Diabetic cardiomyopathy (DCM) is a medical condition characterized by cardiac remodeling and dysfunction in individuals with diabetes mellitus. Sarcoplasmic reticulum (SR) and mitochondrial Ca²⁺ overload in cardiomyocytes have been recognized as biological hallmarks in DCM; however, the specific factors underlying these abnormalities remain largely unknown. In this study, we aimed to investigate the role of a cardiac-specific long noncoding RNA, D830005E20Rik (Trdn-as), in DCM. Our results revealed the remarkably upregulation of Trdn-as in the hearts of the DCM mice and cardiomyocytes treated with high glucose (HG). Knocking down Trdn-as in cardiac tissues significantly improved cardiac dysfunction and remodeling in the DCM mice. Conversely, Trdn-as overexpression resulted in cardiac damage resembling that observed in the DCM mice. At the cellular level, Trdn-as induced Ca²⁺ overload in the SR and mitochondria, leading to mitochondrial dysfunction. RNA-seq and bioinformatics analyses identified calsequestrin 2 (Casq2), a primary calcium-binding protein in the junctional SR, as a potential target of Trdn-as. Further investigations revealed that Trdn-as facilitated the recruitment of METTL14 to the Casq2 mRNA, thereby enhancing the m⁶A modification of Casq2. This modification increased the stability of Casq2 mRNA and subsequently led to increased protein expression. When Casq2 was knocked down, the promoting effects of Trdn-as on Ca²⁺ overload and mitochondrial damage were mitigated. These findings provide valuable insights into the pathogenesis of DCM and suggest Trdn-as as a potential therapeutic target for this condition.
Animals ; Diabetic Cardiomyopathies/pathology* ; RNA, Long Noncoding/genetics* ; Myocytes, Cardiac/metabolism* ; Mice ; Calsequestrin/genetics* ; Calcium/metabolism* ; Male ; Sarcoplasmic Reticulum/metabolism* ; Methyltransferases/metabolism* ; Mice, Inbred C57BL ; Mitochondria, Heart/metabolism* ; Disease Models, Animal ; Mitochondria/metabolism*

Due to physiological and metabolic characteristics and disease characteristics being different from adults, such as the high incidence of infectious diseases, children are at a high-risk of acute kidney injury (AKI) related to anti-bacterial agents, and there were many challenges in prevention and treatment. The hot issues in the research of anti-bacterial agent-related AKI in children, including the study of nephrotoxicity mechanism of anti-bacterial agents, the exploration of early biomarkers, and the optimization of treatment strategies, are expounded in this article. In the future, it is recommended to strengthen researches on child specificity, deepen basic researches, optimize monitoring plans for pediatric treatment drugs, and establish a pediatric drug monitoring system, so as to provide references for rational clinical drug use of the anti-bacterial agents and further researches on AKI.

Due to physiological and metabolic characteristics and disease characteristics being different from adults, such as the high incidence of infectious diseases, children are at a high-risk of acute kidney injury (AKI) related to anti-bacterial agents, and there were many challenges in prevention and treatment. The hot issues in the research of anti-bacterial agent-related AKI in children, including the study of nephrotoxicity mechanism of anti-bacterial agents, the exploration of early biomarkers, and the optimization of treatment strategies, are expounded in this article. In the future, it is recommended to strengthen researches on child specificity, deepen basic researches, optimize monitoring plans for pediatric treatment drugs, and establish a pediatric drug monitoring system, so as to provide references for rational clinical drug use of the anti-bacterial agents and further researches on AKI.

