New strategy of bone marrow mesenchymal stem cells against oxidative stress injury via Nrf2 pathway: oxidative stress preconditioning.

Clinically, bone marrow mesenchymal stem cells (BMSCs) have been used in treatment of many diseases, but the local oxidative stress (OS) of lesion severely limits the survival of BMSCs, which reduces the efficacy of BMSCs transplantation. Therefore, enhancing the anti-OS stress ability of BMSCs is a key breakthrough point. Preconditioning is a common protective mechanism for cells or body. Here, the aim of this study was to investigate the effects of OS preconditioning on the anti-OS ability of BMSCs and its mechanism. Fortunately, OS preconditioning can increase the expression of superoxide dismutase, catalase, NQO1, and heme oxygenase 1 through the nuclear factor erythroid 2-related factor 2 pathway, thereby decreased the intracellular reactive oxygen species (ROS) levels, relieved the damage of ROS to mitochondria, DNA and cell membrane, enhanced the anti-OS ability of BMSCs, and promoted the survival of BMSCs under OS.

PFKP is a prospective prognostic, diagnostic, immunological and drug sensitivity predictor across pan-cancer.

Phosphofructokinase, platelet (PFKP) is a rate-limiting enzyme of glycolysis that plays a decisive role in various human physio-pathological processes. PFKP has been reported to have multiple functions in different cancer types, including lung cancer and breast cancer. However, no systematic pancancer analysis of PFKP has been performed; this type of analysis could elucidate the clinical value of PFKP in terms of diagnosis, prognosis, drug sensitivity, and immunological correlation. Systematic bioinformation analysis of PFKP was performed based on several public datasets, including The Cancer Genome Atlas (TCGA), Cancer Cell Line Encyclopedia (CCLE), Genotype-Tissue Expression Project (GTEx), and Human Protein Atlas (HPA). Prospective carcinogenesis of PFKP across cancers was estimated by expression analysis, effect on patient prognosis, diagnosis significance evaluation, and immunity regulation estimation. Then, pancancer functional enrichment of PFKP was also assessed through its effect on the signaling score and gene expression profile. Finally, upstream expression regulation of PFKP was explored by promoter DNA methylation and transcription factor (TF) prediction. Our analysis revealed that high expression of PFKP was found in most cancer types. Additionally, a high level of PFKP displayed a significant correlation with poor prognosis in patients across cancers. The diagnostic value of PFKP was performed based on its positive correlation with programmed cell death-ligand 1 (PD-L1). We also found an obvious immune-regulating effect of PFKP in most cancer types. PFKP also had a strong negative correlation with several cancer drugs. Finally, ectopic expression of PFKP may depend on DNA methylation and several predicated transcription factors, including the KLF (KLF transcription factor) and Sp (Sp transcription factor) families. This pancancer analysis revealed that a high expression level of PFKP might be a useful biomarker and predictor in most cancer types. Additionally, the performance of PFKP across cancers also suggested its meaningful role in cancer immunity regulation, even in immunotherapy and drug resistance. Overall, PFKP might be explored as an auxiliary monitor for pancancer early prognosis and diagnosis.

A Robust Prognostic Gene Signature Based on eRNAs-Driven Genes in Prostate Cancer.

Prostate cancer (PCa) is the second most common malignancy in men, but its exact pathogenetic mechanisms remain unclear. This study explores the effect of enhancer RNAs (eRNAs) in PCa. Firstly, we screened eRNAs and eRNA -driven genes from The Cancer Genome Atlas (TCGA) database, which are related to the disease-free survival (DFS) of PCa patients;. screening methods included bootstrapping, Kaplan-Meier (KM) survival analysis, and Pearson correlation analysis. Then, a risk score model was established using multivariate Cox analysis, and the results were validated in three independent cohorts. Finally, we explored the function of eRNA-driven genes through enrichment analysis and analyzed drug sensitivity on datasets from the Genomics of Drug Sensitivity in Cancer database. We constructed and validated a robust prognostic gene signature involving three eRNA-driven genes namely MAPK15, ZNF467, and MC1R. Moreover, we evaluated the function of eRNA-driven genes associated with tumor microenvironment (TME) and tumor mutational burden (TMB), and identified remarkable differences in drug sensitivity between high- and low-risk groups. This study identified a prognostic gene signature, which provides new insights into the role of eRNAs and eRNA-driven genes while assisting clinicians to determine the prognosis and appropriate treatment options for patients with PCa.

