Supplementary Material for: Resveratrol Inhibits Proliferation and Induces Apoptosis through the Hedgehog Signaling Pathway in Pancreatic Cancer Cell

<p><i>Purpose:</i> To investigate the effect and possible mechanisms of resveratrol on pancreatic cancer cells in vitro. <i>Methods:</i> After being treated with resveratrol, cell viability, cell cycle phase distribution and apoptosis rate of pancreatic cancer cells were measured by CCK-8 assay and flow cytometer, respectively. The effects of resveratrol on the Hedgehog pathway were studied by real-time RT-PCR and Western blotting. By interfering Gli1 expression in PANC-1 cells and overexpressing Gli1 in BxPC-3 cells, we detected the expressions of Gli1-targeted genes, such as Ptc1, CCND1 and BCL-2, compared with resveratrol experimental group. We further used the luciferase reporter assay to explore the correlation between resveratrol and Gli1. <i>Results:</i> Resveratrol inhibited the growth of pancreatic cancer cells in a dose- and time-dependent manner. Compared with control group, the cells in the G0/G1 phase and the apoptosis rate were significantly increased. Low concentration of resveratrol decreased the expression of the Hedgehog pathway members including Gli1, Ptc1 and Smo. The expression of downstream target genes of the Hedgehog pathway such as Gli1, Ptc1, CCND1 and BCL-2 were significantly decreased after 12.5 µ<i>M</i> resveratrol treatment, which demonstrated a similar change of gene expression when Gli1 was knocked down by the RNAi technique in PANC-1 cells. Resveratrol also downregulated the expression of Gli1, Ptc1, CCND1 and BCL-2 in Gli1-overexpressed BxPC-3 cells. Results of the luciferase assay showed that resveratrol did not act on the Gli1 promoter directly. <i>Conclusion:</i> Resveratrol can inhibit pancreatic cancer cell survival and its mechanisms might be partly via the Hedgehog signaling pathway.</p

Supplementary Material for: Comprehensive analysis of the relationship between cuproptosis-related gene GCSH and prognosis, tumor microenvironment infiltration, and therapy response in endometrial cancer

Background: Cuproptosis, a novel form of cell death regulated by protein lipoylation and implicated in mitochondrial metabolism. However, the impact of the cuproptosis-related gene γ-glutamylcysteine synthetase (GCSH) on endometrial cancer (EC) prognosis, tumor immune microenvironment, and therapeutic response remains to be further researched. Methods: The differential expression of GCSH between endometrial cancer and normal tissues was analyzed using multiple public databases. Additionally, cancer and adjacent tissues were prospectively collected from 17 EC patients, and immunohistochemical analysis was performed to further investigate GCSH expression differences. The relationship between GCSH and the prognosis and clinicopathological characteristics of EC patients was evaluated, and a nomogram was constructed to predict patient survival based on GCSH expression. Then, Gene set variation analysis (GSVA) was utilized to explore the potential biological functions of GCSH in EC. The impact of GCSH on the tumor microenvironment (TME) was estimated. Finally, the effect of GCSH on the response to immunotherapy and chemotherapeutic drugs in EC was investigated. Results: The expression of GCSH was significantly upregulated in EC. High GCSH expression was associated with poor prognosis in EC patients. Enrichment analysis showed that high GCSH was associated with immune suppression. Furthermore, high GCSH was found to be associated with a non-inflamed TME, leading to decreased infiltration levels of immune cells. Finally, it was observed that patients with high GCSH were insensitive to both immunotherapy and chemotherapeutic drugs. Conclusion: This study revealed the role of GCSH in TME, response to therapy, and clinical prognosis in EC, which provided novel insights for the therapeutic application in EC

Supplementary Material for: Comprehensive analysis of the relationship between cuproptosis-related gene GCSH and prognosis, tumor microenvironment infiltration, and therapy response in endometrial cancer