Objective:To characterize the minimal inhibitory concentration (MIC) of macrolides against clinical Mycoplasma pneumoniae (MP) isolates and to investigate the significance of 23S rRNA mutations. Methods:Cross-sectional study.A total of 288 clinical MP strains preserved in the laboratory from 2016 to 2021 were taken for macrolide resistance gene testing and the evaluation of in vitro susceptibility to Azithromycin and Erythromycin.MIC 50 and MIC 90 values were calculated separately for macrolide-susceptible and -resistant strains. Results:All 288 MP strains underwent the test of in vitro susceptibility to Azithromycin, while 86 of them were additionally tested for Erythromycin.Among these strains, 22 strains were Azithromycin-sensitive, and 266 strains were Azithromycin-resistant.A2063G mutations were detected in 260 (97.7%) strains, while A2064G mutations were detected in 6 (2.3%) strains.Azithromycin-resistant strains had an MIC 50 of 128.000 μg/mL and an MIC 90 of 512.000 μg/mL, with the MIC ranging between 16.000 and 512.000 μg/mL.Seven strains were sensitive and 79 strains were resistant to Erythromycin.Among Erythromycin-resistant strains, A2063G mutations were detected in 73 (92.4%) strains, while A2064G mutations were detected in 6 (7.6%) strains.Erythromycin-resistant strains had an MIC 50 of 256.000 μg/mL and an MIC 90 of 512.000 μg/mL, with the MIC ranging between 64.000 and 1 024.000 μg/mL. Conclusions:A2063G and A2064G mutations in the 23S rRNA gene of MP are associated with high-level in vitro resistance to Azithromycin and Erythromycin, significantly limiting the clinical effectiveness of these antibiotics.Early resistance gene testing is recommended for suspected MP patients, which can help optimize the treatment, improve prognosis, and prevent resistance spread.

Objective:To characterize the minimal inhibitory concentration (MIC) of macrolides against clinical Mycoplasma pneumoniae (MP) isolates and to investigate the significance of 23S rRNA mutations. Methods:Cross-sectional study.A total of 288 clinical MP strains preserved in the laboratory from 2016 to 2021 were taken for macrolide resistance gene testing and the evaluation of in vitro susceptibility to Azithromycin and Erythromycin.MIC 50 and MIC 90 values were calculated separately for macrolide-susceptible and -resistant strains. Results:All 288 MP strains underwent the test of in vitro susceptibility to Azithromycin, while 86 of them were additionally tested for Erythromycin.Among these strains, 22 strains were Azithromycin-sensitive, and 266 strains were Azithromycin-resistant.A2063G mutations were detected in 260 (97.7%) strains, while A2064G mutations were detected in 6 (2.3%) strains.Azithromycin-resistant strains had an MIC 50 of 128.000 μg/mL and an MIC 90 of 512.000 μg/mL, with the MIC ranging between 16.000 and 512.000 μg/mL.Seven strains were sensitive and 79 strains were resistant to Erythromycin.Among Erythromycin-resistant strains, A2063G mutations were detected in 73 (92.4%) strains, while A2064G mutations were detected in 6 (7.6%) strains.Erythromycin-resistant strains had an MIC 50 of 256.000 μg/mL and an MIC 90 of 512.000 μg/mL, with the MIC ranging between 64.000 and 1 024.000 μg/mL. Conclusions:A2063G and A2064G mutations in the 23S rRNA gene of MP are associated with high-level in vitro resistance to Azithromycin and Erythromycin, significantly limiting the clinical effectiveness of these antibiotics.Early resistance gene testing is recommended for suspected MP patients, which can help optimize the treatment, improve prognosis, and prevent resistance spread.

