TRIM62 silencing represses the proliferation and invasion and increases the chemosensitivity of hepatocellular carcinoma cells by affecting the NF-κB pathway.

Tripartite interaction motif 62 (TRIM62), which is associated with the tumorigenic process, has been found to be aberrantly expressed in numerous cancers. However, the relationship between TRIM62 and hepatocellular carcinoma (HCC) has not been explored. In this work, the expression, function, and potential mechanism of TRIM62 in HCC were investigated. High TRIM62 levels were exhibited in HCC tissue, which contributed to a reduced overall survival. Loss-of-function experiments showed that TRIM62-silencing HCC cells proliferated more slowly, had reduced invasive ability and epithelial-mesenchymal transition, and were more sensitive to chemotherapeutic drug sorafenib. TRIM62 silencing resulted in the suppression of nuclear factor-kappa B (NF-κB), whereas the forced expression of TRIM62 enhanced NF-κB activation. The reduction of NF-κB could reverse TRIM62-overexpression-induced oncogenic effects in HCC cells. Importantly, TRIM62-silencing HCC cells had reduced tumorigenicity in nude mice. In summary, our data indicate that TRIM62 is highly expressed in HCC and acts as a potential tumor promoter. This work confirms that TRIM62 suppression displays remarkable tumor suppressive effects in HCC by affecting NF-κB activation.

Hereditary Spherocytosis With Liver Transplantation After Cirrhosis: A Case Report.

The clinical manifestations of hereditary spherocytosis are similar to those of various hemolytic anemias, which causes hereditary spherocytosis to be difficult to diagnose clinically. In this case, we obtained the peripheral blood of a patient and family members, and through a whole exome test of the 6,297 genetic phenotypes confirmed by OMIM, we found a heterozygous nonsense mutation (c.4117C>T, P.Q1373X) in the SPTB gene. Combined with the patient's clinical data, the diagnosis was hereditary spherocytosis. Compared with the public population sequence database, the mutation was found to be unique. Through protein structure prediction analysis and literature studies, we found that the mutation may cause SPTB mRNA instability, resulting in insufficient spectrin protein synthesis and affecting the integrity and flexibility of the red blood cell membrane skeleton. This case report found that SPTB gene mutations may cause liver dysfunction and cirrhosis in addition to hereditary spherocytosis, and this finding expands the phenotypic spectrum of SPTB. This study confirmed that NGS can be used to diagnose hereditary spherocytosis. Identifying mutated genes can not only accurately treat diseases, but also avoid potential genetic risks and improve prenatal and postnatal care.

Case Report: Use of Liposomal Amphotericin B in Low Doses in Patients With Visceral Leishmaniasis.

Background: No consensus has been reached regarding the optimal therapy for visceral leishmaniasis (VL), which affects ~12 million people worldwide. Case Presentation: This report described four cases of VL encountered in the First Affiliated Hospital of Xi'an Jiaotong University between October 2019 and December 2020. Of the four patients, one patient experienced relapse after antimonial treatment, and the remaining patients had primary VL (including one patient with impaired kidney function and one patient with hemophagocytic syndrome). All patients received a novel treatment protocol, namely the low-dose L-AmB therapy, which was characterized by a low initial dose, cautious dose escalation, and low-dose therapy as maintenance. All patients were cured without severe complications, and there was no further recurrence during follow-up. Conclusions: This case series demonstrated the safety and efficacy of the low-dose L-AmB therapy for VL patients, providing novel treatment protocol for the VL.

Metagenomic Next-Generation Sequencing in the Diagnosis of HHV-1 Reactivation in a Critically Ill COVID-19 Patient: A Case Report.

Background: Secondary infections pose tremendous challenges in Coronavirus disease 2019 (COVID-19) treatment and are associated with higher mortality rates. Clinicians face of the challenge of diagnosing viral infections because of low sensitivity of available laboratory tests. Case Presentation: A 66-year-old woman initially manifested fever and shortness of breath. She was diagnosed as critically ill with COVID-19 using quantitative reverse transcription PCR (RT-qPCR) and treated with antiviral therapy, ventilator and extracorporeal membrane oxygenation (ECMO). However, after the condition was relatively stabled for a few days, the patient deteriorated with fever, frequent cough, increased airway secretions, and increased exudative lesions in the lower right lung on chest X-rays, showing the possibility of a newly acquired infection, though sputum bacterial and fungal cultures and smears showed negative results. Using metagenomic next-generation sequencing (mNGS), we identified a reactivation of latent human herpes virus type 1 (HHV-1) in the respiratory tract, blood and gastrointestinal tract, resulting in a worsened clinical course in a critically ill COVID-19 patient on ECMO. Anti-HHV-1 therapy guided by these sequencing results effectively decreased HHV-1 levels, and improved the patient's clinical condition. After 49 days on ECMO and 67 days on the ventilator, the 66-year-old patient recovered and was discharged. Conclusions: This case report demonstrates the potential value of mNGS for evidence-based treatment, and suggests that potential reactivation of latent viruses should be considered in critically ill COVID-19 patients.

