GPR168 functions as a tumor suppressor in mouse melanoma by restraining Akt signaling pathway.

Malignant melanoma (MM) is a malignant tumor associated with high mortality rates and propensity for metastasis. Despite advancement in treatment, the incidence of MM continue to rise globally. GPR168, also known as MrgprF, is a MAS related GPR family member. The low expression of GPR168 has also been reported in many malignant tumors including MM. In the study, the statistical analysis from The Cancer Genome Atlas (TCGA) revealed a significant down regulation of GPR168 in melanoma compared to normal melanocytes, underscoring its importance in MM. The aim of the present study is to investigate the affect of GPR168 overexpression and elucidate its molecular mechanisms in MM cells. In addition, we used mouse melanoma B16-F10 cell line and xenograph tumor model to explore the function of GPR168 in melanoma. Our findings demonstrate that GPR168 overexpression could inhibit B16-F10 cell proliferation, migration, and xenografts tumor growth. Further, mechanistic studies revealed that GPR168 affected B16-F10 progress through Akt signal pathway with the decreased expression of p-Akt, p-GSK-3ß, ß-catenin, Myc, CyclinD1 and CDK4. In order to validate these findings, a rescue experiment was formulated employing GPR168 polyclonal antibody (Anti-GPR168 pAbs) to block GPR168 functionality. The addition of Anti-GPR168 pAbs into the culture medium restored both cell proliferation and migration. In conclusion, the overexpression of GPR168 in mouse melanoma B16-F10 cells suppressed proliferation and migration through the Akt signaling pathway. These findings collectively propose GPR168 as a promising novel tumor suppressor in MM, suggesting its potential as a therapeutic target in future interventions.

Qa-1b functions as an oncogenic factor in mouse melanoma cells.

BACKGROUND: Malignant melanoma is one kind of rare cancer in human and animals, which occurs not only in skin but also in the mucous membranes of the nose, mouth, anus, digestive tract, and in the uvea (choroid) of the eye. In order to develop therapies, a better understanding of the genetic landscape and signal pathways are needed. Numerous studies highlighted the importance of NKG2A-HLA-E in tumor immunotherapy, but the function and mechanism of HLA-E in tumor cells have seldom been studied alone. The statistical analyses from publicly available database Oncomine and The Cancer Genome Atlas (TCGA) showed that HLA-E is highly expressed in melanoma compared to non-transformed counterparts. In addition, melanoma patients with HLA-E high expression had a worse OS rate than the patients with low expression. These data indicate the importance of HLA-E in human melanoma. Qa-1b is the homolog of HLA-E in mouse. OBJECTIVE: To investigate the function and mechanism of Qa-1b in mouse melanoma. METHODS: Mouse melanoma cell line B16-F10 and allogenic melanoma model were used to investigate the function and mechanism of Qa-1b in melanoma. RESULTS: Qa-1b was highly expressed in B16-F10 compared with normal mouse epidermal cells. Qa-1b knockdown inhibited B16-F10 cell proliferation and migration in vitro and allogenic melanoma process in vivo. Furthermore, Qa-1b knockdown promoted cell cycle arrest at G0/G1 phase and cell apoptosis through Ras-Raf-MAPK signal pathway. CONCLUSION: Qa-1b functions as an oncogenic factor and it can be used as a new therapeutic target in melanoma.

Long-term glycemic variability and risk of stroke in patients with diabetes: a meta-analysis.

OBJECTIVES: Long-term glycemic variability has been related to increased risk of vascular complication in patients with diabetes. However, the association between parameters of long-term glycemic variability and risk of stroke remains not fully determined. We performed a meta-analysis to systematically evaluate the above association. METHODS: Medline, Embase, and Web of Science databases were searched for longitudinal follow-up studies comparing the incidence of stroke in diabetic patients with higher or lower long-term glycemic variability. A random-effect model incorporating the potential heterogeneity among the included studies were used to pool the results. RESULTS: Seven follow-up studies with 725,784 diabetic patients were included, and 98% of them were with type 2 diabetes mellitus (T2DM). The mean follow-up duration was 7.7 years. Pooled results showed that compared to those with lowest category of glycemic variability, diabetic patients with the highest patients had significantly increased risk of stroke, as evidenced by glycemic variability analyzed by fasting plasma glucose coefficient of variation (FPG-CV: risk ratio [RR] = 1.24, 95% confidence interval [CI] 1.11 to 1.39, P < 0.001; I2 = 53%), standard deviation of FPG (FPG-SD: RR = 1.16, 95% CI 1.02 to 1.31, P = 0.02; I2 = 74%), HbA1c coefficient of variation (HbA1c-CV: RR = 1.88, 95% CI 1.61 to 2.19 P < 0.001; I2 = 0%), and standard deviation of HbA1c (HbA1c-SD: RR = 1.73, 95% CI 1.49 to 2.00, P < 0.001; I2 = 0%). CONCLUSIONS: Long-term glycemic variability is associated with higher risk of stroke in T2DM patients.

