Expression Of Hepatocyte Growth Factor Research Articles

Maternal and umbilical cord serum-derived exosomes enhance endothelial cell proliferation and migration.

We investigated the role of exosomes derived from maternal and umbilical cord blood in the regulation of angiogenesis. We report here that both maternal exosomes (MEs) and umbilical exosomes (UEs) significantly enhance HUVEC proliferation, migration, and tube formation. Importantly, ME-treated HUVECs (MEXs) displayed significantly increased migration, but not proliferation or tube formation, compared with UE-treated HUVECs (UEXs). We found that the expression of a subset of migration-related microRNAs (miRNAs), including miR-210-3p, miR-376c-3p, miR-151a-5p, miR-296-5p, miR-122-5p, and miR-550a-5p, among others, were significantly increased or decreased in UEs, and this altered expression was likely correlated with the differential regulation of HUVEC migration. We also found that the mRNA expression of hepatocyte growth factor (HGF) was up-regulated in MEXs and UEXs and, moreover, that inhibiting HGF partially abolished the enhanced cell migration induced by UEs. Our results suggest that both MEs and UEs greatly enhanced endothelial cell (EC) functions and differentially regulated EC migration, which was mostly attributed to the different expression profiles of exosomal miRNA. These findings highlight the importance of exosomes in the regulation of angiogenesis during pregnancy. Exosomal miRNAs, in particular, may be of great significance for the regulation of angiogenesis in maintaining normal pregnancy.-Jia, L., Zhou, X., Huang, X., Xu, X., Jia, Y., Wu, Y., Yao, J., Wu, Y., Wang, K. Maternal and umbilical cord serum-derived exosomes enhance endothelial cell proliferation and migration.

Preoperative erythropoietin treatment improves survival following major hepatic resection in a cirrhotic rat model

Major hepatic resection of a cirrhotic liver may result in a fatal clinical course. Preoperative erythropoietin (EPO) treatment has been shown to have protective properties and to stimulate liver regeneration. This study aims to investigate the effect of preoperative EPO on survival following major hepatic resection in a cirrhotic rat model. Cirrhotic liver was induced by intraperitoneal injection of thioacetamide (200mg/kg/mL) in 72 Lewis rats. Each 36 rats received EPO (1IU/g, every second day, 5 times preoperatively) or saline (control) and major hepatectomy (removal of the left and half of the median lobe) was performed. Biochemical and immunohistochemical parameters, cytokines and overall survival were compared following surgery. Rats that received preoperative EPO had decreased hepatic aspartate aminotransferase, alanine aminotransferase and interleukin (IL)-1β expression, 48hours following surgery. They had increased hepatocyte growth factor and vascular endothelial growth factor expression at 1hour, increased IL-6 expression at 24, 48 and 120hours and increased Ki-67, 120hours following surgery. Overall, survival was significantly improved among EPO-treated rats (P=0.034). Preoperative EPO treatment has a protective effect and stimulates liver regeneration, leading to improved overall survival following major hepatectomy in a cirrhotic rat model.

Polyphyllin I inhibits gastric cancer cell proliferation by downregulating the expression of fibroblast activation protein alpha (FAP) and hepatocyte growth factor (HGF) in cancer-associated fibroblasts

The aim of this study was to identify the anti-cancer mechanism of Polyphyllin I (PPI) on gastric cancer cells via its activity on cancer-associated fibroblasts (CAFs). We cultured purified gastric CAFs obtained from fresh human gastric cancer tissue and examined the effect of Polyphyllin I on CAF proliferation using a colorimetric viability assay. In addition, we established a nude mouse xenograft model to examine the effect of Polyphyllin I administration on tumorigenesis. Using Western analysis, we quantified protein expression of the CAF-derived cytokines fibroblast activation protein alpha (FAP), secreted protein acidic and cysteine rich (SPARC), stromal cell-derived factor 1 (SDF-1), hepatocyte growth factor tenascin-C (TNC), and hepatocyte growth factor (HGF) in both in vitro and in vivo models. We found that Polyphyllin I inhibits the proliferation of CAFs in a concentration-dependent manner. Following treatment with 2 μg/ml PPI for 24 h in vitro, the expression of FAP, SDF-1 and HGF protein in CAFs was significantly lower than that in the control group, but there was no significant difference in SPARC and TNC protein expression between the two groups. In the nude mouse xenograft model, the tumor inhibition rate was 45.5% when PPI was administered early and 29.4% with administration in the third week. The expression of FAP and HGF in the xenografts was significantly decreased, while the expression of SPARC, SDF-1, and TNC was largely unaltered. Altogether, these data suggest that Polyphyllin I can inhibit the proliferation of gastric cancer cells by downregulating the expression of FAP and HGF in CAFs in vivo.