Quarantine at home may not enough!-from the epidemiological data in Shaanxi Province of China.

OBJECTIVES: A pneumonia associated with 2019 novel coronavirus (2019-nCoV, subsequently named SARS-CoV2) emerged worldwide since December, 2019. We aimed to describe the epidemiological characteristics of 2019 coronavirus disease (COVID-19) in Shaanxi province of China. RESULTS: 1. Among the 245 patients, 132 (53.9%) were males and 113 (46.1%) were females. The average age was 46.15 ± 16.43 years, ranging from 3 to 89 years. 2. For the clinical type, 1.63% (4/245) patients were mild type, 84.90% (208/245) were moderate type, 7.76% (19/245) were severe type, 5.31% (13/245) were critical type and only 0.41% (1/245) was asymptomatic. 3. Of the 245 patients, 116 (47.35%) were input case, 114 (46.53%) were non-input case, and 15 (6.12%) were unknown exposure. 4. 48.57% (119/245) cases were family cluster, involving 42 families. The most common pattern of COVID-19 family cluster was between husband and wife or between parents and children.

Phosphatidylinositol 4-phosphate adaptor protein 2 accelerates the proliferation and invasion of hepatocellular carcinoma cells by enhancing Wnt/ß-catenin signaling.

Phosphatidylinositol 4-phosphate adaptor protein 2 (FAPP2) has been recently identified as a tumor-associated regulator that is closely related to tumorigenesis. Yet, the precise role of FAPP2 in hepatocellular carcinoma (HCC) is still largely unknown. This study was designed to determine the function and molecular mechanisms of FAPP2 in HCC. Elevated expression of FAPP2 commonly occurred in the tumor tissue of HCC compared with normal controls. High expression of FAPP2 was also detected in HCC cell lines and its knockdown markedly decreased the proliferation, colony formation and invasion of HCC cells. Upregulation of FAPP2 by using a FAPP2 expression vector markedly promoted the proliferation, colony formation and invasion of HCC cells. FAPP2 was found to promote the activation of Wnt/ß-catenin signaling. Importantly, inhibition of Wnt/ß-catenin signaling abrogated the FAPP2 overexpression-conferred oncogenic effect in HCC cells. In addition, xenograft tumor experiments revealed that knockdown of FAPP2 significantly decreased the tumorigenicity of HCC cells in vivo. Taken together, the data of our study reported a tumor-promotion function of FAPP2 in HCC and demonstrate that knockdown of FAPP2 was capable of suppressing HCC cell proliferation and invasion through downregulation of Wnt/ß-catenin signaling. This study indicated that FAPP2 might be an attractive candidate anticancer target for HCC.

TRIM66 confers tumorigenicity of hepatocellular carcinoma cells by regulating GSK-3ß-dependent Wnt/ß-catenin signaling.

Tripartite motif 66 (TRIM66) protein, a member of the tripartite motif (TRIM) protein superfamily, has emerged as an oncogenic protein that is closely related to carcinogenesis in multiple cancers. However, whether TRIM66 plays a role in the progression of hepatocellular carcinoma (HCC) remains unknown. This study was aimed to investigate TRIM66 expression and its potential biological function in HCC cell lines. Here we showed that TRIM66 expression was significantly upregulated in HCC cell lines compared with normal control cells. Loss-of-function experiments by RNA interfering knockdown of TRIM66 showed that TRIM66 inhibition significantly reduced the proliferation, colony formation, and invasion of HCC cells, whereas gain-of-function by overexpression of TRIM66 exhibited the opposite effect. Further investigation showed that TRIM66 was involved in regulating glycogen synthase kinase-3ß (GSK-3ß) phosphorylation and ß-catenin expression. Knockdown of TRIM66 impeded the activation of Wnt signaling, while overexpression of TRIM66 promoted Wnt signaling activation. Moreover, inhibition of GSK-3ß by specific inhibitor partially reversed TRIM66 inhibition-mediated antitumor effect, while knockdown of ß-catenin blocked the oncogenic effect of TRIM66 overexpression in HCC cells. Additionally, in vivo experiments using a xenograft tumor model showed that TRIM66 knockdown blunted the tumorigenicity of HCC cells associated with downregulation of ß-catenin expression. Overall, our results showed that TRIM66 functioned as an oncogenic protein in HCC by promoting the activation of Wnt/ß-catenin signaling. Our study suggests that TRIM66 is a potential target for HCC treatment.