Gender differences in the association between sleep duration and body mass index, percentage of body fat and visceral fat area among chinese adults: a cross-sectional study.

BACKGROUND: The relationship between sleep duration and anthropometric indices are still unclear. This study aimed to explore the association between sleep duration and body mass index (BMI), percentage of body fat (PBF) and visceral fat area (VFA) among Chinese adults, further to explore gender difference in it. METHODS: We analyzed part of the baseline data of a cohort study among adult attendees at two health-screening centers in China. Sleep duration was self-reported and categorized into short (< 7 h/day), optimal (7-9 h/day) and long sleep (≥ 9 h/day). BMI, PBF and VFA were assessed by bioelectric impedance analysis. Demographic characteristics, chronic diseases and medication history, physical activity, smoking and alcohol drinking behaviors were measured by an investigator-administrated questionnaire. RESULTS: A total of 9059 adult participants (63.08% were females) were included in the analysis. The participants aged from 19 to 91 years with the mean age of 45.0 ± 14.6 years. Short sleep was independently associated with elevated odds of general obesity (defined using BMI) and visceral obesity (defined using VFA) among the total study population, and gender differences were observed in these associations. Among women, short sleep was associated with 62% increased odds of general obesity (OR = 1.62, 95% CI: 1.24-2.12) and 22% increased odds of visceral obesity (OR = 1.22, 95% CI: 1.02-1.45). Among men, long sleep duration was associated with 21% decreased odds of visceral obesity (OR = 0.79, 95% CI: 0.64-0.99). No association was observed between sleep duration and PBF in both sexes. CONCLUSIONS: Sleep duration was associated with increased odds of general and visceral obesity, and this association differed between men and women. No association was observed between sleep duration and PBF among either males or females.

Gene Expression Characteristics of Liver Tissue Reveal the Underlying Pathogenesis of Hepatocellular Carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is high-mortality primary liver cancer and the most common malignant tumor in the world. This study is based on a hepatocellular carcinoma-related dysfunction module designed to explore the dysregulation of genes in liver cancer tissue. METHODS: By downloading the relevant data on the GEO database, we performed a differential analysis of healthy liver tissue and liver cancer tissues as well as healthy liver tissue and hepatocellular carcinoma tissue and then obtained two sets of differential genes and combined them. We performed a cointerpretation analysis of these differential genes and constructed related functional disorder modules. A hypergeometric test was performed to calculate the potential regulatory effects of multiple factors on the module, and a series of ncRNA and TF regulators were identified. We obtained a total of 4479 differentially expressed genes in hepatocellular carcinoma, and these genes were clustered into ten hepatocellular carcinoma-related functional interpretation disorder modules. RESULTS: Enrichment analysis revealed that these modular genes are mainly involved in signal transduction including cell cycle, TGF-beta signal transduction, and p53 signal transduction. Depending on the predictive analysis of multidimensional regulators, 323 ncRNAs and 52 TF-mediated hepatocellular carcinoma-related dysregulation modules were found to regulate disease progression. CONCLUSIONS: Based on a series of investigations, it was found that miR-30b-5p may participate in the peroxisome signal transduction by downregulating ABCD3-mediated module 1, thereby promoting the development and progression of hepatocellular carcinoma. Our research results not only provide a theoretical basis for biologists to study hepatocellular carcinoma further but also offer new methods and new ideas for the personalized care and treatment of hepatocellular carcinoma.

MiR-195-5p Ameliorates Cerebral Ischemia-Reperfusion Injury by Regulating the PTEN-AKT Signaling Pathway.