MicroRNA-210 promotes angiogenesis in acute myocardial infarction.

MicroRNA-210 (miRNA-210) has been reported to be associated with angiogenesis and may serve important roles in acute myocardial infarction (AMI), which remain unclear. The present study sought to evaluate the efficacy of miRNA-210 in AMI and to examine the potential associated mechanisms. AMI models were established in Sprague-Dawley rats. The expression of miRNA-210 was upregulated via transfection with lentivirus-mediated agonists and quantitative analysis was performed using the reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Immunoblotting and RT-qPCR were separately used to detect the expression levels of hepatocyte growth factor (HGF) in heart samples, while only the protein expression level of β-myosin heavy chain (β-MHC) was assessed. The expression of HGF in human umbilical vein endothelial cells under hypoxic conditions was silenced by transfecting with small interfering RNA, as demonstrated by the determination of associated protein expression levels. The microvessel density (MVD) of the infarcted myocardium was selected to be the angiogenesis efficacy endpoint, which was evaluated by platelet endothelial cell adhesion molecule immunostaining. Markedly increased expression of HGF was observed among the AMI rats receiving miRNA-210 agonists, demonstrated via quantitative analyses using RT-qPCR or western blotting. Promotion of angiogenesis was observed with the increased MVD. Improved cardiac function in the rats was subsequently noted, as they exhibited improved left ventricular fractional shortening and left ventricular ejection fraction percentages, which may result from improved cardiac contractility indicated by attenuating the increase in β-MHC protein expression. Overexpression of miRNA-210 appeared to be an advantageous therapeutic tool for treating AMI, primarily due to its promoting effects on angiogenesis in the infarcted myocardium by stimulating HGF expression and inducing improved left ventricular remodeling, leading to improved cardiac function.

Expression of angiogenic factors in sclerosing stromal tumours of the ovary

Sclerosing stromal tumours (SSTs) are rare benign tumours and are generally observed in individuals in their teens to 20 s. Many cases are suspected as malignant tumours pre-operatively, owing to their high vascularity on diagnostic imaging. Herein, we aimed to evaluate the expressions of various angiogenic factors in SSTs. The expressions of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and hepatocyte growth factor (HGF) were examined by immunohistochemical staining. Upon immunohistochemistry, overexpressions of VEGF, bFGF, and HGF in the ovarian stroma were observed in SSTs. In the normal ovary, the expressions were strong around the vessels and weak in the stroma of the ovary, while a Sertoli–Leydig cell tumour showed weak staining with VEGF, locally strong staining with bFGF and negative staining with HGF. This is the first report about angiogenic factors such as bFGF, and HGF in SST.Impact statementWhat is already known on this subject? Sclerosing stromal tumour (SST) is a rare benign tumour with high vascularity. The high vascularity of SSTs are reported to be related to the overexpression of vascular endothelial growth factor (VEGF). But there is no study on the expressions of other angiogenic factors in SST.What do the results this study add? In the immunohistochemical analysis, VEGF, bFGF and HGF were found to be widely stained in SSTs. This results suggest the possibility that the high vascularity seen on diagnostic imaging, and the many small vessels observed microscopically may be related to the expressions of VEGF, bFGF and HGF in SSTs.What are the implications of these findings for clinical practice and/or further research? Our results suggest the possibility that the combined expression pattern of VEGF, bFGF, and HGF may be used as marker of SSTs.

Exosomes serve as nanoparticles to suppress tumor growth and angiogenesis in gastric cancer by delivering hepatocyte growth factor siRNA.

Exosomes derived from cells have been found to mediate signal transduction between cells and to act as efficient carriers to deliver drugs and small RNA. Hepatocyte growth factor (HGF) is known to promote the growth of both cancer cells and vascular cells, and the HGF‐cMET pathway is a potential clinical target. Here, we characterized the inhibitory effect of HGF siRNA on tumor growth and angiogenesis in gastric cancer. In addition, we showed that HGF siRNA packed in exosomes can be transported into cancer cells, where it dramatically downregulates HGF expression. A cell co‐culture model was used to show that exosomes loaded with HGF siRNA suppress proliferation and migration of both cancer cells and vascular cells. Moreover, exosomes were able to transfer HGF siRNA in vivo, decreasing the growth rates of tumors and blood vessels. The results of our study demonstrate that exosomes have potential for use in targeted cancer therapy by delivering siRNA.

