Identification of m6A modification patterns and RBM15 mediated macrophage phagocytosis in pancreatic cancer: An integrative analysis.

Pancreatic cancer (PC) responds weakly to conventional immunotherapy. RNA N6-methyladenosine (m6A) modification has an essential role in the immune response, while its potential role in PC tumor microenvironment (TME) immune cell infiltration remains unknown. In this study, we thoroughly assessed the m6A modification patterns of 472 PC samples using 19 m6A regulators, and we systematically correlated these modification patterns with TME immune cell infiltration characteristics. We also created the m6Ascore and evaluated the m6A modification patterns of individual tumors, identified three different m6A modification patterns, and explored the role of the important m6A "writer" RBM15 in the regulation of macrophage function in PC. Two independent PC cohorts confirmed that patients with higher m6Ascore showed significant survival benefit. We verified that knockdown of RBM15 has the ability to inhibit PC growth and to promote macrophage infiltration and enhance phagocytosis of PC cells by macrophages. In conclusion, m6A modifications play a non-negligible role in the formation of TME diversity and complexity in PC. We reveal that inhibition of RBM15 suppresses PC development and modulates macrophage phagocytosis, and provide a more effective immunotherapeutic strategy for PC.

Extended Ligation of the Hepatic Vein May Yield a Similar Effect to Liver Venous Deprivation in a Rat Model.

AIMS: To validate the hypothesis that hepatic vein ligation (HVL) alone may produce similar results to liver venous deprivation (LVD or HVL + portal vein ligation [PVL]). METHODS: Rats were assigned to five groups, namely, the control group; the R group in which the right median hepatic vein (RMHV) was ligated; the M group in which the middle median hepatic vein (MMHV) was ligated; the RM group in which both the RMHV and MMHV were ligated (R + MMHVL, extended ligation of the hepatic veins); and the LVD group in which both the right median portal vein and the RMHV were ligated. The liver hypertrophy effect and liver enzymes were determined. Methylene blue staining and retrograde pressurized perfusion assays were performed to investigate the hemodynamic changes. RESULTS: The RM and LVD groups exhibited similar significant hypertrophy in the future liver remnants when compared to that of the control group, and almost no additional hypertrophy effect was observed in the R and M groups. There was a remarkable elevation in serum transaminase levels in both groups. The methylene blue staining experiment indicated that pressure-dependent collaterals formed between the contiguous drainage areas, and the R + MMHVL procedure blocked the outflow of the right median lobe. CONCLUSION: Extended ligation of the hepatic vein (R + MMHVL) resulted in a similar hypertrophy effect and hepatic damage to those of LVD (HVL + PVL) treatment in a rat model, and intrahepatic venovenous collaterals play key roles.

Establishment of a Rat Model of Liver Venous Deprivation: Simultaneous Portal and Hepatic Vein Ligation.

Background and Aims: The aim was to establish a liver venous deprivation (LVD) model in rats, compare hepatic hypertrophy between LVD and associated liver partition and portal vein ligation for staged hepatectomy (ALPPS), and explore the underlying mechanisms. Methods: The LVD or extended-LVD (e-LVD) group received portal vein ligation (PVL) combined with hepatic vein ligation (HVL). The ALPPS or e-ALPPS group received PVL plus parenchyma ligation. Liver regeneration was assessed by measuring the liver weight and performing pathological analysis. Liver functions and the sphingosine kinase 1 (SPHK1)/sphingosine-1-phosphate (S1P)/sphingosine-1-phosphate receptor 1 (S1PR1) pathway were also investigated. Results: All future liver remnants (FLRs) in the ALPPS, e-ALPPS, LVD, and e-LVD groups exhibited significant hypertrophy compared with the control group. The LVD and e-LVD procedures induced similar liver hypertrophy than that in the corresponding ALPPS groups. Furthermore, the LVD and e-LVD methods led to obvious cytolysis in the venous-deprived lobes as well as a noticeable increase in serum transaminase levels, while no necrosis was observed in the ALPPS and e-ALPPS groups. SPHK1/S1P/S1PR1 pathway were distinctly activated after operation, especially in congestive/ischemic livers. Conclusions: We describe the first rat model of LVD and e-LVD with simultaneously associated HVL and PVL. Compared with the ALPPS technique, the LVD or e-LVD procedure had a comparable overall effect on the hypertrophy response and a stronger effect on liver function. The SPHK1/S1P/S1PR1 pathway was involved in the LVD- or ALPPS-induced liver remodeling.