BACKGROUND: MiR-195-5p has been shown to play crucial roles in tumor inhibition, but its biological functions in cerebral ischemia-reperfusion (I/R) injury are unclear. METHODS: To mimic cerebral I/R injury, mice were induced by transient middle cerebral artery occlusion (MCAO). Human brain microvascular endothelial cells (HBMVECs) were treated with oxygen-glucose deprivation (OGD) to mimic I/R injury in vitro. The expression of miR-195-5p and PTEN was detected by qRT-PCR or Western blot. Cell viability was evaluated by CCK-8 assay. Cell apoptosis was detected by flow cytometer. Cell death was detected using specific lactate dehydrogenase (LDH) cytotoxicity kit. Infarct volume in mice brains was evaluated by TTC staining. Histopathological analysis was performed by HE staining and TUNEL staining. The interaction between miR-195-5p and PTEN was determined by TargetScan and luciferase reporter assay. RESULTS: MiR-195-5p was significantly downregulated and PTEN was upregulated during cerebral I/R injury both in vitro and in vivo. Overexpression of miR-195-5p efficiently enhanced cell viability, while reduced LDH release and apoptotic rate of OGD-treated HBMVECs in vitro. MiR-195-5p could negatively regulate the expression of PTEN by directly binding to its 3'-UTR. Overexpression of PTEN obviously attenuated the protective effect of miR-195-5p mimics on cell viability, LDH release and apoptosis in OGD-treated HBMVECs. Meanwhile, overexpression of miR-195-5p increased the expression levels of p-AKT in OGD-treated HBMVECs, while this effect was reversed by overexpression of PTEN. Moreover, overexpression of miR-195-5p efficiently ameliorated brain injury of mice after MCAO treatment in vivo. CONCLUSION: Overexpression of miR-195-5p ameliorated cerebral I/R injury by regulating the PTEN-AKT signaling pathway, providing a potential therapeutic target for cerebral I/R injury.

Prognostic Value of the Neo-Immunoscore in Renal Cell Carcinoma.

Objective: This study evaluated the prognostic value of the newly-built Immunoscore (neo-Immunoscore) in patients with renal cell carcinoma (RCC). Methods: Eighty-two patients with RCC were enrolled in this study. Their 3- and 5-year survival rates and overall survival (OS) were evaluated. The clinicopathologic data of the 82 patients were collected and analyzed. CD3, CD4, CD8, CD45RO, Foxp3, tumor necrosis factor receptor type II (TNFR2), programmed death ligand-1 (PD-L1), CD68, programmed death-1 (PD-1), cytokeratin (CK), and indoleamine 2,3-dioxygenase (IDO) were separated into two panels and stained using multiplex fluorescent immunohistochemistry methods. An immunologic prediction model of RCC patients, the neo-Immunoscore (neo-IS), was constructed using a Cox regression model. For the prognostic prediction of RCC, the neo-IS with the immunoscore (IS) proposed by the Society for Immunotherapy of Cancer (SITC) were compared by receiver operator characteristic (ROC) curve analysis. Survivals between the neo-ISlow and neo-IShigh groups were analyzed using the Kaplan-Meier method. Multivariate Cox regression survival analysis was applied to analyze independent indicators. Results: The Cox regression model allowed the establishment of a neo-IS based on three features: CD 3 CT + , CD4+Foxp3+CD45RO CT + , and CD8+PD- 1 IM + . Compared to that of the IS proposed by the SITC, the neo-IS obtained a better prediction. The 3- and 5-year survival rates in neo-IShigh RCC patients were significantly higher than those in neo-ISlow RCC patients (94.7 vs. 77.4%, P = 0.035 and 94.7 vs. 64.5%, P = 0.002, respectively). The OS in the neo-ISlow group was significantly shorter than that in the neo-IShigh group (73 vs. 97 months, P = 0.000). In comparisons of the neo-IS with clinical pathological factors, we found that the risk stratification and neo-IS were independent factors for the prognosis of patients with RCC. Moreover, the OS rate of neo-IShigh RCC patients with low- and intermediate- risk was higher than that of neo-ISlow patients. Conclusion: The newly-constructed IS model more precisely predicted the survival of patients with RCC and may supplement the prognostic value of risk stratification.

[Effect of Buyang Huanwut decoction on apoptosis of splenocytes in rats with sepsis].