Expression of the hepatocyte growth factor and c-Met in colon cancer: correlation with clinicopathological features and overall survival.

The hepatocyte growth factor (HGF)/c-Met signaling system has been implicated in the development and progression of colon cancer, but the relationship between the expression of HGF or c-MET and clinicopathologic features remains controversial. In the study, we analyzed the expression of HGF and c-Met in colon cancer and assessed the influence of the expression of this growth factor and its receptor on clinical and histological parameters and patient survival. We investigated the mRNA expression of HGF and c-Met with real-time PCR in 90 unselected colon carcinomas and the corresponding normal mucosa. Furthermore, HGF and c-Met protein expression was investigated with immunohistochemistry in all the samples. The mRNA and protein expression levels of HGF and c-Met were significantly higher in colon cancer than in matched normal mucosa. The protein level in most of the cases investigated was correlated with the mRNA level. Overexpression of HGF and c-Met, at both protein and mRNA levels, was correlated with depth of invasion, lymph node metastases and overall AJCC stage. According to univariate analysis, the mean survival time was shorter in the HGF-positive and c-Met-positive groups. Multivariate Cox analysis showed that high M stage and the expression of c-Met independently had a negative impact on overall survival. The HGF/c-Met signaling pathway may be involved in the pathogenesis and progression of colon cancer. C-Met overexpression can be used as a useful parameter to evaluate the prognosis of colon cancer.

Effect of adipose tissue-derived mesenchymal stem cell transplantation in treatment of liver fibrosis and possible mechanism

Objective: To investigate the effect of adipose tissue-derived mesenchymal stem cell (ADSC) transplantation in the treatment of liver fibrosis rats and possible mechanism. Methods: Subcutaneous adipose tissue in the inguinal region of rats was collected to isolate ADSCs. The rats with liver fibrosis induced by intraperitoneally injected carbon tetrachloride were divided into cell transplantation group and phosphate buffer saline (PBS) injection group, and the rats which were fed normally were enrolled as negative control group. The rats in the cell transplantation group were given tail vein injection of ADSCs, and those in the PBS injection group were given injection of 0.5 ml PBS. At 7 days after transplantation, blood samples were collected from the inferior vena cava to evaluate liver function; liver tissue was collected to measure the protein expression of hepatocyte growth factor (HGF) and alpha-smooth muscle actin (α-SMA); Masson trichrome staining was used to evaluate intrahepatic collagen deposition. Hepatic stellate cells (HSCs) were collected from the rats with liver fibrosis, and indirect co-culture of HSCs and ADSCs was performed in vitro to analyze the influence of ADSCs on the proliferation and apoptosis of HSCs. The independent samples t-test was used for comparison between groups, and an analysis of variance was used for comparison of means between multiple samples. Results: ADSCs were found in liver tissue in the transplantation group, and compared with the PBS injection group, the transplantation group had significant alleviation in hepatocyte necrosis, vacuolization, and area of fibrosis and significant reductions in the serum levels of aminotransferases, while there was no significant difference in the level of albumin between the two groups. Compared with the PBS injection group, the transplantation group had significant upregulation in the protein expression of HGF and significant downregulation in the protein expression of α-SMA (both P < 0.05). In vitro co-culture for 72 hours showed that ADSCs inhibited the proliferation of HSCs, and there was a significant difference between the co-culture group and the control group with HSCs cultured alone. Caspase-3 immunostaining showed that after co-culture for 72 hours, there was a significant difference in the apoptosis rate of HSCs between the co-culture group and the control group with HSCs cultured alone (23.42% ± 3.02% vs 14.82% ± 3.93%). Conclusion: ADSC transplantation can upregulate the expression of HGF in the liver, promote the apoptosis of HSCs, and thus alleviate liver fibrosis.

Fibroblast-Specific β-Catenin Signaling Dictates the Outcome of AKI.

