whole process of HCC growth. The proportion of BM-EPCs in vessels increased gradually, from 17 to 21% with tumor growth. Moreover, injected GFP+-EPCs also specifically homed to tumor tissue and incorporated into tumor vessels directly. In this initial study, we demonstrated that BM-EPCs play a prominent role in HCC neovascularization. Blockade of BM-EPC-mediated vasculogenesis may improve the efficacy of current anti-vascularization therapy for patients with HCC.

USP39 promotes the growth of human hepatocellular carcinoma in vitro and in vivo

Ubiquitin specific protease 39 (USP39) plays an important role in mRNA splicing. In the present study, we investigated the role of USP39 in regulating the growth of hepatocellular carcinoma (HCC). We detected USP39 expression in more than 100 HCC clinical samples. The USP39 expression was significantly higher in the tumor tissues compared to the adjacent normal tissues, and was strongly associated with the pathological grade of HCC. USP39 knockdown inhibited cell proliferation and colony formation in vitro in the HepG2 cells, while upregulation of USP39 promoted tumor cell growth. FCM assay showed that USP39 knockdown led to G2/M arrest and induced apoptosis in the HepG2 cells. USP39 knockdown by shRNA inhibited xenograft tumor growth in nude mice. Moreover, USP39 knockdown led to the upregulation of p-Cdc2 and downregulation of p-Cdc25c and p-myt1, while the expression of total Cdc2, Cdc25c and myt1 was not changed in the USP39-knockdown cells. We also found that p-Cdc2 was decreased in the USP39-overexpressing cells and was upregulated in the xenografted tumors derived from the HepG2/KD cells from nude mice. Meanwhile, the expression levels of FoxM1 and its target genes PLK1 and cyclin B1 were decreased in the USP39-knockdown cells. These results suggest that USP39 may contribute to FoxM1 splicing in HCC tumor cells. Our data indicate that USP39 knockdown inhibited the growth of HCC both in vitro and in vivo through G2/M arrest, which was partly achieved via the inhibition of FoxM1 splicing.

Interleukin-10 secreted by mesenchymal stem cells attenuates acute liver failure through inhibiting pyroptosis

Recently, the benefit of mesenchymal stem cells (MSCs) as a cell‐based therapy for acute liver failure (ALF) has gained much attention, although the mechanism of action of MSCs in the treatment of ALF remains elusive. Pyroptosis is a novel form of programmed cell death with an intense inflammatory response. The aim of the present study was to explore the soluble cytokines secreted by MSCs and their therapeutic effects through inhibiting pyroptosis in ALF.

Interleukin-10 Contributes to Therapeutic Effect of Mesenchymal Stem Cells for Acute Liver Failure via Signal Transducer and Activator of Transcription 3 Signaling Pathway

