LincRNA ZNF529-AS1 inhibits hepatocellular carcinoma via FBXO31 and predicts the prognosis of hepatocellular carcinoma patients.

BACKGROUND: Invasion and metastasis of hepatocellular carcinoma (HCC) is still an important reason for poor prognosis. LincRNA ZNF529-AS1 is a recently identified tumour-associated molecule that is differentially expressed in a variety of tumours, but its role in HCC is still unclear. This study investigated the expression and function of ZNF529-AS1 in HCC and explored the prognostic significance of ZNF529-AS1 in HCC. METHODS: Based on HCC information in TCGA and other databases, the relationship between the expression of ZNF529-AS1 and clinicopathological characteristics of HCC was analysed by the Wilcoxon signed-rank test and logistic regression. The relationship between ZNF529-AS1 and HCC prognosis was evaluated by Kaplan‒Meier and Cox regression analyses. The cellular function and signalling pathways involved in ZNF529-AS1 were analysed by GO and KEGG enrichment analysis. The relationship between ZNF529-AS1 and immunological signatures in the HCC tumour microenvironment was analysed by the ssGSEA algorithm and CIBERSORT algorithm. HCC cell invasion and migration were investigated by the Transwell assay. Gene and protein expression were detected by PCR and western blot analysis, respectively. RESULTS: ZNF529-AS1 was differentially expressed in various types of tumours and was highly expressed in HCC. The expression of ZNF529-AS1 was closely correlated with the age, sex, T stage, M stage and pathological grade of HCC patients. Univariate and multivariate analyses showed that ZNF529-AS1 was significantly associated with poor prognosis of HCC patients and could be an independent prognostic indicator of HCC. Immunological analysis showed that the expression of ZNF529-AS1 was correlated with the abundance and immune function of various immune cells. Knockdown of ZNF529-AS1 in HCC cells inhibited cell invasion and migration and inhibited the expression of FBXO31. CONCLUSION: ZNF529-AS1 could be a new prognostic marker for HCC. FBXO31 may be the downstream target of ZNF529-AS1 in HCC.

KIF4A as a novel prognostic biomarker in cholangiocarcinoma.

ABSTRACT: Cholangiocarcinoma (CCA) is one of the most common malignant tumors. Although gene-targeted therapies have significantly improved the outcome of many cancers, the results are still not satisfactory for patients with CCA. Owing to the lack of an effective biomarker for guiding clinical treatment and monitoring prognosis in patients with CCA, the purpose of this study was to identify a new biomarker that could help predict the outcome of patients with CCA using bioinformatics tools.Gene expression data were collected from three publicly available datasets, comprising 263 patients with CCA and 22 healthy controls. Differentially expressed genes were obtained using the limma package (FDR < 0.05, |Log2FC|>1), and the respective protein-protein interaction revealed five relevant genes in the STRING dataset (TOP2A, BUB1, RRM2, TYMS, and KIF4A). The immunohistochemistry and PCR were used to analyze the difference in KIF4A expression in CCA.Kinesin Family Member 4A (KIF4A) was the only gene significantly associated with overall patient survival (P .035), with higher KIF4A expression being associated with poor survival rates. Moreover, KIF4A was significantly correlated with the infiltration of activated memory T cells (P = .0198) and activated mast cells (P = .008) in the tumor microenvironment. Increase in KIF4A expression affected the infiltration degree of the immune cells, which may be involved in the regulation of immune tolerance by CCA cells. The results indicated that the expression of KIF4A in CCA was higher than that in paracancerous tissues.Taken together, these findings suggest that KIF4A could be a potential new biomarker in CCA for predicting the response of patients to targeted immunotherapies.

Follistatin Like 5 (FSTL5) inhibits epithelial to mesenchymal transition in hepatocellular carcinoma.