AKI is a devastating condition with high morbidity and mortality. The pathologic features of AKI are characterized by tubular injury, inflammation, and vascular impairment. Whether fibroblasts in the renal interstitium have a role in the pathogenesis of AKI is unknown. In this study, we investigated the role of fibroblast-specific β-catenin signaling in dictating the outcome of AKI, using conditional knockout mice in which β-catenin was specifically ablated in fibroblasts (Gli1-β-cat-/-). After ischemia-reperfusion injury (IRI), Gli1-β-cat-/- mice had lower serum creatinine levels and less morphologic injury than Gli1-β-cat+/+ littermate controls. Moreover, we detected fewer apoptotic cells, as well as decreased cytochrome C release; reduced expression of Bax, FasL, and p53; and increased phosphorylation of Akt, in the Gli1-β-cat-/- kidneys. Gli1-β-cat-/- kidneys also exhibited upregulated expression of proliferating cell nuclear antigen and Ki-67, which are markers of cell proliferation. Furthermore, Gli1-β-cat-/- kidneys displayed suppressed NF-κB signaling and cytokine expression and reduced infiltration of inflammatory cells. Notably, loss of β-catenin in fibroblasts induced renal expression of hepatocyte growth factor (HGF) and augmented the tyrosine phosphorylation of c-met receptor after IRI. In vitro, treatment with Wnt ligands or ectopic expression of active β-catenin inhibited HGF mRNA and protein expression and repressed HGF promoter activity. Collectively, these results suggest that fibroblast-specific β-catenin signaling can control tubular injury and repair in AKI by modulating HGF expression. Our studies uncover a previously unrecognized role for interstitial fibroblasts in the pathogenesis of AKI.

Effects of acupuncture at opposite acupoints on expression of hepatocyte growth factor in rats with skeletal muscle contusion

To observe the effects of acupuncture at opposite acupoints on expression of hepatocyte growth factor (HGF) in rats with skeletal muscle contusion, and to explore the mechanism of acupuncture at opposite acupoints on skeletal muscle contusion. Fifty-four Sprague Dawley (SD) rats were randomly divided into a blank group (6 rats), a model group (24 rats) and an opposing needling group (24 rats). The model group and opposing needling group were further divided into 1-day subgroup, 3-day subgroup, 5-day subgroup and 7-day subgroup, 6 rats in each one. No intervention was given in the blank group, while the model of skeletal muscle contusion was established in the model group and opposing needling group by self-made contusion device. 24 hours after contusion, electroacupuncture (EA) was applied at "Zusanli" (ST 36) and the corresponding points of ashi points at health side for 15 min, once a day. The subgroups of opposing needling group were treated for 1 day, 3 days, 5 days and 7 days, respectively. No treatment was given in the model group. Samples were collected in the subgroups 1 day, 3 days, 5 days and 7 days after treatment. The morphological change of injured gastrocnemius muscle was observed by using microscope after HE staining. The positive cell rate of proliferating cell nuclear antigen (PCNA) was detected by immunohistochemistry. The expression levels of HGF protein and PCNA protein were observed by Western blot. ① The results of HE staining showed that, 1 day after contusion, the inflammatory cells of gastrocnemius muscle in the opposing needling group were less than those in the model group; 3 days and 5 days after contusion, myoblasts and myotubes in the opposing needling group were more than those in the model group; 7 days after contusion, the neonatal muscle cells in the opposing needling group were more than those in the model group. ② The results of immunohistochemistry showed that, 1 day, 3 days and 5 days after contusion, the positive cell rate of PCNA in the opposing needling group was significantly higher than that in the model group (all P<0.001); 7 days after contusion, the positive cell rate of PCNA in the opposing needling group was significantly less than that in the model group (P<0.001). ③ The results of Western blot showed that, 1 day, 3 days and 5 days after contusion, the expression of HGF protein and PCNA protein in the opposing needling group was significantly higher than that in the model group (all P<0.05); 7 days after contusion, the expression of HGF protein and PCNA protein in the opposing needling group was significantly lower than that in the model group (all P<0.05). Acupuncture at opposite acupoints could regulate the expression of HGF and promote the activation, proliferation, migration and differentiation of muscle satellite cells in rats with skeletal muscle contusion, which could speed up the process of skeletal muscle injury repair.

Undifferentiated Adipose Tissue Stem Cell Transplantation Promotes Hepatic Regeneration, Ameliorates Histopathologic Damage of the Liver, and Upregulates the Expression of Liver Regeneration- and Liver-Specific Genes in a Rat Model of Partial Hepatectomy.