Background:Mesenchymal stem cells (MSCs) transplantation has been proven to have therapeutic potential for acute liver failure (ALF). However, the mechanism remains controversial. Recently, modulation of inflammation by MSCs has been regarded as a crucial mechanism. The aim of the present study was to explore the soluble cytokines secreted by MSCs and their therapeutic effects in ALF. Methods:MSCs isolated from Sprague-Dawley rats were identified by fluorescence-activated cell sorting analysis. Conditioned medium derived from MSCs (MSCs-CM) was collected and analyzed by a cytokine microarray. MSCs and MSCs-CM were transplanted into rats with D-galactosamine-induced ALF. Liver function, survival rate, histology, and inflammatory factors were determined. Exogenous recombinant rat interleukin (IL)-10, anti-rat IL-10 antibody, and AG490 (signal transducer and activator of transcription 3 [STAT3] signaling pathway inhibitor) were administered to explore the therapeutic mechanism of MSCs-CM. Statistical analysis was performed with SPSS version 19.0, and all data were analyzed by the independent-sample t-test. Results:There are statistical differences of the survival curve between ALF+MSCs group and ALF+Dulbeccos modified Eagles medium (DMEM) group, as well as ALF+MSCs-CM group and ALF+DMEM group (all P < 0.05). Serum alanine aminotransferase (ALT) level in the ALF+MSCs and ALF+MSCs-CM groups was lower than that in the ALF+DMEM group (865.53±52.80 vs. 1709.75±372.12 U/L and 964.72±414.59 vs. 1709.75±372.12 U/L, respectively, all P < 0.05); meanwhile, serum aspartate aminotransferase (AST) level in the ALF+MSCs and ALF+MSCs-CM groups was lower than that in the ALF+DMEM group (2440.83±511.94 vs. 4234.35±807.30 U/L and 2739.83±587.33 vs. 4234.35±807.30 U/L, respectively, all P < 0.05). Furthermore, MSCs or MSCs-CM treatment significantly reduced serum interferon-&ggr; (IFN-&ggr;), IL-1&bgr;, IL-6 levels and increased serum IL-10 level compared with DMEM (all P < 0.05). Proteome profile analysis of MSCs-CM indicated the presence of anti-inflammatory factors and IL-10 was the most distinct. Blocking of IL-10 confirmed the therapeutic significance of this cytokine. Phosphorylated STAT3 was upregulated after IL-10 infusion and inhibition of STAT3 by AG490 reversed the therapeutic effect of IL-10. Conclusions:The factors released by MSCs, especially IL-10, have the potential for therapeutic recovery of ALF, and the STAT3 signaling pathway may mediate the anti-inflammatory effect of IL-10.

Bone marrow cells: Important role on neovascularization of hepatocellular carcinoma.

Background and Aim:  Present antivascular therapies including embolization to hepatocellular carcinoma (HCC) were not as satisfying as expected. The aim was to explore whether or not bone marrow cells (BMCs) played an important role on neovascularization in HCC.

MicroRNA-122 Inhibits Lipid Droplet Formation and Hepatic Triglyceride Accumulation via Yin Yang 1

Background/Aims: An increase in intracellular lipid droplet formation and hepatic triglyceride (TG) content usually results in nonalcoholic fatty liver disease. However, the mechanisms underlying the regulation of hepatic TG homeostasis remain unclear. Methods: Oil red O staining and TG measurement were performed to determine the lipid content. miRNA expression was evaluated by quantitative PCR. A luciferase assay was performed to validate the regulation of Yin Yang 1 (YY1) by microRNA (miR)-122. The effects of miR-122 expression on YY1 and its mechanisms involving the farnesoid X receptor and small heterodimer partner (FXR-SHP) pathway were evaluated by quantitative PCR and Western blot analyses. Results: miR-122 was downregulated in free fatty acid (FFA)-induced steatotic hepatocytes, and streptozotocin and high-fat diet (STZ-HFD) induced nonalcoholic steatohepatitis (NASH) in mice. Transfection of hepatocytes with miR-122 mimics before FFA induction inhibited lipid droplet formation and TG accumulation in vitro. These results were verified by overexpressing miR-122 in the livers of STZ-HFD-induced NASH mice. The 3’-untranslated region (3’UTR) of YY1 mRNA is predicted to contain an evolutionarily conserved miR-122 binding site. In silico searches, a luciferase reporter assay and quantitative PCR analysis confirmed that miR-122 directly bound to the YY1 3’UTR to negatively regulate YY1 mRNA in HepG2 and Huh7 cells. The (FXR-SHP) signaling axis, which is downstream of YY1, may play a key role in the mechanism of miR-122-regulated lipid homeostasis. YY1-FXR-SHP signaling, which is negatively regulated by FFA, was enhanced by miR-122 overexpression. This finding was also confirmed by overexpression of miR-122 in the livers of NASH mice. Conclusions: The present results indicate that miR-122 plays an important role in lipid (particularly TG) accumulation in the liver by reducing YY1 mRNA stability to upregulate FXR-SHP signaling.

