N6-methyladenosine (m6A) modification, as a widespread and high-profile type of epigenetic modification, can dynamically and reversibly regulate the whole process of RNA metabolism. This modification governs RNA stability, splicing, and translation via m6A regulators, which are categorized as Writers, Erasers, and Readers. m6A modification also plays a critical role in the development of tumors. Disruptions in the homeostasis of m6A modification levels lead to dysregulation of m6A regulators. Consequently, these dysregulated regulators modulate the proliferation, migration, and invasion of tumor cells and interfere with the normal functions of suppressor genes and oncogenes. This article aims to comprehensively elucidate the specific regulatory impacts of m6A modification on tumor-related gene expression. It focuses on the regulatory mechanisms of m6A modification on mRNA stability. Additionally, it explores the influence of m6A on the molecular translation of key signaling pathways. Moreover, it investigates the indirect regulatory effects mediated by non-coding RNAs (ncRNAs), etc. The intention is to offer a novel analysis of the pathogenesis of cancer at a new level, and also provide a theoretical basis for finding new effective treatment methods.

Objective:To establish a model for the combined detection of serum copeptin and inflammatory markers in acute stroke (AS), and to explore the value of copeptin and inflammatory marker detection in the clinical diagnosis and prognosis assessment of AS.Methods:A total of 75 patients were diagnosed with acute ischemic stroke (AIS) [46 males, age (64.1±11.7) years] and 45 patients with acute intracerebral hemorrhage (ICH) [28 males, age (61.0±13.9) years] who were admitted to the First Affiliated Hospital of Hunan University of Chinese Medicine through the emergency department from January 1 to July 31, 2024, were included as the observation group. Meanwhile, 60 healthy individuals [39 males, age (64.4±8.2) years] were selected as the control group (HC). The differences in serum copeptin levels and inflammatory markers among different groups were compared. ROC curves were drawn to analyze the value of copeptin and inflammatory markers in the clinical diagnosis and prognosis assessment of AIS. The Kaplan-Meier method was used to draw survival curves to analyze the in-hospital survival rates of patients in different groups. Cox regression analysis was conducted to identify the risk factors affecting the prognosis of AIS patients.Results:The level of copeptin was significantly elevated in AS, with the results showing ICH>AIS>HC ( H=100.11, P<0.001). Copeptin demonstrated the highest efficacy in the early diagnosis of AIS and ICH (AUC=0.893, sensitivity 89.3%, specificity 75.0%; AUC=0.986, sensitivity 95.6%, specificity 93.3%) and the assessment of prognosis (AUC=0.997, sensitivity 100%, specificity 96.8%; AUC=0.907, sensitivity 86.7%, specificity 86.7%), outperforming other single indicators. The combined detection of copeptin with the neutrophil-to-lymphocyte ratio (NLR) and the systemic immune-inflammation index (SIIRI) was the best combination for the early diagnosis of AIS and ICH (AUC=0.937, sensitivity 77.3%, specificity 98.3%; AUC=0.989, sensitivity 95.6%, specificity 95.0%) and for the assessment of prognosis (AUC=0.996, sensitivity 100%, specificity 96.8%; AUC=0.944, sensitivity 86.7%, specificity 90.0%). Kaplan-Meier survival curves showed that AIS patients in the low-value group of copeptin and NLR had a higher survival rate during hospitalization than those in the high-value group ( HR 54.46, 7.608, P<0.01, respectively), and ICH patients in the low-value group of copeptin, SIIRI, SIRI, and SII had a higher survival rate during hospitalization than those in the high-value group ( HR 12.67, 7.923, 3.567, 5.925, P<0.05); Cox regression showed that copeptin, NLR, NIHSS, and mRS were independent risk factors affecting the prognosis of patients with AIS ( HR 1.421, 1.368, 1.158, and 1.188, respectively, P<0.05), copeptin and SIIRI were independent risk factors affecting the prognosis of ICH ( HR 1.308, 1.113, P<0.05), and GCS was a protective factor affecting ICH prognosis ( HR=0.741, P<0.05). Conclusion:Copeptin and inflammatory indicators can reflect the severity of different subtypes of stroke. The single or combined detection shows good value in the clinical application of AS. The combination of copeptin-NLR and copeptin-SIIRI is respectively the best comprehensive biomarker combination for the early diagnosis and prognosis assessment of AIS and ICH.