Protein succinylation associated with the progress of hepatocellular carcinoma.

Although post-translational modification is critical to tumorigenesis, how succinylation modification of lysine sites influences hepatocellular carcinoma (HCC) remains obscure. 90 tumours and paired adjacent normal tissue of liver cancer were enrolled for succinylation staining. 423 HCC samples with 20 genes related to succinylation modification from TCGA were downloaded for model construction. Statistical methods were employed to analyse the data, including the Non-Negative Matrix Factorization (NMF) algorithm, t-Distributed Stochastic Neighbour Embedding (t-SNE) algorithm, and Cox regression analysis. The staining pan-succinyllysine antibody staining indicated that tumour tissues had a higher succinyllysine level than adjacent tissues (p < 0.001), which could be associated with a worse prognosis (p = 0.02). The survival was associated with pathological stage, tumour recurrence status and succinyllysine intensity in the univariate or multivariable cox survival analysis model. The risk model from 20 succinyllysine-related genes had the best prognosis prediction. The high expression of succinylation modification in HCC contributed to the worse patient survival prognosis. Model construction of 20 genes related to succinylation modification (MEAF6, OXCT1, SIRT2, CREBBP, KAT5, SIRT4, SIRT6, SIRT7, CPT1A, GLYATL1, SDHA, SDHB, SDHC, SDHD, SIRT1, SIRT3, SIRT5, SUCLA2, SUCLG1 and SUCLG2) could be reliable in predicting prognosis in HCC.

The Value of Early Positive Nucleic Acid Test and Negative Conversion Time of SARS-CoV-2 RNA in the Clinical Outcome of COVID-19 Patients.

Background: The outbreak of coronavirus disease (COVID-19) poses a great threat to global public health. At present, the number of newly confirmed COVID-19 cases and deaths is increasing worldwide. The strategy of comprehensive and scientific detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) through quantitative real-time polymerase chain reaction (qRT-PCR) for special populations and environments provides great support for the prevention and control of this pandemic in China. Our study focused on determining the factors associated with the length of time from symptom onset to the first positive nucleic acid test of throat swabs in COVID-19 patients, evaluating the effect of early positive nucleic acid detection on the disease severity and its significance in prognosis, and predicting the factors associated with the time from positive SARS-CoV-2 RNA test to negative conversion (negative conversion of SARS-CoV-2 virus) in COVID-19 patients. Methods: This study included 116 hospitalized patients with COVID-19 from January 30, 2020 to March 4, 2020 in Wuhan, China. Throat swab samples were collected for qRT-PCR testing of SARS-CoV-2 RNA, and all patients included in this study were positive for this test. Results: The multivariate Cox proportional hazards model showed that disease severity (HR = 0.572; 95% CI 0.348-0.942; p = 0.028) was a protective factor for the time from symptom onset to positive nucleic acid detection. Meanwhile, the time from symptom onset to positive nucleic acid detection (HR = 1.010; 95% CI 1.005-1.020; p = 0.0282) was an independent risk factor for the delay in negative conversion time of SARS-CoV-2 virus. However, the severity of the disease (HR=1.120; 95% CI 0.771-1.640; p = 0.544) had no correlation with the negative conversion time of SARS-CoV-2 virus. Conclusions: Patients with more severe disease had a shorter time from symptom onset to a positive nucleic acid test. Prolonged time from symptom onset to positive nucleic acid test was an independent risk factor for the delay in negative conversion time of SARS-CoV-2 virus, and the severity of the disease had no correlation with negative conversion time of SARS-CoV-2 virus.

Identification of Prognostic Genes in Hepatocellular Carcinoma.