microRNA-520f inhibits hepatocellular carcinoma cell proliferation and invasion by targeting TM4SF1.

microRNAs (miRNAs) are reported to play crucial roles in tumorigenesis. Dysregulation of miR-520f has been implicated to be involved in several cancer progressions. However, the biological functions of miR520f in hepatocellular carcinoma (HCC) remain unclear. Thus, the molecular mechanism underlying miR-520f on HCC development was investigated in this study. Here, we found that miR-520f was remarkably down-regulated in human HCC samples and cell lines compared to paired normal tissues and cell lines as detected by qRT-PCR. Furthermore, the deregulated miR-520f was strongly associated with larger tumor size, advanced TNM stage, and metastasis in HCC patients. Functional investigations revealed that overexpression of miR-520f significantly suppressed cell proliferation, invasion and migration, caused cell cycle arrested at G0/G1 phase, and promoted cell apoptosis in HCC cells according to MTT, colony formation, transwell, and flow cytometry assays, respectively, whereas, downregulation of miR-520f exhibited inverse effects. Transmembrane-4 L-Six family member-1 (TM4SF1) was identified as a direct target of miR-520f, and an inverse relationship was found between miR-520f and TM4SF1 mRNA levels in HCC specimens. Rescue experiments suggested that restoration of TM4SF1 partially abolished miR-520f-meidated cell proliferation and invasion inhibition in HCC cells through regulating P13K/AKT and p38 MAPK signaling pathways. In conclusion, these data indicated that miR-520f acted as tumor suppressor in HCC proliferation and invasion by targeting TM4SF1, which might provide potential therapeutic evidence for HCC patients.

Pterostilbene alleviates polymicrobial sepsis-induced liver injury: Possible role of SIRT1 signaling.

Liver injury occurs frequently during sepsis. Pterostilbene (Pte), a natural dimethylated analog of resveratrol from blueberries, exerts anti-inflammatory and anti-apoptotic effects in various diseases. However, the role of Pte in sepsis-induced liver injury and its underlying mechanisms remain unknown. The current study aimed to evaluate the protective effects of Pte on sepsis-induced liver injury and its potential mechanisms. Sepsis was induced using cecal ligation and puncture (CLP) in C57BL/6 mice. Mice were administered Pte (5, 10, 15mg/kg, i.p.) at 0.5h, 2h, and 8h after CLP induction. The pathological changes of the liver were evaluated using hematoxylin and eosin (H&E) staining. The serum levels of alanine transaminase (ALT) and aspartate aminotransferase (AST) were measured. The levels of tumor necrosis factor-alpha (TNF-α), interleukin (IL-6), myeloperoxidase (MPO), p38 mitogen-activated protein kinase (p38MAPK), Bax, and B-cell lymphoma 2 (Bcl-2) were also evaluated. Pte treatment attenuated the CLP-induced liver injury, as evidenced by the attenuated histopathologic injuries and the decreased serum aminotransferase levels. Pte reduced the serum inflammatory cytokine (TNF-α and IL-6) levels and hepatic mRNA levels of TNF-α and IL-6. Pte also reduced MPO activity and p38MAPK activation in the liver. Additionally, Pte significantly inhibited Bax expression and increased Bcl-2 expression. Moreover, Pte increased the expression of sirtuin-1 (SIRT1) and reduced the expression of acetylated forkhead box O1 (Ac-FoxO1), acetylated Ac-p53, and acetylated nuclear factor-kappa beta (Ac-NF-κB). However, SIRT1 small interfering RNA (siRNA) abolished Pte's effects on the expression levels of those protein. Notably, Pte improved the survival rate in septic mice. In conclusion, Pte alleviates sepsis-induced liver injury by reducing inflammatory response and inhibiting hepatic apoptosis, and the potential mechanism is associated with SIRT1 signaling activation.