[Oxidative stress preconditioning alleviates oxidative stress-induced damage of bone marrow mesenchymal stem cells].

Objective To investigate the effect of oxidative stress preconditioning on the oxidative stress-induced damage of bone marrow mesenchymal stem cells (BMSCs). Methods BMSCs were isolated and cultured by density gradient centrifugation combined with adherence method. The cells were divided into three groups: control group (control medium), oxidative damage group (treated with 1000 µmol/L H2O2 for 24 hours), and preconditioning group (preincubated with 50 µmol/L H2O2 for 8 hours before treatment with 1000 µmol/L H2O2 for 24 hours). DCFH-DA staining was used to analyze the level of reactive oxygen species (ROS). Mitochondrial membrane potential was measured by JC-1 staining. DNA damage was detected by TUNEL. Malondialdehyde (MDA) content was detected by thiobarbituric acid (TBA) method, and superoxide dismutase (SOD) activity was detected by water soluble tetrazolium-1 (WST-1) assay. CCK-8 assay was used to detect cell viability and flow cytometry to detect cell apoptosis. Results Compared with the oxidative damage group, the preconditioning group had reduced ROS level, reduced DNA damage, higher mitochondrial membrane potential, significantly decreased MDA content, increased SOD activity, increased cell viability, and significantly decreased apoptosis. Conclusion Oxidative stress preconditioning can enhance the anti-oxidative stress ability of BMSCs and promote its survival under oxidative stress.

[Risk factors for delirium in intensive care unit and its duration].

OBJECTIVE: To screen risk factors for delirium and its duration in intensive care unit (ICU) patients. METHODS: 1 200 patients admitted to ICU of the Second Hospital of Shanxi Medical University from May 2017 to May 2019 were enrolled. The gender, age, anesthesia mode, duration of mechanical ventilation and hypoxia, acute physiology and chronic health evaluation II (APACHE II) score, sedative drug use, and length of ICU stay were recorded. The occurrence and duration of ICU delirium were recorded. Multivariate Logistic regression analysis and multiple linear regression analysis were used to analyze the factors with statistical significance differences between the groups for screening the risk factors for delirium and its duration in ICU patients. RESULTS: 397 of 1 200 patients developed delirium, the incidence of ICU delirium was 33.1%. The duration of delirium in 189 patients (47.6%) was 1.0 day, and the duration of delirium in 397 delirium patients was 2.0 (1.5, 2.5) days. (1) Analysis of risk factors for delirium: univariate analysis showed that there was no significant difference in the incidence of ICU delirium among patients with different genders or ages. The incidence of ICU delirium in patients with duration of mechanical ventilation or hypoxia 4-9 days and ≥ 10 days was higher than that in patients with ≤ 3 days. The incidence of ICU delirium of general anesthesia and internal medicine patients was higher than that of patients with lumbar anesthesia. The incidence of ICU delirium in patients with APACHE II score ≥ 20 was higher than that in patients with ≤ 10 and 11-19. The patients with length of ICU stay > 9 days had a higher ICU delirium incidence than those ≤ 8 days. Increased incidence of ICU delirium in sedative patients was found as compared with those who did not use sedatives. Multivariate Logistic regression analysis showed that APACHE II score [odds ratio (OR) = 5.491, 95% confidence interval (95%CI) was 4.361-6.913, P < 0.001], the length of ICU stay (OR = 2.679, 95%CI was 1.822-3.941, P < 0.001) and the use of sedatives (OR = 2.479, 95%CI was 1.821-3.374, P < 0.001) were risk factors for ICU delirium. (2) Analysis of risk factors of ICU delirium duration: univariate analysis showed that there was no significant difference in ICU delirium duration in patients with different genders or ages. The duration of ICU delirium in patients with duration of mechanical ventilation or hypoxia ≥ 10 days was longer than that in patients with ≤ 3 days and 4-9 days. The duration of ICU delirium in general anesthesia and non-surgical patients was higher than that in patients with spinal anesthesia. The ICU delirium duration in patients with APACHE II score ≥ 20 was longer than that in patients with ≤ 10 and 11-19. The duration of ICU delirium in patients with the length of ICU stay > 9 days was longer than that in patients with ≤ 8 days. The duration of ICU delirium in patients on sedatives was longer than those not taking sedatives. Multiple linear regression analysis showed that the duration of ICU delirium increased by an average of 0.061 days (ß = 0.061, 95%CI was 0.032-0.090, P < 0.001) for each additional day of hypoxia (hypoxia duration was divided into three grades of ≤ 3, 4-9 and ≥ 10 days). For every one increase in APACHE II score (APACHE II score was divided into three grades of ≤ 10, 11-19 and ≥ 20), duration of ICU delirium extended an average of 0.058 days (ß = 0.058, 95%CI was 0.048-0.068, P < 0.001). ICU delirium duration increased by an average of 0.065 days in patients with length of ICU stay > 9 days as compared with those ≤ 8 days (ß = 0.065, 95%CI was 0.056-0.075, P < 0.001). On average, the duration of ICU delirium was prolonged by 0.362 days in patients on sedatives as compared with those who did not use sedatives (ß = 0.362, 95%CI was 0.234-0.490, P < 0.001). CONCLUSIONS: APACHE II score, the length of ICU stay and the use of sedatives were common risk factors for ICU delirium and its duration. The hypoxic duration was risk factors for ICU delirium duration.