OBJECTIVE: To observe the change in number of spleen T lymphocyte, B lymphocyte, apoptotic splenocytes and the expression of Bcl-2 and Bax protein in rats with sepsis; to study the effect of Buyang Huanwu decoction (BYHWD) on the immune function in rats with sepsis. METHODS: One hundred and eighty healthy male Wistar rats were randomly divided into four groups: sham operation group (n=30), model group (n=50), BYHWD treatment group (n=50) and BYHWD prevention group (n=50) . Sepsis model was reproduced by cecal ligation and puncture (CLP). The rats of BYHWD treatment group and BYHWD prevention group were given BYHWD (1 g/ml) 15 ml/kg at 30 minutes after surgery, and 3 days before surgery, once a day, respectively. The rats were sacrificed and their spleens were harvested at 6, 12, 24, 48, 96 hours after reproduction of the model. Morphological changes in spleen, the expression of T lymphocyte, B lymphocyte, apoptotic cells, Bcl-2 and Bax were determined in all the groups. RESULTS: By light microscopy, it was found that white pulp became atrophic and splenic nodules destroyed after CLP. The pathology was most obvious in model group, followed by BYHWD treatment group, and least obvious in prevention group. CD4(+) T lymphocyte, CD40 B lymphocyte and Bcl-2 protein expression in model group were obviously reduced compared with those of sham operation group, but the number of apoptotic cells and Bax protein expression were elevated, reaching their nadir or peak 24 hours after the surgery [the optical densities (A value): CD4(+) T lymphocyte: 18.28±4.57 vs. 98.60±18.18; CD40 B lymphocyte: 26.96±6.26 vs. 104.87±30.97; Bcl-2 protein expression: 20.23±11.75 vs. 149.67±5.24; apoptotic cells: 241.75±44.79 vs. 14.67±5.24; Bax protein expression: 128.75±44.79 vs. 5.34±4.26, all P<0.01], then they gradually increase or decrease. CD8(+) T lymphocyte count did not significantly changed . The results showed that BYHWD could markedly elevate the level of CD4(+) T lymphocyte and CD40 B lymphocyte, and lower the protein expression of apoptotic cells and Bax. There were no significant changes in the CD8(+) T lymphocyte and Bcl-2 protein expression. Furthermore, the results in the BYHWD prevention group were better than those in BYHWD treatment group (A value: CD4(+) T lymphocyte: 94.12±15.45 vs. 72.37±8.00; B lymphocyte: 90.46±13.34 vs. 55.66±4.23; apoptotic cells: 27.63±9.91 vs. 40.83±16.09; Bax protein expression: 11.63±5.91 vs. 30.83±16.09, P<0.05 or P<0.01). Ninety-six hours after CLP, the above indexes gradually approached to the level of the sham operation group. Correlation analysis showed that cell apoptosis and Bax were positively correlated (r=0.522, P=0.000), but cell apoptosis and Bcl-2 showed negative correlation (r=-0.659, P=0.000). CONCLUSION: BYHWD improves the immunological suppression in rats with sepsis by lowering apoptosis of CD4(+) T lymphocyte and B lymphocyte in spleen, and its underlying mechanism may be that BYHWD produce a decrease of apoptosis through Bax.

[Study on apoptosis of splenic lymphocytes in rats with sepsis].

OBJECTIVE: To observe the change in number of spleen T lymphocytes and B lymphocytes and apoptosis of splenocytes in rats with sepsis, and explore the mechanism of immune imbalance in sepsis. METHODS: Eighty healthy male Wistar rats were randomly divided into two groups: sham operation (n=30) and sepsis model (n=50). Sepsis model was reproduced by cecal ligation and puncture (CLP). The rats were sacrificed and their spleens were obtained at 6, 12, 24, 48, 96 hours after the model was reproduced. The pathological changes in spleen were observed by optical microscopy. Immunohistochemistry method was used to examine the positive expression of CD4(+)/CD8(+)T lymphocyte and B lymphocyte, Bax and Bcl-2. Apoptotic cells of spleen were detected by means of terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL). RESULTS: By optical microscopy, white pulps were atrophied and splenic nodules were destroyed after CLP. At 6, 12, 24, 48, 96 hours after CLP, the number of CD(+)T lymphocyte, B lymphocyte and Bcl-2 positive expressed cells in sepsis model group were obviously reduced as compared with sham operation group (all P<0.01). CD8(+) positive lymphocytes were not significant changed (all P>0.05). Apoptotic splenocytes and Bax positive cells were obviously increased compared with those of sham operation group (all P<0.01). Correlation analysis showed that Bcl-2 expression and cell apoptosis was negatively correlated (r=-0.659, P<0.01), but Bax positive cells and apoptosis showed positive correlation (r=0.522, P<0.01). CONCLUSION: Disturbance of immunological function exists in the rats with sepsis at 24 hours after CLP. The number of CD4(+) T lymphocyte, B lymphocyte was obviously reduced, apoptotic splenocytes were obviously increased. Bax and Bcl-2 play an important role in apoptosis of splenocytes in sepsis.