Targeting HMGB1-TLR4 signaling by miR-216a-5p elevation alleviates the inflammatory behavioral hypersensitivity.

Neuroinflammation induced by microglial activation has a critical role in inflammatory pain. In this study, we detected the function of miR-216a-5p in the progression of inflammatory behavioral hypersensitivity. Here, decreases of miR-216a-5p and up-regulation of high-mobility group box1 (HMGB1) were observed in complete freund's adjuvant (CFA)-induced inflammatory pain model in mice and LSP-activated BV2 microglia. HMGB1 was identified as a target of miR-216a-5p by luciferase reporter system. Ectopic expression of miR-216a-5p suppressed microglial marker IBA-1 expression and subsequent pro-inflammatory cytokine releases (IL-1ß, IL-6 and TNF-α) from LPS-activated microglia. Additionally, LPS exposure enhanced the protein expression levels of HMGB1, TLR4 and p-p65 NF-kB in microglia, which were abrogated following miR-216a-5p overexpression. Intriguingly, transfection of HMGN1 cDNA into BV2 microglial cells reversed the inhibitory effects of miR-216a-5p elevation on microglial activation-triggered inflammatory response. Intrathecal delivery of LV-miR-216a-5-p ameliorated CFA-evoked mechanical and thermal hyperalgesia in mice. Concomitantly, overexpressing miR-216a-5p also restrained the inflammatory response and microglia activation in CFA-induced inflammatory mouse models, concomitant with the decreases in the expression of HMGB1, TLR4 and p-p65 NF-kB in spinal cord. Thus, these findings highlight that miR-216a-5p may alleviate inflammatory behavioral hypersensitivity by blocking microglia-mediated neuroinflammation via targeting the HMGB1-TLR4-NF-kB pathway, supporting miR-216a-5p as a potential therapeutic avenue for inflammatory pain.

Manifestations of liver injury in 333 hospitalized patients with coronavirus disease 2019 / 中华消化杂志

Objective:To investigate the manifestations of liver injury in hospitalized patients with coronavirus disease 2019 (COVID-19), to investigate the prognosis indicators of the disease, and to provide the reference for clinical diagnosis and treatment.Methods:From January 10 to February 14, 2020, at Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, the data of 333 hospitalized patients with COVID-19 were collected. The changes of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBil), direct bilirubin (DBil), indirect bilirubin (IBil) and albumin of the first liver function test after admission and the reexaminations of liver function test during hospitalization period in patients with liver injury were retrospectively analyzed. Student t test and Chi-square test were used for statistical analysis. Results:Liver injury occurred in 39.6% (132/333) of COVID-19 patients. There was no statistically significant difference in the rate of liver injury between patients in intensive care unit (ICU) and in general ward (45.6%, 26/57 vs. 38.4%, 106/276; χ2=1.026, P>0.05). 67.4% (89/132) of COVID-19 patients with liver injury presented with increased ALT or AST level on admission. During hospitalization, the level of ALT was higher than that of the first examination after admission ((60.28±50.44) U/L vs. (42.25±32.21) U/L), and the difference was statistically significant ( t=-3.230, P<0.05). The levels of ALT and AST of 71.2% (94/132) patients were both <80 U/L, which indicated that most of the patients showed mild liver injury. The patients with elevated level of TBil, DBil and IBil accounted for 3.9% (13/333), 5.4% (18/333) and 2.4% (8/333) of the COVID-19 patients, respectively. The albumin level of COVID-19 patients with liver injury during hospitalization was lower than that of the first examination after admission ((31.8±5.1) g/L vs. (33.7±5.4) g/L), and the difference was statistically significant ( t=2.712, P<0.05). The albumin levels at first examination on admission and reexamination during hospitalization of patients in ICU were both significantly lower than those of patients in general ward ((29.3±3.7) g/L vs. (34.8±5.1) g/L and (27.6±2.8) g/L vs. (32.9±5.1) g/L), and the differences were statistically significant ( t=4.928 and 4.783, both P<0.05). Conclusions:The incidence of liver injury in COVID-19 patients is high. A slight increase in aminotransferase levels is particularly common. Bilirubin abnormality is relatively rare and mild. The level of albumin may be one of the indicators for the severity and prognosis of COVID-19.

The glucose metabolic frame of conditions induce the overproduction of a co-culture pigment: induction mechanism of the pigment and development of a specific eutrophic-oligotrophic transition cultivation system.

Induction mechanism of a potential red pigment (RPc) was investigated in the present paper. A typical competition relationship exists between Penicillium sp. HSD07B and Candida tropicalis during co-culture, and C. tropicalis converts glucose into glycerol, organic acids and other substances, resulting in a stricter glucose limitation and the secretion of RPc. Moreover, a novel eutrophic-oligotrophic transition cultivation system (E-OTCS) was developed to produce red pigment during monoculture of Penicillium sp. HSD07B. However, the monoculture pigment (RPm) is different from RPc in components, and RP3 and RP4 only occur in RPm when glycerol is supplied. In addition, the additions of glycerol and organic acids to glucose exhaustion medium can significantly improve the pigment yield. These facts not only prove the feasibility of producing RPm using E-OTCS, but also reveal that, besides glucose exhaustion, the accumulation of metabolites of glucose including glycerol and organic acids is also an important factor influencing the production of RPc.

New candidate tumor-suppressor gene KLF6 and its splice variant KLF6 SV2 counterbalancing expression in primary hepatocarcinoma.