N6-methyladenosine (m6A) modification, as a widespread and high-profile type of epigenetic modification, can dynamically and reversibly regulate the whole process of RNA metabolism. This modification governs RNA stability, splicing, and translation via m6A regulators, which are categorized as Writers, Erasers, and Readers. m6A modification also plays a critical role in the development of tumors. Disruptions in the homeostasis of m6A modification levels lead to dysregulation of m6A regulators. Consequently, these dysregulated regulators modulate the proliferation, migration, and invasion of tumor cells and interfere with the normal functions of suppressor genes and oncogenes. This article aims to comprehensively elucidate the specific regulatory impacts of m6A modification on tumor-related gene expression. It focuses on the regulatory mechanisms of m6A modification on mRNA stability. Additionally, it explores the influence of m6A on the molecular translation of key signaling pathways. Moreover, it investigates the indirect regulatory effects mediated by non-coding RNAs (ncRNAs), etc. The intention is to offer a novel analysis of the pathogenesis of cancer at a new level, and also provide a theoretical basis for finding new effective treatment methods.

Objective:To establish a model for the combined detection of serum copeptin and inflammatory markers in acute stroke (AS), and to explore the value of copeptin and inflammatory marker detection in the clinical diagnosis and prognosis assessment of AS.Methods:A total of 75 patients were diagnosed with acute ischemic stroke (AIS) [46 males, age (64.1±11.7) years] and 45 patients with acute intracerebral hemorrhage (ICH) [28 males, age (61.0±13.9) years] who were admitted to the First Affiliated Hospital of Hunan University of Chinese Medicine through the emergency department from January 1 to July 31, 2024, were included as the observation group. Meanwhile, 60 healthy individuals [39 males, age (64.4±8.2) years] were selected as the control group (HC). The differences in serum copeptin levels and inflammatory markers among different groups were compared. ROC curves were drawn to analyze the value of copeptin and inflammatory markers in the clinical diagnosis and prognosis assessment of AIS. The Kaplan-Meier method was used to draw survival curves to analyze the in-hospital survival rates of patients in different groups. Cox regression analysis was conducted to identify the risk factors affecting the prognosis of AIS patients.Results:The level of copeptin was significantly elevated in AS, with the results showing ICH>AIS>HC ( H=100.11, P<0.001). Copeptin demonstrated the highest efficacy in the early diagnosis of AIS and ICH (AUC=0.893, sensitivity 89.3%, specificity 75.0%; AUC=0.986, sensitivity 95.6%, specificity 93.3%) and the assessment of prognosis (AUC=0.997, sensitivity 100%, specificity 96.8%; AUC=0.907, sensitivity 86.7%, specificity 86.7%), outperforming other single indicators. The combined detection of copeptin with the neutrophil-to-lymphocyte ratio (NLR) and the systemic immune-inflammation index (SIIRI) was the best combination for the early diagnosis of AIS and ICH (AUC=0.937, sensitivity 77.3%, specificity 98.3%; AUC=0.989, sensitivity 95.6%, specificity 95.0%) and for the assessment of prognosis (AUC=0.996, sensitivity 100%, specificity 96.8%; AUC=0.944, sensitivity 86.7%, specificity 90.0%). Kaplan-Meier survival curves showed that AIS patients in the low-value group of copeptin and NLR had a higher survival rate during hospitalization than those in the high-value group ( HR 54.46, 7.608, P<0.01, respectively), and ICH patients in the low-value group of copeptin, SIIRI, SIRI, and SII had a higher survival rate during hospitalization than those in the high-value group ( HR 12.67, 7.923, 3.567, 5.925, P<0.05); Cox regression showed that copeptin, NLR, NIHSS, and mRS were independent risk factors affecting the prognosis of patients with AIS ( HR 1.421, 1.368, 1.158, and 1.188, respectively, P<0.05), copeptin and SIIRI were independent risk factors affecting the prognosis of ICH ( HR 1.308, 1.113, P<0.05), and GCS was a protective factor affecting ICH prognosis ( HR=0.741, P<0.05). Conclusion:Copeptin and inflammatory indicators can reflect the severity of different subtypes of stroke. The single or combined detection shows good value in the clinical application of AS. The combination of copeptin-NLR and copeptin-SIIRI is respectively the best comprehensive biomarker combination for the early diagnosis and prognosis assessment of AIS and ICH.