BACKGROUND: Epithelial to mesenchymal transition (EMT) is a key process in determining distant metastasis and intra-hepatic dissemination of hepatocellular carcinoma (HCC). Follistatin (FST) family members are considered to be an attractive therapeutic targets and prognostic indicators in cancers. As a derivative of FST, Follistatin Like 5 (FSTL5) may play a similar role in HCC cells. This study aimed to investigate the expression and function of FSTL5 in HCC and its role in EMT. METHODS: FSTL5, E-cadherin and vimentin in HCC, and paracancerous tissues were detected by immunohistochemistry. Correlation of FSTL5 expression with overall survival was assessed. The proliferation and invasion of HCC cell lines SK-Hep1 and MHCC-LM3 were analyzed by cell counting kit-8 and Transwell assays. The expression of FSTL5, E-cadherin, and vimentin in HCC cells was examined by polymerase chain reaction and Western blot analysis. T-test was used to analyze the difference in proliferation and invasion ability between groups. The Spearman rank correlation test was used to detect the correlation between the expression of FSTL5 and E-cadherin or vimentin. RESULTS: The expression of FSTL5 in HCC was lower than that in paracancerous tissues (9.97% vs. 82.55%, χ = 340.15, P < 0.001). Patients with high FSTL5 expression had a better prognosis (χ = 8.22, P = 0.004) and smaller tumor diameter (χ = 45.52, P < 0.001), less lymph node metastasis (χ = 5.58, P = 0.02), earlier tumor node metastasis stage (χ = 11.29, P = 0.001), a reduced number of tumors (χ = 5.05, P = 0.02), lower alpha-fetoprotein value (χ = 24.36, P < 0.001), more probability of hepatitis carrying (χ = 40.9, P < 0.001), and better liver function grade (χ = 5.21, P = 0.02). Immunohistochemistry showed that FSTL5 expression in HCC tissues was positively correlated with E-cadherin expression (r = 0.38, P < 0.001) and negatively correlated with vimentin expression (r = -0.385, P < 0.001). Furthermore, over-expression of FSTL5 up-regulated the expression of E-cadherin and down-regulated the expression of vimentin in SK-Hep1 (negative control [NC] vs. FSTL5-interfering group [Lv-FSTL5]: E-cadherin [t = 45.03, P < 0.001], vimentin [t = 67, P < 0.001]) and MHCC-LM3 (NC vs. Lv-FSTL5: E-cadherin [t = 50, P < 0.001], vimentin [t = 72.75, P < 0.001]) cells at mRNA level. The same as protein level. In addition, the over-expression of FSTL5 inhibited the proliferation (NC vs. Lv-FSTL5: SK-Hep1, 3 d [t = 7.324, P = 0.018], 4 d [t = 6.23, P = 0.021], 5 d [t = 10.21, P = 0.003]; MHCC-LM3, 3 d [t = 4.32, P = 0.037], 4 d [t = 7.49, P = 0.012], 5 d [t = 9.3661, P = 0.009]) and invasion (NC vs. Lv-FSTL5: SK-Hep1, t = 21.57, P < 0.001; MHCC-LM3, t = 18.04, P < 0.001) of HCC cells. CONCLUSIONS: Down-regulation of FSTL5 may contribute to EMT of HCC, and FSTL5 is a potential target in the treatment of HCC.

Lentivirus-mediated overexpression of HSDL2 suppresses cell proliferation and induces apoptosis in cholangiocarcinoma.

BACKGROUND: Cholangiocarcinoma (CCA) is a malignant tumor of the bile duct epithelium, including intrahepatic, perihilar, and distal CCA based on anatomical location. Hydroxysteroid dehydrogenase-like 2 (HSDL2) belongs to the SDR subfamily of oxidoreductases, and it is involved in glioma oncogenesis, as it can promote cell proliferation and inhibit cell apoptosis. The purpose of this study was to explore the underlying molecular mechanisms of HSDL2 in the process of CCA. METHODS: HSDL2 expression levels were observed in CCA and adjacent (normal control) tissues by analyzing data from The Cancer Genome Atlas and Gene Expression Omnibus databases. A receiver operating characteristic curve analysis was carried out. In vitro, we overexpressed HSDL2 in RBE cells (a human CCA cell line) using a stable lentivirus-mediated transduction strategy. We then used quantitative real-time-PCR and Western blotting methods to detect the efficiency of HSDL2 overexpression. Cell proliferation was assessed using a Celigo Image Cytometer, MTT assays, and the expression of PCNA. Cell apoptosis was assessed by flow cytometry analysis, caspase3/7 activity, and the expression of the apoptotic markers BCL-2 and BAX. RESULTS: We observed a downregulation of HSDL2 in CCA tissues based on The Cancer Genome Atlas and Gene Expression Omnibus data analysis. The receiver operating characteristic curve analysis showed that HSDL2 could be an excellent efficacy biomarker for CCA. In vitro, HSDL2 overexpression largely suppressed the proliferation of RBE cells. In addition, apoptosis was induced by HSDL2 overexpression. CONCLUSION: The results of the data analysis indicated that, compared with adjacent tissues, HSDL2 was downregulated in CCA tissues, and overexpressing HSDL2 in CCA cells suppressed growth and proliferation, which involved activating apoptosis. This helps to understand the underlying HSDL2-related molecular mechanisms in the process of CCA.