Background: Previous studies have demonstrated the important role of tumor stem cells (TSCs) in the development of hepatocellular carcinoma (HCC); however, TSC-related genetic markers have not been investigated. Aim: The aim of the present study was to identify stem cell-related signature genes to predict the prognosis of HCC, using The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Methods: In total, 423 liver HCC tissue samples, including 373 tumor and 50 adjacent normal tissue samples from TCGA, and 115 primary tumor and 52 adjacent non-tumor tissue samples from the GEO GSE76427 database, were used in the present study. The non-negative matrix factorization (NMF) algorithm, t-distributed stochastic neighbor embedding (t-SNE) algorithm and Cox regression analysis were combined for model construction and validation. Results: Overall, six clusters were identified using the NMF and t-SNE algorithms with 470 stem cell-related genes. The results demonstrated that patients in cluster 5 had the worst prognosis. For multivariate Cox survival analysis, 15 genes with optimal lambda values were chosen and eight genes were incorporated into the final regression model using the optimal Akaike information criterion value. Validation of the risk model using the aforementioned eight signature genes demonstrated the models strong reliability and stable predictive performance. Conclusion: The results of the present study indicated that the eight-gene (Hes family BHLH transcription factor 5, KIT ligand, methyltransferase-like 3, proteasome 26S subunit non-ATPase 1, Ras-related protein Rab-10, treacle ribosome biogenesis factor 1, YTH N6-methyladenosine RNA binding protein 2 and Zinc Finger CCCH-Type Containing 13) signature constructed by the model may be reliable in predicting the prognosis of patients with HCC.

Coronavirus disease 2019 (COVID-19) in a Chinese renal transplant recipient: a case report.

Since December 2019, a novel coronavirus that caused viral pneumonia broke out and became global pandemic. Coronavirus disease 2019 (COVID-19) is caused by the SARS-CoV-2 virus. Reports on the clinical manifestations in solid organ transplant (SOT) recipients are rare. We report the clinical features and treatment of a Chinese renal transplant recipient with COVID-19. A 46-year-old Chinese woman, who had a renal transplant in 2006 due to chronic glomerulonephritis, was admitted to Renmin Hospital of Wuhan University for fever, cough, and expectoration for more than 10 days and diarrhea for 3 days. At admission, her body temperature was 38.2 °C and pulse oxygen saturation was 96% under oxygen inhalation. There were decreased breath sounds bilaterally. Laboratory data revealed normal leucocyte count, a normal percentage of neutrophils, a normal percentage of lymphocytes, decreased lymphocyte count, elevated procalcitonin and C-reactive protein (CRP), and increased levels of urea, creatinine, and estimated glomerular filtration rate. COVID-19 was confirmed by nasopharyngeal swab and sputum which were positive for SARS-CoV-2 by real-time reverse transcription PCR (RT-PCR). Chest CT revealed multiple patchy and flake ground-glass shadows in bilateral lung fields, and strip shadows in bilateral lower lobes. Treatment included antiviral (umifenovir, hydroxychloroquine), antibacterial (moxifloxacin), and other support therapies. Her symptoms, laboratory data, and chest CT showed trends of gradual improvement, while nasopharyngeal swabs were always positive for SARS-CoV-2. She was finally discharged from hospital on her 70th day of hospitalization when 2 consecutive nasopharyngeal swabs were negative for SARS-CoV-2. This is a rare report on COVID-19 in a renal transplant recipient, which can help enhance the understanding and treatment of COVID-19 in renal transplant recipients.

Clinical characteristics and peripheral immunocyte subsets alteration of 85 COVID-19 deaths.

OBJECTIVES: To retrospectively evaluate the clinical and immunological characteristics of patients who died of COVID-19 and to identify patients at high risk of death at an early stage and reduce their mortality. RESULTS: Total white blood cell count, neutrophil count and C-reactive protein were significantly higher in patients who died of COVID-19 than those who recovered from it (p < 0.05), but the total lymphocyte count, CD4 + T cells, CD8 + T cells, B cells and natural killer cells were significantly lower when compared in the same groups. Multiple logistic regression analysis showed that increased D-dimer, decreased CD4 + T cells and increased neutrophils were risk factors for mortality. Further multiple COX regression demonstrated that neutrophil ≥ 5.27 × 109/L increased the risk of death in COVID-19 patients after adjustment for age and gender. However, CD4 + T cells ≥ 260/µL appeared to reduce the risk of death. CONCLUSION: SARS-CoV-2 infection led to a significant decrease of lymphocytes, and decreased CD4 + T cell count was a risk factor for COVID-19 patients to develop severe disease and death. METHODS: This study included 190 hospitalized COVID-19 patients from January 30, 2020 to March 4, 2020 in Wuhan, China, of whom 85 died and 105 recovered. Two researchers independently collected the clinical and laboratory data from electronic medical records.