Objective Adipose tissue stem cells (ADSCs) present a promising therapeutic method to alleviate liver failure (LF). The purpose of this prospective study was to evaluate the efficacy of undifferentiated ADSC transplantation on liver regeneration and on the expression of liver regeneration- and liver-specific genes, following 60% partial hepatectomy (PHx). Methods Sixty female rats were subjected to PHx and were transplanted with 106 or 2 × 106 ADSCs, either into the portal vein (PV) or into the hepatic parenchyma. Animals of the control group were not transplanted and served as controls. Animals were sacrificed on the 4th, the 7th, or the 15th postoperative day (POD). Results The transplanted ADSCs were successfully engrafted into the liver parenchyma and ameliorated the histopathologic damage on the 7th and 15th POD. All transplanted animals demonstrated a significantly higher liver regeneration rate on the 4th and 7th POD, compared with the control group. The expression of hepatocyte growth factor, α-fetoprotein, tyrosine aminotransferase, hepatocyte nuclear factor 4a, and cytochrome P450 1A2 was significantly upregulated, compared with the control group. Conclusions Although undifferentiated, ADSC transplantation significantly enhanced the liver regeneration process. These findings may be proven clinically valuable, especially in cases of acute LF.

Transplantation of Telocytes Attenuates Unilateral Ureter Obstruction-Induced Renal Fibrosis in Rats

Background/Aims: Previous studies imply that telocytes may have a protective effect on fibrosis in various organs, including the liver, colon, and heart. The effect of telocytes on renal fibrosis remains unknown. Herein, this study was designed to investigate the effect of telocytes on renal fibrosis and the potential mechanisms involved. Methods: In a unilateral ureteral obstruction (UUO)-induced renal fibrosis model, telocytes were injected via the tail vein every other day for 10 days. The degree of renal damage and fibrosis was determined using histological assessment. The expression of collagen I, fibronectin, epithelial-mesenchymal transition markers, and Smad2/3 phosphorylation was examined by western blot analyses. Real-time PCR and enzyme-linked immunosorbent assay were performed in vivo to detect the levels of transforming growth factor (TGF)-β1 and various growth factors. Results: Telocytes attenuated renal fibrosis, as evidenced by reduced interstitial collagen accumulation, decreased expression of fibronectin and collagen I, upregulation of E-cadherin, and downregulation of α-smooth muscle actin. Furthermore, telocytes decreased serum TGF-β1 levels, suppressed Smad2/3 phosphorylation, and increased the expression of hepatocyte growth factor (HGF) in rat kidney tissue following UUO. Blockage of HGF counteracted the protective effect of telocytes on UUO-treated kidneys. Through the detection of HGF mRNA levels in vitro, we found that telocytes had no effect on HGF expression compared with renal fibroblasts. Conclusion: Telocytes attenuated UUO-induced renal fibrosis in rats, likely through enhancing the expression of HGF in an indirect manner.

Mesenchymal Stem Cells Attenuate the Renal Tubular Epithelial Cells Epithelial Mesenchymal Transition by Restoring the Reciprocal Balance between Hepatocyte Growth Factor and Transforming Growth Factor-β1

Objective: Mesenchymal stem cells (MSCs) have recently shown promise for the treatment of various types of chronic kidney disease models. However, the mechanism of this effect is still not well understood. Our study is aimed to investigate the effect of MSCs on transforming growth factor-β1 (TGF-β1)-induced EMT in renal tubular epithelial cells (HK-2 cells) and the underlying mechanism related to the reciprocal balance between hepatocyte growth factor (HGF) and TGF-β1.Methods: HK-2 cells were treated with TGF-β1-then co-cultured with MSCs. The induced EMT was assessed by cellular morphology and the expressions of alpha-smooth muscle actin (α-SMA) and EMT-related proteins. MTT assay and flow cytometry were employed to detect the effect of TGF-β1 and MSCs on HK-2 cell proliferation and apoptosis. SiRNA against hepatocyte growth factor (siHGF) was transfected to decrease the expression of HGF to identify the role of HGF in MSCs inhibiting HK-2 EMT.Results: TGF-β1 decreased HGF expression, induced EMT, suppressed proliferation and promoted apoptosis in HK-2 cells; but when co-cultured with MSCs all the outcomes were reversed. However, after treated with siHGF, all the benefits taken from MSCs vanished.Conclusion: TGF-β1 was a motivating factor of renal cell EMT and it suppressed the HGF expression. However, MSCs provided protection against EMT by increasing HGF level and decreasing TGF-β1 level. Our results also demonstrated HGF is one of the critical factor in MSCs anti-fibrosis.