Neuroendocrine tumor of the common bile duct: a case report and review of the literature.

We report a rare case of neuroendocrine tumor (NET) in the common bile duct (CBD). The patient is a 56-year-old female who presented to our department with symptoms of fever but without jaundice. A preoperative examination showed a tumor in the CBD. The tumor volume was almost 5.5 × 4.5 × 4 cm3, which is the biggest NET in the CBD reported on PubMed. The imaging results (computed tomography [CT] and magnetic resonance imaging [MRI]) were not consistent with CBD adenocarcinoma. The tumor appeared to oppress the growth of the CBD rather than originate in the bile duct wall; combined with the low blood bilirubin index and lack of jaundice symptoms, the preoperative diagnosis was not clear. We performed a radical resection of the cholangiocarcinoma. The patient recovered well before going home. The pathology was NET (Grade 2). The patient showed no recurrence to date, without intravenous chemotherapy (8 months).

Synchronous multiple primary gallbladder and gastric malignancies: Report of two cases and review of the literature.

Multiple primary malignancies (MPM) are rare. In particular, synchronous gallbladder and gastric malignancies are extremely rare, are associated with a concealed onset and atypical symptoms, and are highly likely to be overlooked or misdiagnosed. The clinical data of two patients with synchronous gallbladder and gastric malignancies are herein reported and integrated with the relevant literature to retrospectively analyze and summarize the pathogenesis and clinical characteristics of MPM. Case 1 was a male 46-year-old patient who underwent laparoscopic cholecystectomy, and succumbed to extensive tumor metastasis 2 months after the operation. Case 2 was an 80-year-old female patient who was treated with distal gastrectomy for gastric cancer, cholecystectomy, gastrojejunostomy and dissection of 5 suprapyloric, 6 subpyloric, 7 left gastric and 8 common hepatic artery lymph nodes, and succumbed to multiple organ failure induced by extensive tumor invasion within 1 week after the operation. Clinical physicians must pay closer attention to early symptoms of MPM in order to make an accurate diagnosis, perform timely radical surgical treatment and achieve favorable therapeutic outcomes, in terms of significantly increasing long-term patient survival rates.

Up-regulated FSTL5 inhibits invasion of hepatocellular carcinoma through the Wnt/ß-catenin/YAP pathway.

Patients with hepatocellular carcinoma (HCC) have a poor survival rate because of its high invasion ability. Therefore, it is necessary to elucidate the mechanisms of HCC migration and invasion. Our previous study showed that follistatin-like 5 (FSTL5), which was associated with the prognosis of HCC patients, acts as an inhibitor of HCC cell proliferation. It also promotes the transition of cell morphology from mesenchymal to epithelial, which is associated with the process of mesenchymal-to-epithelial transition. In this study, we used two HCC cell lines (SK-Hep1 and SMMC-7721) to explore the effect of FSTL5 on HCC invasion and migration. We found that up-regulated FSTL5 restrained HCC invasion and migration by transwell, wound healing, detachment, and attachment assays. Decreased expression of YAP was found upon over-expression of FSTL5, as well as inhibition of the Wnt/ß-catenin signaling pathway. YAP is a downstream gene of the Wnt/ß-catenin signaling pathway and plays an important role in HCC metastasis. Thus, we speculate that FSTL5 inhibits the invasion of HCC through the Wnt/ß-catenin/YAP pathway. In conclusion, FSTL5 exerts an inhibitory effect on HCC metastasis and proliferation through the Wnt/ß-catenin/YAP pathway and may be a target gene for anti-tumor therapy.

Multiple injections of human umbilical cord-derived mesenchymal stromal cells through the tail vein improve microcirculation and the microenvironment in a rat model of radiation myelopathy.