Background: Cuproptosis, a novel form of cell death regulated by protein lipoylation and implicated in mitochondrial metabolism. However, the impact of the cuproptosis-related gene γ-glutamylcysteine synthetase (GCSH) on endometrial cancer (EC) prognosis, tumor immune microenvironment, and therapeutic response remains to be further researched. Methods: The differential expression of GCSH between endometrial cancer and normal tissues was analyzed using multiple public databases. Additionally, cancer and adjacent tissues were prospectively collected from 17 EC patients, and immunohistochemical analysis was performed to further investigate GCSH expression differences. The relationship between GCSH and the prognosis and clinicopathological characteristics of EC patients was evaluated, and a nomogram was constructed to predict patient survival based on GCSH expression. Then, Gene set variation analysis (GSVA) was utilized to explore the potential biological functions of GCSH in EC. The impact of GCSH on the tumor microenvironment (TME) was estimated. Finally, the effect of GCSH on the response to immunotherapy and chemotherapeutic drugs in EC was investigated. Results: The expression of GCSH was significantly upregulated in EC. High GCSH expression was associated with poor prognosis in EC patients. Enrichment analysis showed that high GCSH was associated with immune suppression. Furthermore, high GCSH was found to be associated with a non-inflamed TME, leading to decreased infiltration levels of immune cells. Finally, it was observed that patients with high GCSH were insensitive to both immunotherapy and chemotherapeutic drugs. Conclusion: This study revealed the role of GCSH in TME, response to therapy, and clinical prognosis in EC, which provided novel insights for the therapeutic application in EC

Supplementary Material for: Comprehensive analysis of the relationship between cuproptosis-related gene GCSH and prognosis, tumor microenvironment infiltration, and therapy response in endometrial cancer

Background: Cuproptosis, a novel form of cell death regulated by protein lipoylation and implicated in mitochondrial metabolism. However, the impact of the cuproptosis-related gene γ-glutamylcysteine synthetase (GCSH) on endometrial cancer (EC) prognosis, tumor immune microenvironment, and therapeutic response remains to be further researched. Methods: The differential expression of GCSH between endometrial cancer and normal tissues was analyzed using multiple public databases. Additionally, cancer and adjacent tissues were prospectively collected from 17 EC patients, and immunohistochemical analysis was performed to further investigate GCSH expression differences. The relationship between GCSH and the prognosis and clinicopathological characteristics of EC patients was evaluated, and a nomogram was constructed to predict patient survival based on GCSH expression. Then, Gene set variation analysis (GSVA) was utilized to explore the potential biological functions of GCSH in EC. The impact of GCSH on the tumor microenvironment (TME) was estimated. Finally, the effect of GCSH on the response to immunotherapy and chemotherapeutic drugs in EC was investigated. Results: The expression of GCSH was significantly upregulated in EC. High GCSH expression was associated with poor prognosis in EC patients. Enrichment analysis showed that high GCSH was associated with immune suppression. Furthermore, high GCSH was found to be associated with a non-inflamed TME, leading to decreased infiltration levels of immune cells. Finally, it was observed that patients with high GCSH were insensitive to both immunotherapy and chemotherapeutic drugs. Conclusion: This study revealed the role of GCSH in TME, response to therapy, and clinical prognosis in EC, which provided novel insights for the therapeutic application in EC

Supplementary Material for: Comprehensive analysis of the relationship between cuproptosis-related gene GCSH and prognosis, tumor microenvironment infiltration, and therapy response in endometrial cancer

Background: Cuproptosis, a novel form of cell death regulated by protein lipoylation and implicated in mitochondrial metabolism. However, the impact of the cuproptosis-related gene γ-glutamylcysteine synthetase (GCSH) on endometrial cancer (EC) prognosis, tumor immune microenvironment, and therapeutic response remains to be further researched. Methods: The differential expression of GCSH between endometrial cancer and normal tissues was analyzed using multiple public databases. Additionally, cancer and adjacent tissues were prospectively collected from 17 EC patients, and immunohistochemical analysis was performed to further investigate GCSH expression differences. The relationship between GCSH and the prognosis and clinicopathological characteristics of EC patients was evaluated, and a nomogram was constructed to predict patient survival based on GCSH expression. Then, Gene set variation analysis (GSVA) was utilized to explore the potential biological functions of GCSH in EC. The impact of GCSH on the tumor microenvironment (TME) was estimated. Finally, the effect of GCSH on the response to immunotherapy and chemotherapeutic drugs in EC was investigated. Results: The expression of GCSH was significantly upregulated in EC. High GCSH expression was associated with poor prognosis in EC patients. Enrichment analysis showed that high GCSH was associated with immune suppression. Furthermore, high GCSH was found to be associated with a non-inflamed TME, leading to decreased infiltration levels of immune cells. Finally, it was observed that patients with high GCSH were insensitive to both immunotherapy and chemotherapeutic drugs. Conclusion: This study revealed the role of GCSH in TME, response to therapy, and clinical prognosis in EC, which provided novel insights for the therapeutic application in EC