HSP4 triggers epithelial-mesenchymal transition and promotes motility capacities of hepatocellular carcinoma cells via activating AKT.

BACKGROUND AND AIMS: Heat shock factor (HSF4) plays a vital role in carcinogenesis and tumour progression. However, its clinical significance implications in hepatocellular carcinoma (HCC) remained elusive. METHODS: RT-PCR and western blot were used to detect the HSF4 expression levels in HCC cells and tissues. Immunohistochemistry staining was performed on a tissue microarray containing 104 HCC patients received radical resection. In vitro effects of HSF4 on proliferation, migration and invasion were determined by colony formation and transwell assays in HCCLM3, Huh7, MHCC97L and SMMC7721 cells. Epithelial-mesenchymal transition (EMT) was identified by RT-PCR, WB and immunofluorescence in HCCLM3 and MHCC97L cells. AKT pathway activation was detected by WB and dual luciferase report system in HCCLM3 and MHCC97L cells. RESULTS: HSF4 expression was higher in primary HCC tissues derived from recurrent patients, and positively correlated with invasiveness potentials of cell lines. Clinically, patients with high HSF4 expression had significant poorer prognosis. In vitro experiments showed HSF4 silencing inhibited HCC cell proliferation, migration and invasion, whereas HSF4 overexpression had inverse effects. Moreover, silence of HSF4 induced an epithelial-like phenotype, whereas the overexpression of HSF4 resulted in a mesenchymal-like phenotype in HCC by activating AKT pathway. Further experiments showed that HSF4 could activate AKT pathway in a hypoxia-inducible factor-1α (HIF-1α) dependent, but transforming growth factor-ß (TGF-ß) independent manner. CONCLUSIONS: HSF4 is upregulated in HCC, resulting in greater proliferation, migration and invasion capacities. Moreover, high HSF4 expression is a promising predictive indicator of poor outcome after radical resection. HSF4 may promote aggressive tumour behaviour by enhancing EMT through activating AKT pathway in a HIF1α-dependent manner.

Long noncoding RNA myocardial infarction-associated transcript regulated the pancreatic stellate cell activation to promote the fibrosis process of chronic pancreatitis.

BACKGROUND: Long noncoding RNAs (lncRNAs) play crucial roles in fibrosis process. In our previous RNA-seq study, we found that lncRNA myocardial infarction-associated transcript (MIAT) was differentially expressed in pancreatic tissues of chronic pancreatitis (CP) patients. However, the function of MIAT in CP remains unknown. This study was aimed to investigate the function and underlying mechanism of MIAT in pancreatic fibrosis. MATERIALS AND METHODS: The expression levels of MIAT, miR-216a-3p, cyclooxygenase 2 (COX-2), α-smooth muscle actin (α-SMA), and collagen I were estimated by Western blot analysis and qualitative reverse transcription polymerase chain reaction. The relationships between miR-216a-3p, MIAT, and COX-2 were confirmed by luciferase reporter assay. The proliferation of human pancreatic stellate cells (HPaSteCs) was detected by cell counting kit-8 assay. RESULTS: We found that MIAT, along with the levels of fibrosis-related proteins α-SMA and collagen I, as well as COX-2 were upregulated, while miR-216a-3p was downregulated in transforming growth factor (TGF)-ß1-stimulated HPaSteCs. Mechanistically, MIAT acted as a molecular sponge for miR-216a-3p. Furthermore, we identified COX-2 as a direct target of miR-126a-3p. Additionally, MIAT overturned the inhibitory effect of miR-216a-3p overexpression and COX-2 knockdown on the activation and proliferation of HPaSteCs. CONCLUSION: Our study provided mechanistic insights into a critical role for MIAT as a miRNA sponge in CP.