BACKGROUND/AIMS: This study aimed to detect the expression of newly discovered zinc finger transcriptional factor KLF6 and its splice variant KLF6 SV2 in primary hepatocarcinoma (PHC) tissues and hepatoma cell strains, and to evaluate their clinicopathologic relationship with PHC. METHODOLOGY: Wild-type KLF6 and KLF6 SV2 mRNA expression was determined by RTPCR in 27 cases of PHC tissues and cell strains of HepG2, SMMC7721 and LO2. Western blotting and immunohistochemical staining were adopted to detect KLF6 protein expression. Positive area ratio of wild-type KLF6 protein expression and its relationship with clinicopathological parameters of PHC was analyzed. RESULTS: Wild-type KLF6 expression in PHC tissues was lower than that in paracancerous tissues. In contrast, KLF6 SV2 mRNA expression was higher in PHC tissues and hepatoma cell strains (p<0.05). Positive area ratio of wild-type KLF6 protein expression was positively correlated with cellular differentiation degree of PHC (p<0.01), but negatively correlated not only with liver cirrhosis, tumor size and extrahepatic metastases (p<0.01), but also with portal vein thrombus and the number of lymph nodes with metastasis (p<0.05). CONCLUSIONS: Wild-type KLF6 deletion and inactivation was involved in the growth, cell differentiation and other physiological processes of PHC. The upregulation of KLF6 splice variant might counterbalance the wildtype KLF6 and contribute to the occurrence and development of PHC.

Expression of metadherin/AEG-1 gene is positively related to orientation chemotaxis and adhesion of human hepatocellular carcinoma cell lines of different metastatic potentials / 华中科技大学学报（医学）（英德文版）

Metastasis contributes to the poor prognosis of hepatocellular carcinoma (HCC). However, the mechanism through which a primary HCC cell develops into a metastatic phenotype is not well understood. The purpose of this study was to examine the correlation between metadherin (MTDH)/astrocyte elevated gene-1 (AEG-1) expression in HCC cell lines of different metastatic potentials and such metastatic phenotypes as orientation chemotaxis and adhesion. MTDH/AEG-1 expression was detected by RT-PCR and western blotting in HCC cell lines (HepG2, Huh7, Sk-HEP-1, MHCC-97H). Distribution of MTDH/AEG-1 was observed by immunofluorescence staining and confocal laser scanning microscopy. The abilities of orientation chemotaxis and adhesion and the index of interaction between HCC cell lines and microvascular endothelial cell lines (MVECs, including HUVECs and HPMECs) were measured by chemotaxis assay and adhesion assay, respectively. The results showed that MTDH/AEG-1 protein expression was significantly higher in high metastatic potential cancer cell lines (Sk-HEP-1, MHCC-97H) than in low metastatic potential cell lines (HepG2, Huh7) (P<0.05). The MTDH/AEG-1 protein was localized in the perinuclear region of HCC cells. Furthermore, the abilities of orientation chemotaxis and adhesion of HCC cells to HPMECs were increased as compared with those of HCC cells to HUVECs (P<0.05). The abilities of orientation chemotaxis and adhesion were much stronger in Sk-HEP-1 and MHCC-97H cells with MTDH/AEG-1 highly expressed than in HepG2 and Huh7 cells with MTDH/AEG-1 lowly expressed (P<0.05). These results suggested that the expression of MTDH/AEG-1 gene in HCC cell lines of different metastatic potentials was closely positively related to the abilities of orientation chemotaxis and adhesion of HCC cells. It was deduced that MTDH/AEG-1 might play a pivotal role in the lung-specific metastasis of HCC, which may be mediated through orientation chemotaxis and adhesion abilities of HCC cells. MTDH/AEG-1 may serve as a potential therapeutic target for HCC.

Expression of metadherin/AEG-1 gene is positively related to orientation chemotaxis and adhesion of human hepatocellular carcinoma cell lines of different metastatic potentials / 华中科技大学学报（医学）（英德文版）

Metastasis contributes to the poor prognosis of hepatocellular carcinoma (HCC). However, the mechanism through which a primary HCC cell develops into a metastatic phenotype is not well understood. The purpose of this study was to examine the correlation between metadherin (MTDH)/astrocyte elevated gene-1 (AEG-1) expression in HCC cell lines of different metastatic potentials and such metastatic phenotypes as orientation chemotaxis and adhesion. MTDH/AEG-1 expression was detected by RT-PCR and western blotting in HCC cell lines (HepG2, Huh7, Sk-HEP-1, MHCC-97H). Distribution of MTDH/AEG-1 was observed by immunofluorescence staining and confocal laser scanning microscopy. The abilities of orientation chemotaxis and adhesion and the index of interaction between HCC cell lines and microvascular endothelial cell lines (MVECs, including HUVECs and HPMECs) were measured by chemotaxis assay and adhesion assay, respectively. The results showed that MTDH/AEG-1 protein expression was significantly higher in high metastatic potential cancer cell lines (Sk-HEP-1, MHCC-97H) than in low metastatic potential cell lines (HepG2, Huh7) (P<0.05). The MTDH/AEG-1 protein was localized in the perinuclear region of HCC cells. Furthermore, the abilities of orientation chemotaxis and adhesion of HCC cells to HPMECs were increased as compared with those of HCC cells to HUVECs (P<0.05). The abilities of orientation chemotaxis and adhesion were much stronger in Sk-HEP-1 and MHCC-97H cells with MTDH/AEG-1 highly expressed than in HepG2 and Huh7 cells with MTDH/AEG-1 lowly expressed (P<0.05). These results suggested that the expression of MTDH/AEG-1 gene in HCC cell lines of different metastatic potentials was closely positively related to the abilities of orientation chemotaxis and adhesion of HCC cells. It was deduced that MTDH/AEG-1 might play a pivotal role in the lung-specific metastasis of HCC, which may be mediated through orientation chemotaxis and adhesion abilities of HCC cells. MTDH/AEG-1 may serve as a potential therapeutic target for HCC.