Mesenchymal Stem Cells Modified with Heme Oxygenase-1 Have Enhanced Paracrine Function and Attenuate Lipopolysaccharide-Induced Inflammatory and Oxidative Damage in Pulmonary Microvascular Endothelial Cells

Background/Aims: Bone marrow-derived mesenchymal stem cell (BM-MSC) transplantation has therapeutic effects on endothelial damage during acute lung injury (ALI). Heme oxygenase-1 (HO-1) can restore homeostasis and implement cytoprotective defense functions in many pathologic states. Therefore, we explored whether transduction of HO-1 into BM-MSCs (MSCs-HO-1) would have an increased beneficial effect on lipopolysaccharide (LPS)-induced inflammatory and oxidative damage in human pulmonary microvascular endothelial cells (PVECs). Methods: MSCs were isolated from rat bone marrow and transfected with the HO-1 gene by a lentivirus vector. The phenotype and multilineage differentiation of MSCs were assessed. MSCs or MSCs-HO-1 were co-cultured with PVECs using a transwell system, and LPS was added to induce PVEC injury. The production of reactive oxygen species (ROS), and the activities of lipid peroxide (LPO), malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GPx) in PVECs were determined by flow cytometry and colorimetric assays, respectively. The levels of human PVEC-derived tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-6 in the supernatants of the co-culture system, and the activity of nuclear transcription factor-κB and NF-E2-related factor 2 (Nrf2) in PVECs were examined by enzyme-linked immunosorbent assay (ELISA). The mRNA expression of TNF-α, IL-1β and IL-6 in PVECs was detected by quantitative real-time polymerase chain reaction (qRT-PCR), HO-1 expression and enzymatic activity in PVECs and the influence of zinc protoporphyrin (ZnPP) or HO-1 small interfering RNA on the above inflammatory and oxidative stress markers were evaluated. In addition, the expression of rat MSC-derived hepatocyte growth factor (HGF) and IL-10 was determined by ELISA and qRT-PCR. Results: MSCs showed no significant changes in phenotype or multilineage differentiation after transduction. LPS strongly increased the production of inflammatory and oxidative stress indicators, as well as decreased the levels of antioxidant components and the activity of Nrf2 in PVECs. MSC co-cultivation ameliorated these detrimental effects in PVECs and MSCs-HO-1 further improved the damage to PVECs induced by LPS when compared with MSCs alone. The beneficial effects of MSCs-HO-1 were dependent on HO-1 overexpression and may be attributed to the enhanced paracrine production of HGF and IL-10. Conclusion: MSCs-HO-1 have an enhanced ability to improve LPS-induced inflammatory and oxidative damage in PVECs, and the mechanism may be partially associated with the enhanced paracrine function of the stem cells. These data encourage further testing of the beneficial effects of MSCs-HO-1 in ALI animal models.

MACC1 and HGF are associated with survival in patients with gastric cancer.

Metastasis-associsated in colon cancer 1 (MACC1), a newly identified oncogene, promotes tumor cell proliferation and invasion. In the present study, the expression of MACC1, hepatocyte growth factor (HGF) and its receptor, MET proto-oncogene (c-Met), was investigated in human gastric cancer tissues and adjacent normal tissues by immunohistochemistry. The association between the expression levels of the proteins and the clinicopathological parameters of the tumors were statistically analyzed. Furthermore, lentiviral particles expressing MACC1 were used to infect the hepatic satellite cell (HSC) line LX2. The expression of α-smooth muscle actin (SMA), HGF, matrix metallopeptidase (MMP)-2 and MMP-9 in human HSCs was examined by western blotting and reverse transcription-quantitative polymerase chain reaction. Transwell assays were used to measure the effect of MACC1-infected or non-infected HSCs on the migration and invasion abilities of MKN45 and MKN74 gastric carcinoma cells in vitro. The results demonstrated that positive protein expression of MACC1, HGF and c-Met was significantly higher in human gastric cancer tissues compared with adjacent normal tissues. Positive expression of MACC1 and c-Met in gastric cancer tissues had no correlation with the sex, age, tumor location and peritoneal metastasis of patients, but was significantly correlated with tumor size, depth of tumor invasion, lymph node metastasis, TNM stage, histological differentiation, and overall (5 years) and disease-free survival (5 years). Positive expression of each MACC1, HGF and c-Met protein was demonstrated to be positively correlated with each other in human gastric cancer tissues. Western blotting results confirmed that MACC1 protein was overexpressed in MACC1-overexpressing lentivirus-infected HSCs. Overexpression of MACC1 significantly increased HGF, MMP-2, MMP-9 and α-SMA expression levels in HSCs. Results from the Transwell assays indicated an increase in the number of MKN45 or MKN74 cells migrating towards MACC1-overexpressing HSCs, compared with control HSCs. These findings suggested that MACC1 may regulate the expression of HGF, MMP-2 and MMP-9 in HSCs, and may thus promote migration and invasion of gastric carcinoma cells. MACC1, HGF and c-Met might cooperatively participate in the malignant progression of gastric cancer. In conclusion, MACC1 might serve as a useful molecular target for the diagnosis, treatment and prognosis of gastric cancer.