[Effects of nicotinamide mononucleotide adenylyl transferase 3 on mitochondrial function and anti-oxidative stress of rabbit bone marrow mesenchymal stem cells via regulating nicotinamide adenine dinucleotide levels].

OBJECTIVE: To investigate the effect of nicotinamide mononucleotide adenosyl transferase 3 (NMNAT3) on the mitochondrial function and anti-oxidative stress of rabbit bone marrow mesenchymal stem cells (BMSCs) under oxidative stress in vitro by regulating nicotinamide adenine dinucleotide (NAD +) levels. METHODS: The bone marrow of femur and tibia of New Zealand white rabbits were extracted. BMSCs were isolated and cultured in vitro by density gradient centrifugation combined with adherent culture. The third generation cells were identified by flow cytometry and multi-directional induction. Overexpression of NMNAT3 gene was transfected into rabbit BMSCs by enhanced green fluorescent protein (EGFP) labeled lentivirus (BMSCs/Lv-NMNAT3-EGFP), and then the expression of NMNAT3 was detected by real-time fluorescence quantitative PCR (qRT-PCR) and Western blot and cell proliferation by cell counting kit 8 (CCK-8) method. BMSCs transfected with negative lentivirus (BMSCs/Lv-EGFP) and untransfected BMSCs were used as controls. The oxidative stress injury cell model was established by using H 2O 2 to treat rabbit BMSCs. According to the experimental treatment conditions, they were divided into 4 groups: Group A was normal BMSCs without H 2O 2 treatment; untransfected BMSCs, BMSCs/Lv-EGFP, and BMSCs/Lv-NMNAT3-EGFP in groups B, C, and D were treated with H 2O 2 simulated oxidative stress, respectively. The effects of NMNAT3 on the mitochondrial function of BMSCs under oxidative stress [changes of mitochondrial membrane potential, NAD + and adenosine triphosphate (ATP) levels], the changes of anti-oxidative stress ability of BMSCs [reactive oxygen species (ROS) and malondialdehyde (MDA) levels, manganese superoxide dismutase (Mn-SOD) and catalase (CAT) activities], and the effects of BMSCs on senescence and apoptosis [senescence associated-ß-galactosidase (SA-ß-gal) staining and TUNEL staining] were detected after 24 hours of treatment. RESULTS: The rabbit BMSCs were successfully isolated and cultured in vitro. The stable strain of rabbit BMSCs with high expression of NMNAT3 gene was successfully obtained by lentiviral transfection, and the expressions of NMNAT3 gene and protein significantly increased ( P<0.05). There was no significant difference in the trend of cell proliferation compared with normal BMSCs. After treatment with H 2O 2, the function of mitochondria was damaged and apoptosis increased in all groups. However, compared with groups B and C, the group D showed that the mitochondrial function of BMSCs improved, the membrane potential increased, the level of NAD + and ATP synthesis of mitochondria increased; the anti-oxidative stress ability of BMSCs enhanced, the levels of ROS and MDA decreased, and the activities of antioxidant enzymes (Mn-SOD, CAT) increased; and the proportion of SA-ß-gal positive cells and the rate of apoptosis decreased. The differences in all indicators between group D and groups B and C were significant ( P<0.05). CONCLUSION: NMNAT3 can effectively improve the mitochondrial function of rabbit BMSCs via increasing the NAD + levels, and enhance its anti-oxidative stress and improve the survival of BMSCs under oxidative stress conditions.