Supplementary Material for: Comprehensive analysis of the relationship between cuproptosis-related gene GCSH and prognosis, tumor microenvironment infiltration, and therapy response in endometrial cancer

Background: Cuproptosis, a novel form of cell death regulated by protein lipoylation and implicated in mitochondrial metabolism. However, the impact of the cuproptosis-related gene γ-glutamylcysteine synthetase (GCSH) on endometrial cancer (EC) prognosis, tumor immune microenvironment, and therapeutic response remains to be further researched. Methods: The differential expression of GCSH between endometrial cancer and normal tissues was analyzed using multiple public databases. Additionally, cancer and adjacent tissues were prospectively collected from 17 EC patients, and immunohistochemical analysis was performed to further investigate GCSH expression differences. The relationship between GCSH and the prognosis and clinicopathological characteristics of EC patients was evaluated, and a nomogram was constructed to predict patient survival based on GCSH expression. Then, Gene set variation analysis (GSVA) was utilized to explore the potential biological functions of GCSH in EC. The impact of GCSH on the tumor microenvironment (TME) was estimated. Finally, the effect of GCSH on the response to immunotherapy and chemotherapeutic drugs in EC was investigated. Results: The expression of GCSH was significantly upregulated in EC. High GCSH expression was associated with poor prognosis in EC patients. Enrichment analysis showed that high GCSH was associated with immune suppression. Furthermore, high GCSH was found to be associated with a non-inflamed TME, leading to decreased infiltration levels of immune cells. Finally, it was observed that patients with high GCSH were insensitive to both immunotherapy and chemotherapeutic drugs. Conclusion: This study revealed the role of GCSH in TME, response to therapy, and clinical prognosis in EC, which provided novel insights for the therapeutic application in EC

Supplementary Material for: ING4 Inhibits Proliferation and Induces Apoptosis in Human Melanoma A375 Cells via the Fas/Caspase-8 Apoptosis Pathway

<p><b><i>Background:</i></b> Inhibitor of growth 4 (ING4) plays a role in regulating the cell cycle, apoptosis, cell invasion and migration, but the mechanisms involved remain to be elucidated. <b><i>Objective:</i></b> To explore how ING4 affects human malignant melanoma A375 cells. <b><i>Methods:</i></b> Recombinant lentiviral vectors (A375/pLenO-GTP-ING4) were constructed and transfected into A375 cells (experimental group). The impact of ING4 on the proliferation and apoptosis of A375 cells was investigated in in vitro and in vivo experiments in mice using the MTT assay and flow cytometry. <b><i>Results:</i></b> In the experimental group, optical density was lower and apoptotic cells were more frequent from days 2-5 (p = 0.000 and p < 0.01); there were smaller xenografts and more apoptotic cells in mice (all p < 0.05); moreover, increased levels of Fas, cleaved caspase-8 and caspase-3, and decreased levels of FasL and procyclic acidic repetitive protein were observed in vitro and in vivo. <b><i>Conclusion:</i></b> ING4 might suppress proliferation and enhance apoptosis in human malignant melanoma cells by activating Fas-induced apoptosis in a caspase-8-dependent pathway.</p

Supplementary Material for: Combination of Mean Platelet Volume/Platelet Count Ratio and the APACHE II Score Better Predicts the Short-Term Outcome in Patients with Acute Kidney Injury Receiving Continuous Renal Replacement Therapy