Roles of Fibroblast Activation Protein and Hepatocyte Growth Factor Expressions in Angiogenesis and Metastasis of Gastric Cancer.

This study aims to explore the roles of fibroblast activation protein (FAP) and hepatocyte growth factor (HGF) expressions in the angiogenesis and metastasis of gastric cancer (GC). From May 2012 to December 2015, 110 GC patients who received surgical treatment in the First Hospital of Qinhuangdao were selected. The HGF and FAP expressions in 110 cases of GC, 130 cases of normal gastric mucosa and 115 cases of gastric ulcer were detected by streptavidin-perosidase (SP) method. Venous blood HGF level of GC patients was tested by enzyme-linked immunosorbent assay (ELISA). The micro-vessel number of the patients in the three groups were calculated and analyzed. In GC group, positive expression rates of FAP and HGF protein were 61.8% and 67.3% respectively, which were both higher than those in normal gastric mucosa and gastric ulcer groups. The micro-vessel numbers in patients of the normal gastric mucosa and gastric ulcer groups are far less than that in GC group. FAP, HGF and micro-vessel density (MVD) were significantly correlated with infiltration depth, tumor-node-metastasis (TNM) staging, lymph node metastasis (LNM) and distant metastasis. The results of ELISA showed that serum HGF level was related to tumor size, infiltration degree, TNM staging, LNM and distant metastasis. FAP and HGF expressions in GC were positively correlated with MVD, and the expressions of FAP and HGF in GC were in positive correlation. Our study provided evidence that high FAP and HGF expressions may be positively correlated with the angiogenesis and metastasis of GC.

MicroRNA-7-5p mediates the signaling of hepatocyte growth factor to suppress oncogenes in the MCF-10A mammary epithelial cell

MicroRNA-7 (miR-7) is a non-coding RNA of 23-nucleotides that has been shown to act as a tumor suppressor in various cancers including breast cancer. Although there have been copious studies on the action mechanisms of miR-7, little is known about how the miR is controlled in the mammary cell. In this study, we performed a genome-wide expression analysis in miR-7-transfected MCF-10A breast cell line to explore the upstream regulators of miR-7. Analysis of the dysregulated target gene pool predicted hepatocyte growth factor (HGF) as the most plausible upstream regulator of miR-7. MiR-7 was upregulated in MCF-10A cells by HGF, and subsequently downregulated upon treatment with siRNA against HGF. However, the expression of HGF did not significantly change through either an upregulation or downregulation of miR-7 expression, suggesting that HGF acts upstream of miR-7. In addition, the target genes of miR-7, such as EGFR, KLF4, FAK, PAK1 and SET8, which are all known oncogenes, were downregulated in HGF-treated MCF-10A; in contrast, knocking down HGF recovered their expression. These results indicate that miR-7 mediates the activity of HGF to suppress oncogenic proteins, which inhibits the development of normal cells, at least MCF-10A, into cancerous cells.

Carcinoma associated fibroblasts derived from oral squamous cell carcinoma promote lymphangiogenesis via c-Met/PI3K/AKT in vitro.

Carcinoma-associated fibroblasts (CAFs) are dominant components of the tumor microenvironment (TME) that promote the development, progression and metastasis of cancer. c-Met is a receptor of the hepatocyte growth factor (HGF), which is involved in lymphangiogenesis. Currently, the roles of CAFs during lymphangiogenesis are unknown. It has been hypothesized that CAFs contribute to lymphangiogenesis of oral squamous cell carcinoma (OSCC) via a HGF/c-Met complex. The expression of HGF in OSCC was determined using CAFs derived from OSCC tissue and it was demonstrated that HGF is overexpressed in OSCC-derived CAFs. It was also revealed that c-Met was highly expressed in human lymphatic endothelial cells (HLECs) when co-cultured with CAFs. Furthermore, it was demonstrated that recombinant human HGF significantly enhanced the proliferation, migration, invasion and tube formation of HLECs. By contrast, the inhibition of c-Met expression suppressed the aforementioned biological activities and also downregulated the expression of c-Met, phosphoinositide 3-kinase and phosphorylated protein kinase B. Taken together, these data demonstrate that c-Met is associated with the regulation of lymphangiogenesis. Thus, the results of the present study indicate that c-Met may be a promising novel therapeutic target to treat patients with OSCC.