Exosomal miR-9-3p suppresses HBGF-5 expression and is a functional biomarker in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is the fifth most common cancer and the third leading cause of cancer-related death worldwide. Exosomes are secreted membrane vesicles that play important roles in various diseases by transporting proteins and RNAs, including microRNAs, between cells. However, the function of exosomal miRNA in HCC has not been fully investigated. METHODS: Exosomes were obtained from the sera by ultracentrifugation and were processed for transmission electron microscopy (TEM). Real time PCR were revealed changes of miRNA between patients and normal donors. Predicted targets of miRNA were described by bioinformatics analysis, luciferase reporter assay was used to confirmed whether miR-9-3p regulates target expression. And then miRNA were over-expressed in HCC cell line to study its function, western blotting were used to test expression of miRNA targets, Cell viability and proliferation were analyzed after over-expressed miR-9-3p using MTT and BrdU assay. RESULTS: Serum exosomes from patients with HCC contained significantly lower levels of the miR-9-3p than did serum exosomes from normal donors, suggesting a potential role for this microRNA in HCC. Bioinformatics analysis identified fibroblast growth factor 5 (HBGF-5), which plays an important role in cell proliferation, as a potential miR-9-3p target mRNA. Luciferase reporter assay confirming that miR-9-3p can directly regulate HBGF-5 expression. Consistent with this finding, overexpression of miR-9-3p in three HCC cell lines significantly downregulated HBGF-5 expression at both the mRNA and protein levels. Finally, overexpression of miR-9-3p reduced HCC cell viability and proliferation, and additionally reduced ERK1/2 expression, suggesting a potential mechanism by which miR-9-3p acts. CONCLUSIONS: These results provide new insight into the functions of miR-9-3p and HBGF-5 in HCC and identify miR-9-3p as a potential therapeutic target for HCC.

Hepatogenic differentiation from human adipose-derived stem cells and application for mouse acute liver injury.

Adipose-derived stem cells (ADSCs) derived from adipose tissue have the capacity to differentiate into endodermal, mesoderm, and ectodermal cell lineages in vitro, which are an ideal engraft in tissue-engineered repair. In this study, human ADSCs were isolated from subcutaneous fat. The markers of ADSCs, CD13, CD71, CD73, CD90, CD105, CD166, CYP3A4, and ALB were detected by immunofluorescence assays. Human ADSCs were cultured in a specific hepatogenesis differentiation medium containing HGF, bFGF, nicotinamide, ITS, and oncostatin M for hepatogenic differentiation. The hepatocyte markers were analyzed using immunofluorescence and real-time PCR after dramatic changes in morphology. Hepatocytes derived from ADSCs or ADSCs were transplanted into the mice of liver injury for observation cells colonization and therapy in liver tissue. The result demonstrated that human ADSCs were positive for the CD13, CD71, CD73, CD90, CD105, and CD166 but negative for hepatocyte markers, ALB and CYP3A4. After hepatogenic differentiation, the hepatocytes were positive for liver special markers, gene expression level showed a time-lapse increase with induction time. Human ADSCs or ADSCs-derived hepatocyte injected into the vein could improve liver function repair and functionally rescue the CCl4-treated mice with liver injury, but the ADSCs transplantation was better than ADSCs-derived hepatocyte transplantation. In conclusion, our research shows that a population of hepatocyte can be specifically generated from human ADSCs and that cells may allow for participation in tissue-repair.

Posterior mediastinal tuberculous lymphadenitis with dysphagia as the main symptom: a case report and literature review.

INTRODUCTION: Mediastinal tuberculous lymphadenitis (MTL) is mostly seen in primary tuberculosis in children, uncommon observed in adults. It usually presents the toxic symptoms of tuberculosis but rarely with symptoms characteristic of esophageal compression, such as dysphagia. Such patients can easily be misdiagnosed as esophageal neoplasm and get delayed or faulty treatment. CASE REPORT: A 32-year-old man presented with dull chest pain of one month and dysphagia of five days. CRP was elevated, and a skin test was strongly positive. At upper endoscopy, a protruding lesion covered by normal mucosa was seen at 26 cm from the upper incisor. Barium swallow showed visible external compressive stricture on the middle-lower esophagus with normal mucosal pattern. Chest computed tomography (CT) scan showed a subcarinal mass adjacent to the esophageal wall in posterior mediastinum. An endoscopic ultrasonography (EUS) revealed a hypoechoic lesion suspected of esophageal stromal tumor in the fourth layer. A tissue was obtained by ultrasound-guided fine-needle aspiration (EUS-FNA), but cytopathology, bacilliculture and PCR test had no special findings. The patient required experimental antitubercular treatment and the protruding lesion shrank gradually during therapy period. CONCLUSIONS: MTL could not be ignored in the differential diagnosis of posterior mediastinal mass with dysphagia. Analyzing and evaluating test results comprehensively is the key to make correct diagnosis and timely treatment. The experimental antituberculous treatment should be used if MTL is highly suspected.