BACKGROUND: At present, no effective clinical treatment is available for the late effects of radiation myelopathy. The aim of the present study was to assess the therapeutic effects of human umbilical cord-derived mesenchymal stromal cells (UC-MSCs) in a rat model of radiation myelopathy. METHODS: An irradiated cervical spinal cord rat model was generated. UC-MSCs were injected through the tail vein at 90, 97, 104 and 111 days post-irradiation. Behavioral tests were performed using the forelimb paralysis scoring system, and histological damage was examined using Nissl staining. The microcirculation in the spinal cord was assessed using von Willebrand factor (vWF) immunohistochemical analysis and laser-Doppler flowmetry. The microenvironment in the spinal cord was determined by measuring the pro-inflammatory cytokines interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) in the serum and the anti-inflammatory cytokines brain-derived neurotrophic factor (BDNF) and glial cell-derived neurotrophic factor (GDNF) in the spinal cord. RESULTS: Multiple injections of UC-MSCs through the tail veil decreased the forelimb paralysis, decreased spinal cord histological damage, increased the number of neurons in the anterior horn of the spinal cord, increased the endothelial cell density and the microvessel density in the white matter and gray matter of the spinal cord, increased the relative magnitude of spinal cord blood flow, down-regulated pro-inflammatory cytokine expression in the serum, and increased anti-inflammatory cytokine expression in the spinal cord. CONCLUSION: Multiple injections of UC-MSCs via the tail vein in a rat model of radiation myelopathy significantly improved the microcirculation and microenvironment through therapeutic paracrine effects.

Radiation-induced endothelial cell loss and reduction of the relative magnitude of the blood flow in the rat spinal cord.

The main purpose of the present study was to examine the time-dependent alterations in the endothelial cell density that occur in the first 180 days after irradiation of the spinal cord and the functional role of these alterations in the spinal cord blood flow. An irradiated cervical spinal cord rat model (C2-T2 segment) was generated using a (60)Co irradiator to deliver 30 Gy. A significant loss of forelimb motor function was observed 180 days post-irradiation. The number of neurons in the anterior horn of the spinal cord began to decrease significantly 3 days post-irradiation compared with normal controls, reaching the lowest number at 90 days post-irradiation. A significant reduction in the endothelial cell density was observed from 14 days post-irradiation in the white matter and from 3 days post-irradiation in the gray matter. The lowest endothelial cell density was reached at 30 days post-irradiation in the white matter and at 60 days post-irradiation in the gray matter. A significant reduction in the microvessel density was observed from 3 days post-irradiation in both the white matter and the gray matter. The lowest microvessel density was reached at 90 days post-irradiation in both the white matter and the gray matter. A significant reduction in the relative magnitude of spinal cord blood flow was observed from 21 days post-irradiation. The lowest relative magnitude of spinal cord blood flow was reached at 90 days post-irradiation. We did not find any evidence of demyelination. The results revealed that a single 30-Gy irradiation dose resulted in impaired forelimb motor function, a decreased number of neurons, and reduced endothelial cell density, microvessel density and relative magnitude of spinal cord blood flow. However, a 30-Gy single-dose irradiation was not sufficient to induce demyelination in the rat spinal cord.

The green tea extract epigallocatechin-3-gallate inhibits irradiation-induced pulmonary fibrosis in adult rats.

The present study evaluated the effect of epigallocatechin-3-gallate (EGCG), the most abundant catechin in green tea, on irradiation-induced pulmonary fibrosis and elucidated its mechanism of action. A rat model of irradiation-induced pulmonary fibrosis was generated using a (60)Co irradiator and a dose of 22 Gy. Rats were intraperitoneally injected with EGCG (25 mg/kg) or dexamethasone (DEX; 5 mg/kg) daily for 30 days. Mortality rates and lung index values were calculated. The severity of fibrosis was evaluated by assaying the hydroxyproline (Hyp) contents of pulmonary and lung tissue sections post-irradiation. Alveolitis and fibrosis scores were obtained from semi-quantitative analyses of hematoxylin and eosin (H&E) and Masson's trichrome lung section staining, respectively. The serum levels of transforming growth factor ß1 (TGF-ß1), interleukin (IL)-6, IL-10, and tumor necrosis factor-α (TNF-α) were also measured. Surfactant protein-B (SPB) and α-SMA expression patterns were evaluated using immunohistochemistry, and the protein levels of nuclear transcription factor NF-E2-related factor 2 (Nrf-2) and its associated antioxidant enzymes heme oxygenase-1 enzyme (HO-1) and NAD(P)H: quinone oxidoreductase-1 (NQO-1) were examined via western blot analysis. Treatment with EGCG, but not DEX, reduced mortality rates and lung index scores, improved histological changes in the lung, reduced collagen depositions, reduced MDA content, enhanced SOD activity, inhibited (myo)fibroblast proliferation, protected alveolar epithelial type II (AE2) cells, and regulated serum levels of TGF-ß1, IL-6, IL-10, and TNF-α. Treatment with EGCG, but not DEX, activated Nrf-2 and its downstream antioxidant enzymes HO-1 and NQO-1. Taken together, these results showed that EGCG treatment significantly inhibits irradiation-induced pulmonary fibrosis. Furthermore, the results suggested promising clinical EGCG therapies to treat this disorder.