Purpose: Isocitrate dehydrogenase 1 (IDH1) mutations are the most common genetic abnormalities in low-grade gliomas and secondary glioblastomas. Glioma patients with these mutations had better clinical outcomes. However, the effect of IDH1 mutation on drug sensitivity is still under debate. Materials and Methods: IDH1-R132H mutant cells were established by lentivirus. IDH1-R132H protein expression was confirmed by western blot. The expression of ataxia telangiectasia mutated (ATM) signaling pathway and apoptosis-related proteins were detected by immunofluorescence and western blot. Temozolomide (TMZ) induced cell apoptosis was detected by flow cytometry. Tumor cell proliferation was detected by Cell Counting Kit-8. In vivo nude mice were used to confirm the in vitro roles of IDH1 mutation. Results: We established glioma cell lines that expressed IDH1-R132H mutation stably. We found that TMZ inhibited glioma cells proliferation more significantly in IDH1 mutant cells compared to wild type. The IC50 of TMZ in IDH1-R132H mutant group was less than half that of wild-type group (p < 0.01). TMZ significantly induced more DNA damage (quantification of γH2AX expression in IDH1 mutation vs. wild type, p < 0.05) and apoptosis (quantification of AnnexinV+propidium iodide–cells in IDH1 mutation versus wild type, p < 0.01) in IDH1 mutant gliomas compared to wild-type gliomas. The ATM-associated DNA repair signal was impaired in IDH1 mutant cells. Inhibiting the ATM/checkpoint kinase 2DNA repair pathway further sensitized IDH1 mutant glioma cells to chemotherapy. We found that IDH1 mutation significantly inhibited tumor growth in vivo (the tumor size was analyzed statistically, p < 0.05). Moreover, we confirmed that gliomas with IDH1 mutation were more sensitive to TMZ in vivo compared to wild type significantly and the results were consistent with the in vitro experiment. Conclusion These results provide evidence that combination of TMZ and ATM inhibitor enhances the antitumor effect in IDH1 mutant gliomas.

Purpose: Isocitrate dehydrogenase 1 (IDH1) mutations are the most common genetic abnormalities in low-grade gliomas and secondary glioblastomas. Glioma patients with these mutations had better clinical outcomes. However, the effect of IDH1 mutation on drug sensitivity is still under debate. Materials and Methods: IDH1-R132H mutant cells were established by lentivirus. IDH1-R132H protein expression was confirmed by western blot. The expression of ataxia telangiectasia mutated (ATM) signaling pathway and apoptosis-related proteins were detected by immunofluorescence and western blot. Temozolomide (TMZ) induced cell apoptosis was detected by flow cytometry. Tumor cell proliferation was detected by Cell Counting Kit-8. In vivo nude mice were used to confirm the in vitro roles of IDH1 mutation. Results: We established glioma cell lines that expressed IDH1-R132H mutation stably. We found that TMZ inhibited glioma cells proliferation more significantly in IDH1 mutant cells compared to wild type. The IC50 of TMZ in IDH1-R132H mutant group was less than half that of wild-type group (p < 0.01). TMZ significantly induced more DNA damage (quantification of γH2AX expression in IDH1 mutation vs. wild type, p < 0.05) and apoptosis (quantification of AnnexinV+propidium iodide–cells in IDH1 mutation versus wild type, p < 0.01) in IDH1 mutant gliomas compared to wild-type gliomas. The ATM-associated DNA repair signal was impaired in IDH1 mutant cells. Inhibiting the ATM/checkpoint kinase 2DNA repair pathway further sensitized IDH1 mutant glioma cells to chemotherapy. We found that IDH1 mutation significantly inhibited tumor growth in vivo (the tumor size was analyzed statistically, p < 0.05). Moreover, we confirmed that gliomas with IDH1 mutation were more sensitive to TMZ in vivo compared to wild type significantly and the results were consistent with the in vitro experiment. Conclusion These results provide evidence that combination of TMZ and ATM inhibitor enhances the antitumor effect in IDH1 mutant gliomas.

To construct a eukaryotic expression plasmid containing the luciferase reporter gene (Fluc) to quickly detect apoptosis. Four amino acids, Asp-Glu-Val-Asp (DEVD), the recognize motif of Caspase-3, were introduced into the middle of the Fluc-C and N fragment. Meanwhile, four amino acids, Asp-Glu-Val-Gly (DEVG), were selected as a negative control. Subsequently, the recombinant gene was cloned into the N and C terminal end of the split intein, and named as pFluc-DEVD and pFluc-DEVG. Then the plasmids were transfected into cells and renilla luciferase was co-transfected in each sample as an internal control for transfection efficiency. Then the apoptosis level was detected by the double luciferase reporter gene and the Western blotting analysis. The results showed that when apoptosis occurred, the content of firefly luciferase expressed in the pFluc-DEVD plasmid transfected group was about 3 times higher than pFluc-DEVG plasmid transfected group. Furthermore, Western blotting detection indicated that the Fluc level was significantly increased in pFluc-DEVD transfected group when pre-treated by apoptosis stimulants. The activation degree of Caspase-3 was closely related to the expression of Fluc, and had a significant statistical difference. These results confirmed that firefly luciferase protein expressed by pFluc-DEVD plasmid can be cleaved by the intracellular Caspase-3 enzyme, and this plasmid can accurately reflect the cell apoptosis level, which provides a useful method for quantitative detection of apoptosis.
Apoptosis ; Genes, Reporter ; Luciferases, Firefly ; Transfection