Preincubation with a low-dose hydrogen peroxide enhances anti-oxidative stress ability of BMSCs.

OBJECTIVE: To investigate the effects of low-concentration hydrogen peroxide pretreatment on the anti-oxidative stress of the bone marrow mesenchymal stem cells (BMSCs). METHODS: Rabbit BMSCs were isolated and cultured by density gradient centrifugation combined with the adherence method. Then, the third generation of well-grown BMSCs was continuously treated with 50-µM hydrogen peroxide (H2O2) for 8 h as the optimal pretreatment concentration and the BMSCs were continuously applied for 24 h with 500 µM H2O2, and the optimal damage concentration was determined as the oxidative stress cell model. The experiment was divided into three groups: control group, high-concentration H2O2 injury group (500 µM), and low-concentration H2O2 pretreatment group (50 µM + 500 µM). In each group, the DCFH-DA fluorescence probe was used to detect the reactive oxygen species (ROS). ELISA was used to detect the activity of superoxide dismutase (SOD) and catalase (CAT), and the TBA method was used to detect malondialdehyde (MDA). The mitochondrial membrane potential was detected by JC-1. The cell viability was detected by CCK-8 method, while flow cytometry and TUNEL/DAPI double staining were performed to detect cell apoptosis. Hence, the effect of H2O2 pretreatment on the anti-oxidative stress of BMSCs was investigated. One-way analysis of variance was performed using SPSS 19.0 statistical software, and P < 0.05 was considered statistically significant. RESULTS: A large number of typical BMSCs were obtained by density gradient centrifugation and adherent culture. The oxidative stress cell model was successfully established by 500-µM H2O2. Compared with the high-concentration H2O2 injury group, the low-concentration H2O2 pretreatment reduced the production of ROS [(62.33 ± 5.05), P < 0.05], SOD and CAT activities significantly increased (P < 0.05), and MDA levels significantly decreased (P < 0.05). The mitochondrial membrane potential fluorescence changes, the ratio of red/green fluorescence intensity of the high-concentration H2O2 injury group was less, and the ratio of the low-concentration H2O2 pretreatment group was significantly higher than that. The ratio of red/green increased by about 1.8 times (P < 0.05). The cell viability and survival rate of BMSCs were significantly increased in low-concentration H2O2 pretreatment group (P < 0.05), and the cell apoptosis rate was significantly decreased (P < 0.05). CONCLUSION: Pretreatment with low-concentration H2O2 can enhance the anti-oxidative stress ability and reduce their apoptosis of BMSCs under oxidative stress.

Preservation of CGRP in myocardium attenuates development of cardiac dysfunction in diabetic rats.

BACKGROUND: Calcitonin gene-related peptide (CGRP) plays an important role in cardiovascular regulation, which was found reduced in serum of diabetic patients. To test the hypothesis that lack of CGRP in myocardium is associated with diabetic cardiac dysfunction, which may be improved by preservation of CGRP in diabetic rats. METHODS AND RESULTS: Diabetes was induced in male Sprague-Dawley rats by streptozotocin (50mg/kg). Two groups of the diabetic rats, one fed with standard laboratory chew and another with the laboratory food plus hot pepper (containing 0.0174% of capsaicin), to stimulate production and release of CGRP. Cardiac functions were evaluated by measurements of intraventricular pressures after 8weeks of development of diabetes. Transient receptor potential vanilloid type 1 (TRPV1), CGRP, ß1-adreneregic receptor and norepinephrine were analyzed. Significantly lower levels of TRPV1 and CGRP were detected in the thoracic dorsal root ganglia (DRG) and myocardium of the diabetic animals, along with significant decline in left ventricular systolic pressure (by 24%) and heart rate (by 25%) and increase of the end-diastolic pressure (by 83%) with obvious reduction of CGRP in the DRG, by 41%, the myocardium (by 30%) and the serum (by 20%). The cardiac performance, the TRPV1 and the CGRP in the diabetic animals fed with hot pepper were well preserved. No any significant change in ß1-adreneregic receptor and norepinephrine was detected. CONCLUSION: The findings may suggest a novel mechanism underlying diabetic cardiac dysfunctions via impairing TRPV1-CGRP pathway in myocardium. Preservation of the TRPV1-CGRP mechanism may prevent the development of cardiac dysfunction in diabetes.