Knockdown of FRAT1 inhibits hypoxia-induced epithelial-to-mesenchymal transition via suppression of the Wnt/ß-catenin pathway in hepatocellular carcinoma cells.

Hypoxia-induced epithelial-to-mesenchymal transition (EMT) in hepatocellular carcinoma (HCC) was investigated. Frequently rearranged in advanced T-cell lymphomas-1 (FRAT1) is a positive regulator of the Wnt/ß-catenin signaling pathway and is overexpressed in many human tumors. However, the expression and role of FRAT1 in HCC has not been elucidated. In this study, we investigated the effect of FRAT1 on EMT process in HCC cells induced by hypoxia. Our results showed that FRAT1 is highly expressed in HCC tissues and cell lines. Hypoxia significantly induced FRAT1 expression in HCC cells. FRAT1 knockdown inhibited hypoxia-induced cell migration/invasion, downregulation of epithelial markers and upregulation of mesenchymal markers. Moreover, FRAT1 knockdown suppressed the expression levels of ß-catenin, cyclin D1 and c-myc in HCC cells under the same hypoxic condition. Our results revealed that FRAT1 is a hypoxia factor that is critical for the induction of EMT in HCC cells. These data suggest a potential role for targeting FRAT1 in the prevention of hypoxia-induced HCC cancer progression and metastasis mediated by EMT.

Effects of hepatitis C virus gene NS2 on the expressions of Bcl-2 and Bax in HepG2 cells.

OBJECTIVE: To study the effects of hepatitis C virus gene nonstructural protein 2 on the expressions of Bcl-2 and Bax in liver hepatocellular cells. METHOD: The study was conducted at the Department of Infectious Diseases, the First Affiliated Hospital of Medical School, Xi'an Jiaotong University, Xi'an, China, from March 2012 to April 2013. Negative controls pEGFP-C3-NS2, pEGFP-C3-C and pEGFP-C3 were transiently transfected into liver hepatocellular cells and expressions of Bcl-2 and Bax were detected by Western blot 24 h post-transfection. SPSS 13 was used for statistical analysis. RESULTS: After transfected with NS2 gene, expression of Bcl-2 in liver hepatocellular cells was slightly higher than the non-transfected cells, and the expression of Bax was significantly higher than the non-transfected cells. CONCLUSION: Hepatitis C virus non-structural protein 2 gene plays a role in adjusting the proto-oncogene Bcl-2 and tumour suppressor gene Bax.

Cloning and functions of the HBxAg-binding protein XBP1.

In the present study the hepatitis B virus X antigen binding protein 1 (XBP1) was cloned by inducing its expression, and its subcellular localization and function were examined. Total RNA was extracted from HepG2 cells and XBP1 was amplified using reverse transcription polymerase chain reaction (RT-PCR), followed by restriction enzyme digestion of the pGBKT7 yeast plasmid and identification by enzyme digestion. The plasmid was transformed into AH109 yeast via the lithium acetate method and protein extracts were prepared. XBP1 protein expression in the eukaryotic cells was determined using polyacrylamide gel electrophoresis and western blot analysis. The gene encoding the XBP1-binding protein was screened in liver cells using yeast two-hybrid technology. We transfected a human hepatocellular carcinoma cell line and observed the intracellular localization of the gene expression protein using a fluorescence microscope, followed by prokaryotic expression and XBP1 gene identification. A 921-bp XBP1 gene fragment was obtained via RT-PCR amplification and 20 proteins with known functions that interact with XBP1 were screened, including metallothionein, smooth muscle cell-related protein, asialoglycoprotein receptor, pyruvate dehydrogenase kinase 1 and a sequence with unknown functions. A green fluorescent protein expression plasmid pEGFP-C1-XBP1 of XBP1 was constructed successfully and its expression protein was localized in the cytoplasm. A 56-kDa recombinant protein was successfully obtained via prokaryotic expression and was demonstrated to have good specificity using western blot analysis. The XBP1 gene, which expresses the XBP1 protein, is located in the cytoplasm and plays a role in the intracellular structure, cell growth, intracellular metabolism and signal transduction pathway, as well as DNA duplication, transcription, recombination and repair.