Objective To investigate the efficacy and safety of mesenchymalstem cells (MSCs)in improving the renal transplantation functional recovery of donation after citizens death (DCD)derived renal donors.Methods A retrospective analysis of 97 cases of DCD renal transplantation was performed in our center from July 2011 to December 2016.Among them,50 cases were treated with MSCs (the treatment group) and 47 cases without MSCs (the control group).In the treatment group,the umbilical cord stem cell suspension (50 mL,approximately 1 106/kg weight) was infused before vascular patency and 1 week,2 weeks,and 3 weeks after renal transplantation.Postoperative renal function within 3 months of recovery,delayed graft function (DGF) incidence and duration,the incidence of pulmonary infection,acute rejection and surgical complications were analyzed.Resuts The incidence rate of DGF and duration in the treatment group were 16.0％ and (14.42 ± 3.95) days,and those in the control group were 27.7％ and (17.85 ± 6.25) days respectively,and there were statistically significant difference.The incidence of acute rejection in the treatment group and the control group was 12.0 and 21.3％ respectively with the difference being statistically significant.The incidence rate of pulmonary infection in the treatment group and control group was 10％ and 12.7％respectively with the different being not statistically significant.Comparison of eGFR at 1st,1st month and 3rd month after operation showed that the eGFR in the control group was statistically significantly lower than that in the treatment group at 1st and 3rd month,but there was no significant difference in eGFR at 1st week between the two groups.There was no significant difference in the surgical complications between the two groups.Conclusion In the treatment group,the incidence rate of DGF was lower,and DGF duration was shorter than in the control group.The acute rejection rate in the control group increased significantly,but there was no significant difference in pulmonary infection and surgical complications between two groups.The recovery trend of renal function in the treatment group was better than that in the control group.

BACKGROUND:Bone mesenchymal stem cells have immunological regulation function both in vitro and in vivo, while the effect of bone marrow mesenchymal stem cells on CD4+T cellimmune function in patients receiving kidney transplantation remains unclear. OBJECTIVE:To explore the monitoring significance of CD4+T-cellimmune function by ImmuKnow assay and to determine the effect of induction therapy with bone marrow mesenchymal stem cells on cellimmune function in patients receiving kidney transplantation. METHODS:From January 2011 to June 2013, 24 patients receiving al ograft renal transplantation with autologous bone marrow mesenchymal stem cells were included and another 48 patients receiving al ograft renal transplantation and Simulect induction therapy with various matched preoperative characters served as controls. In both groups, adenosine triphosphate levels in CD4+T cells in the peripheral blood were determined by the ImmuKnow assay preoperatively and at 14, 30, 60, 90, 180 days postoperatively, as wel as during acute rejection and infection episodes. RESULTS AND CONCLUSION:During the 180 days postoperatively, fewer patients in the bone marrow mesenchymal stem cellgroup had acute rejection and injection than the Simulect group, but no significant differences were observed. Postoperative adenosine triphosphate levels in CD4+T cells were significantly lower than those determined preoperatively in both groups (P<0.05), while no significant differences were observed between the two groups. A total of 12 patients in the bone marrow mesenchymal stem cellgroup and 26 patients in the Simulect group had infection episodes, and the adenosine triphosphate levels in CD4+T cells during the infection episodes were lower than clinical stable patients in both groups (P<0.01). For patients receiving renal transplantation, induction therapy with bone marrow mesenchymal stem cells can effectively decrease the cellimmune function, which can be reflected by the adenosine triphosphate levels in CD4+T cells in the peripheral blood determined by the ImmuKnow assay.

Objective To evaluate the values of cell-mediated immune function in elderly renal allograft recipients.Method The levels of immuknowTM ATP was sequentially monitored by means of Cylex immuknowTM assay in 52 elderly renal allograft recipients including 11 with infection and 8 with acute rejection.Results No statistically significant difference was found between stable allograt function and uremia (P＞0.05).The levels of immuknowTM ATP during infection was significantly lower than those with stable allograft function with acute rejection (P ＜ 0.01).The levels of immuknowTM ATP during acute rejection was significantly higher than those with stable allograft function with infection (P＜0.01).Conclusion Sequential monitoring of immuknowTM ATP is helpful for elderly renal allograft recipients in individualized immunosuppression therapy.Cylex immuknowTM assay can be used as a potent tool for assessment of high risk in infection and rejection.