<b><i>Background/Aims:</i></b> Both the Acute physiology and Chronic Health Evaluation (APACHE II) score and mean platelet volume/platelet count Ratio (MPR) can independently predict adverse outcomes in critically ill patients. This study was aimed to investigate whether the combination of them could have a better performance in predicting prognosis of patients with acute kidney injury (AKI) who received continuous renal replacement therapy (CRRT). <b><i>Methods:</i></b> Two hundred twenty-three patients with AKI who underwent CRRT between January 2009 and December 2014 in a Chinese university hospital were enrolled. They were divided into survivals group and non-survivals group based on the situation at discharge. Receiver Operating Characteristic (ROC) curve was used for MPR and APACHE II score, and to determine the optimal cut-off value of MPR for in-hospital mortality. Factors associated with mortality were identified by univariate and multivariate logistic regression analysis. <b><i>Results:</i></b> The mean age of the patients was 61.4 years, and the overall in-hospital mortality was 48.4%. Acute cardiorenal syndrome (ACRS) was the most common cause of AKI. The optimal cut-off value of MPR for mortality was 0.099 with an area under the ROC curve (AUC) of 0.636. The AUC increased to 0.851 with the addition of the APACHE II score. The mortality of patients with of MPR > 0.099 was 56.4%, which was significantly higher than that of the control group with of ≤ 0.099 (39.6%, P= 0.012). Logistic regression analysis showed that average number of organ failure (OR = 2.372), APACHE II score (OR = 1.187), age (OR = 1.028) and vasopressors administration (OR = 38.130) were significantly associated with poor prognosis. <b><i>Conclusion:</i></b> Severity of illness was significantly associated with prognosis of patients with AKI. The combination of MPR and APACHE II score may be helpful in predicting the short-term outcome of AKI

Supplementary Material for: Corticosteroid for IgA Nephropathy: Are They Really Therapeutic?

<b><i>Background:</i></b> IgA nephropathy (IgAN) is a common chronic glomerular disease that, in most patients, slowly progresses to end-stage kidney disease. The therapy with corticosteroid in IgAN is still a worldwide problem that is confusing the clinicians. <b><i>Methods:</i></b> MEDLINE, EMBASE, the Cochrane Library, and article reference lists were searched for randomized controlled trials (RCTs) that compared corticosteroids with placebo and any other non-immunosuppressive agents in treating IgAN. Twelve RCTs involving 1,057 patients were included. <b><i>Results:</i></b> Overall, we found that steroids had statistically significant effects in preventing the decline in renal function (relative risk 0.42, 95% CI 0.25–0.71, <i>p</i> < 0.001) and reducing proteinuria (SMD: –0.58 g/day, 95% CI –0.80 to –0.36 g/day) in patients with IgAN. The association between glucocorticoid and risk of kidney outcome was not modified by steroids’ type (prednisone or methylprednisone), dose (≤30 or > 30 mg/day), duration (≤8 or > 8 months), or serum creatinine (< 1.10 or ≥1.10 mg/dL). But steroids increased the risk of side effects such as gastrointestinal and endocrinium symptoms. <b><i>Conclusion:</i></b> This study provides the clear beneficial effects of the steroids therapy on the kidney function and proteinuria, although it should be used with caution

Erratum: Development and Validation of a New Dry Weight Estimation Method Using Single Frequency Bioimpedance in Hemodialysis Patients

<i>Background:</i> We proposed a new method to estimate dry weight (DW) using single frequency bioimpedance. <i>Methods:</i> We hypothesized that the change in whole body resistance at 50 kHz (R₅₀) was proportional to the ultrafiltration volume (UFV) during a hemodialysis (HD) session. When the targeted resistance estimated in healthy subjects was reached, the patient achieved his/her DW. UFV and R₅₀ were monitored in 40 HD patients. Another 43 HD patients were stratified into 2 groups to validate this method. <i>Results:</i> The change in whole body resistance was proportional to UFV in each of the 40 HD patients. In the DW_decrease group, pre-dialysis systolic blood pressure (n = 29, 154.5 ± 22.8 vs. 146.9 ± 22.3, p < 0.05) and antihypertensive medicine (4.7 ± 3.6 vs. 3.3 ± 2.2, p < 0.05) decreased without adverse symptoms change. In the DW_increase group, the number of adverse symptoms in 1 week (n = 14, 26 vs. 6, p < 0.05) decreased without a change in systolic blood pressure. <i>Conclusion:</i> This method may become a convenient and cheaper way to estimate DW in HD patients