Determination of urine tumor necrosis factor, IL-6, IL-8, and serum IL-6 in patients with hemorrhagic fever with renal syndrome

OBJECTIVE: The aim of this study was to explore the role of cytokines in the pathogenesis of hemorrhagic fever with renal syndrome (HFRS). METHODS: Double-antibody sandwich ELISA was used to determine serum interleukin (IL)-6, urine tumor necrosis factor (TNF), IL-6, and IL-8 levels in 56 patients with HFRS. RESULTS: Serum IL-6, urine TNF, IL-6, and IL-8 concentrations in HFRS patients were significantly higher than those in the control group (p < 0.001). the concentrations increased at fever stage, then continued to increase during the hypotension stage and peaked at the oliguria stage. the concentrations of serum IL-6, urine TNF, IL-6, and IL-8 increased according to the severity of the disease, and differed greatly among different types of the disease. serum IL-6 had remarkable relationships with serum specific antibodies. it was positively related to serum 12-microglobulin (β-mg), blood ureanitrogen (bun), and creatinine (Cr). significant positive relationships were also found both between urine IL-6 and TNF, and between IL-6 and IL-8 (r = 0.5768, p < 0.05; r = 0.3760, p < 0.01). CONCLUSION: TNF, IL-6, and IL-8 were activated during the course of the disease. IL-6 was associated with the immunopathological lesions caused by the hyperfunction of the humoral immune response. IL-6, IL-8 and TNF were involved in renal immune impairment. determining them might, to a certain extent, be useful in predicting the prognosis and outcome of patients with hfrs.

Determination of urine tumor necrosis factor, IL-6, IL-8, and serum IL-6 in patients with hemorrhagic fever with renal syndrome.

OBJECTIVE: The aim of this study was to explore the role of cytokines in the pathogenesis of hemorrhagic fever with renal syndrome (HFRS). METHODS: Double-antibody sandwich ELISA was used to determine serum interleukin (IL)-6, urine tumor necrosis factor (TNF), IL-6, and IL-8 levels in 56 patients with HFRS. RESULTS: Serum IL-6, urine TNF, IL-6, and IL-8 concentrations in HFRS patients were significantly higher than those in the control group (p<0.001). The concentrations increased at fever stage, then continued to increase during the hypotension stage and peaked at the oliguria stage. The concentrations of serum IL-6, urine TNF, IL-6, and IL-8 increased according to the severity of the disease, and differed greatly among different types of the disease. Serum IL-6 had remarkable relationships with serum specific antibodies. It was positively related to serum ß2-microglobulin (ß2-MG), blood urea nitrogen (BUN), and creatinine (Cr). Significant positive relationships were also found both between urine IL-6 and TNF, and between IL-6 and IL-8 (r=0.5768, p<0.05; r=0.3760, p<0.01). CONCLUSION: TNF, IL-6, and IL-8 were activated during the course of the disease. IL-6 was associated with the immunopathological lesions caused by the hyperfunction of the humoral immune response. IL-6, IL-8 and TNF were involved in renal immune impairment. Determining them might, to a certain extent, be useful in predicting the prognosis and outcome of patients with HFRS.

NS5ATP9 gene regulated by NF-kappaB signal pathway.

NS5ATP9 was previously identified as p15 PAF [proliferating cell nuclear antigen (PCNA)-associated factor] to bind with PCNA. We earlier identified the promoter region of NS5ATP9 and found NS5ATP9 is a NS5A up-regulation gene. However little is known about how it is regulated. To investigate the gene regulation of NS5ATP9, we screened NS5ATP9 promoter binding proteins using phage display and verified by electrophoretic mobility shift assay (EMSA). We found that the nuclear protein rhNF-kappaB (p50) could bind to the NS5ATP9 promoter and the binding region contained within a 156 bp (nucleotides -5 to -161bp) immediately upstream of the transcription initiation site. Our results suggest that NF-kappaB could participate in the regulation of NS5ATP9 gene expression in carcinogenesis.

NS5ATP9, a gene up-regulated by HCV NS5A protein.

Non-structural protein 5A (NS5A) appears to interact with a variety of cellular proteins and play an important role in mediating cell growth, cellular signaling pathways and pathogenesis of hepatitis C virus (HCV). NS5ATP9 was identified as a NS5A trans-activated protein in suppression subtractive hybridization (SSH), and the regulation was confirmed by luciferase reporter assay and quantitative real time PCR (qRT-PCR). A minimal promoter region contained within the 211bp (nucleotides -161 to +50bp) immediately upstream of the transcription initiation site. NS5ATP9 is a NS5A up-regulation gene which may play a role in the pathogenesis of HCV-associated hepatocellular carcinoma.