Combined treatment with MSC transplantation and neutrophil depletion ameliorates D-GalN/LPS-induced acute liver failure in rats

BACKGROUND The imbalance of immunity is an important pathogenesis of acute liver failure (ALF). Neutrophils are the hallmark of acute inflammation, which have an essential role in immune regulation. Mesenchymal stem cell (MSC) transplantation is a promising therapy in ALF treatment. Recent studies indicated a considerable connection between MSCs and neutrophils in immune regulation. AIM To investigate changes in neutrophils in ALF rats after MSC transplantation, and to explore the therapeutic effect and mechanism of the combined treatment with MSC transplantation and neutrophil depletion in ALF. METHODS We employed monotherapy and the combination therapy with MSCs and anti-PMN serum in D-galactosamine (D-GalN)/lipopolysaccharides (LPS)-induced ALF rats. Rats were sacrificed at 6, 12 and 24h, respectively. Blood samples and liver tissues were collected. Hepatic injury, inflammatory cytokines (TNF-α, IL-1β and IL-10), chemokines (CXCL1 and CXCL2), the number and activity of neutrophils and animal survival were assessed at fixed times. RESULTS MSC transplantation can effectively improve the liver function of ALF rats and reduce the number and activity of neutrophils in both peripheral blood and liver. Compared with MSC transplantation alone, anti-PMN treatment and co-treatment had a better result in diminishing neutrophils. The co-treatment also exhibited a better therapeutical effect in ALF rats compared with monotherapy. In this process, the expressions of inflammatory cytokines in the liver were consistent with liver function. CONCLUSIONS The regulation of the neutrophil-related microenvironment is affected in D-GalN/LPS-induced ALF rats after MSC transplantation. The combined treatment with MSC transplantation and neutrophil depletion may have a better therapeutic effect in ALF rats.

USP39 promotes colorectal cancer growth and metastasis through the Wnt/β-catenin pathway

In the present study, we first examined the expression of USP39 protein using tissue array containing 90 colorectal cancer (CRC) tissues and 9 clinical samples, and observed that it has significantly higher expression in cancer tissues as compared to the corresponding adjacent normal tissues. Also, we tested USP39 expression level in four CRC cancer cell lines and identified that it indeed had higher expression in all these CRC cell lines. In addition, its knockdown inhibited not only the cell growth of SW480 and HT29 cells, but also the cell migration and invasion. Further analysis of its molecular mechanism suggested that the expression of four crucial proteins of Wnt/β-catenin pathway, including β-catenin, TCF4, MMP2 and MMP9 was reduced as a result of USP39 knockdown. Taken together, all these findings demonstrated that USP39 protein plays an important role in the growth and metastasis of colorectal cancer mainly through Wnt/β-catenin pathway.

Hepatic expression of Yin Yang 1 (YY1) is associated with the non-alcoholic fatty liver disease (NAFLD) progression in patients undergoing bariatric surgery

BackgroundThis study is to investigate the association between the hepatic expression of Yin Yang 1 (YY1) and the progression of non-alcoholic fatty liver disease (NAFLD) in patients undergoing bariatric surgery.MethodsObese patients undergoing bariatric surgery were included. Liver tissues were subjected to the quantitative real-time PCR, Western blot analysis, and immunohistochemical assay, to determine the expression levels of YY1.ResultsTotally 88 patients were included. According to the NAFLD activity score (NAS), these patients were divided into the control (n = 12), steatosis (n = 20), non-defining NASH (n = 38), and NASH (n = 18) groups. Significant differences in the serum glucose, insulin, ALT, AST, and HOMA-IR levels were observed among these different NAFLD groups. Hepatic YY1 expression had correlation with serum glucose, insulin, HOMA-IR, ALT, AST, triglycerides, HDL, and GGT. Immunohistochemical analysis showed that, compared with the control group, the expression levels of YY1 were significantly higher in the non-defining NASH and NASH groups. In addition, multivariate regression model showed that the serum ALT and YY1 levels were strongly associated with the NAFLD activity.ConclusionsSeveral factors are associated with NAFLD progression, including the expression of YY1. Our findings contribute to understanding of the pathogenesis of NAFLD.Trial registrationNCT03296605, registered on September 28, 2017.