Pre-culture in endothelial growth medium enhances the angiogenic properties of adipose-derived stem/stromal cells.

Considerable progress has been made on the development of adipose-derived stem/stromal cells (ASCs) as pro-angiogenic therapeutic tools. However, variable clinical results highlight the need for devising strategies to enhance their therapeutic efficacy. Since ASCs proliferate and stabilize newly formed vessels during the angiogenic phase of adipose tissue formation, we hypothesized that mimicking an angiogenic milieu during culture of ASCs would enhance their capacity to support endothelial cell survival and angiogenesis. To test this, we compared the effect of an endothelial growth medium (EGM-2) and conventional media (αMEM) on the progenitor and angiogenic properties of ASCs. ASCs cultured in EGM-2 (ASC-EGM) displayed the highest clonogenic efficiency, proliferative potential and multilineage potential. After co-culture under growth factor starvation, only ASC-EGM attenuated luciferase-expressing human umbilical vein endothelial cells (HUVECluc) apoptosis and supported the formation of endothelial cords in a dose-dependent manner. These effects were recapitulated by the conditioned medium of ASC-EGM, which displayed a 100-fold higher expression of hepatocyte growth factor in comparison with ASC-αMEM. Next, HUVECluc and ASCs were co-transplanted subcutaneously into immunodeficient mice, and the survival of HUVECluc was monitored by bioluminescent imaging. After 60days, the survival of HUVECluc transplanted alone was equivalent to that of HUVECluc co-transplanted with ASC-αMEM (15.0±0.7 vs. 13.0±0.5%). Strikingly, co-transplantation with ASC-EGM increased HUVECluc survival to 105.0±3.5%, and the resulting organoids displayed functional vasculature with the highest human-derived vascular area. These findings demonstrate that pre-conditioning of ASCs in endothelial growth medium augment their pro-angiogenic properties and could enhance their therapeutic efficacy against ischemic diseases.

Interleukin-1 receptor antagonist inhibits angiogenesis via blockage IL-1α/PI3K/NF-κβ pathway in human colon cancer cell.

PurposeThis article investigates the relationship between cancer cells and stromal cells in carcinoma cell living microenvironment and elucidates the mechanism that interleukin-1 receptor antagonist (IL-1RA) blocks metastatic potential in colon cancer.MethodsWestern blot and RT-PCR assay were used to determine the expression of hepatocyte growth factor (HGF) and IL-1α in colon carcinoma cells and stromal cells. Effect of IL-1RA and HGF on metastatic potential of colon cancer cells were examined by proliferation, invasion, and angiogenesis assays. The interactional role of IL-1RA and HGF were detected by ELISA assay, invasion, and angiogenesis assay making up a co-culture system consisting of stromal and colon cancer cells in cells living microenvironment.ResultsIL-1α was expressed in human umbilical vein endothelial cells (HUVECs) and HT-29 and WiDr (colon cancer cell lines with higher liver metastatic potential). HGF was expressed only in fibroblast. HGF secretion from fibroblasts was significantly inhibited by IL-1RA (P<0.01). Furthermore, IL-1RA could significantly inhibit migration, proliferation, and angiogenesis of HUVECs (P<0.01). In the double co-culture system, there is a high liver metastatic potential of colon cancer cell line (HT-29) because it can secrete autocrine IL-1α, significantly enhanced angiogenesis compared with low liver metastatic cell line (CaCo-2) (P<0.01), which does not secrete IL-1α. On the contrary, blockage of autocrine IL-1α by IL-1RA might significantly decrease metastatic potential of colon carcinoma cells through downregulation of IL-1α/PI3K/NF-κB pathway.ConclusionIL-1 receptor antagonist (IL-1RA) is an important inhibitor in metastatic process of colon carcinoma cell. Based on the above results, we suggest that IL-1RA may be a promising new therapeutic approach in inhibiting colon cancer with IL-1-producing patients.