Objective:To investigate the efficacy and safety of Rituximab (RTX) in the treatment of children with frequently relapsing/steroid-dependent nephrotic syndrome (FRNS/SDNS) and to analyze the factors influencing the efficacy.Methods:Case series study.The clinical data of children with FRNS/SDNS who received B-cell-guided RTX (single dose: 375 mg/m 2, maximum dose: 500 mg, one additional dose when peripheral blood CD19 + B lymphocytes ≥0.01) in the First Affiliated Hospital of Zhengzhou University from September 2019 to March 2022 were retrospectively collected.The frequency of relapse and cumulative dose of glucocorticoids before and after RTX treatment were compared.The Kaplan-Meier method was used to analyze relapse-free survival rate and FRNS/SDNS-free survival rate after RTX treatment.The influencing factors of relapse were analyzed using the Cox proportional hazards regression model. Results:Totally 47 children were enrolled, including 35 males and 12 females; the age of first application of RTX was 10.2 (6.9, 13.0) years; 33 children had used one type of immunosuppressant before, and 14 children had used two or more types of immunosuppressant before; the dose of RTX treatment was 3.0 (2.0, 3.0). The frequency of relapse[0(0, 0.55) times/year vs.1.62 (1.09, 2.40) times/year] and cumulative dose of glucocorticoids[0.12 (0.05, 0.21) mg/(kg·d) vs.0.40 (0.20, 0.56) mg/(kg·d)] after RTX treatment significantly decreased compared with previous immunosuppressive treatment ( Z=-5.56, -5.54, all P<0.001). The relapse-free survival rates at 6, 12, 18 and 24 months after treatment were 80.9%, 72.3%, 68.1% and 68.1%, respectively, and the FRNS/SDNS-free survival rates were 93.6%, 89.4%, 89.4% and 89.4%, respectively.Univariate Cox regression analysis showed that the high frequency of relapse during previous immunosuppressive therapy was a risk factor for relapse after RTX treatment ( P<0.05). Of the 14 children who relapsed, 6 occurred in children whose CD19 + B lymphocytes<0.01, and the frequency of relapse after RTX treatment was significantly higher than those whose CD19 + B lymphocytes≥0.01 ( Z=-2.84, P=0.005). No severe adverse reactions occurred during RTX treatment and follow-up. Conclusions:The B-cell-guided RTX is effective and safe in the treatment of FRNS/SDNS in children.The high frequency of relapse during previous immunosuppressive therapy is a risk factor for relapse after RTX treatment, and relapse in the state of B lymphocyte depletion predicts poor outcomes of RTX treatment.

Objective:We genetically modified our non-replicating vaccinia virus NTV to improve its immunogenicity.Methods:We constructed NTV-modified strain NTV-ΔF1L-C7L by homologous recombination of vaccinia virus based on CRISPR-Cas9 technology by inserting the C7L gene while deleting the F1L gene. The recombinant virus NTV-ΔF1L-C7L was then immunized with 10 7 PFU in BALB/c mice, and the levels of humoral and cellular immunity induced by NTV-ΔF1L-C7L were detected by ELISA and ELISpot method, respectively, and the levels of neutralizing antibodies were determined by the phage-reduced neutralization assay. Results:The PCR and western- blot identification proved that the F1L gene of the constructed NTV-modified strain NTV-ΔF1L-C7L was missing, while the C7L gene was inserted back in the region, and the C7L gene could be expressed normally, indicating that the recombinant virus was constructed correctly. After immunization of mice with NTV-ΔF1L-C7L, ELISA result showed that the recombinant virus NTV-ΔF1L-C7L induced a higher level of IgG antibody than NTV; ELISpot result also showed that the recombinant virus was able to induce a higher level of IFN-γ; and the result of plaque reduction neutralization test showed that the recombinant virus was able to induce a higher level of IFN-γ antibody than that of NTV.Conclusions:We correctly constructed the NTV gene-modified strain NTV-ΔF1L-C7L, which induced stronger humoral and cellular immunity compared with NTV, and provided reference data for the research and development of replacement products for smallpox or monkeypox vaccines.

Humans ; Cytomegalovirus ; Cytomegalovirus Infections ; Antigens, Viral ; Immunity ; Autoimmune Diseases ; CD8-Positive T-Lymphocytes

Objective:Through the study of the cell biological characteristics and virulence in mice in vivo of three non-replicating vaccinia virus modified strains to provide reference for the development of smallpox/monkeypox vaccine replacement products.Methods:Replicating vaccinia virus Tiantan strain VTT and non-replicating Tiantan Vaccinia Strain NTV were studied in BHK-21/CEF and its modified strains NTV-C7L, NTV-△F1L-C7L, NTV-K1L were amplified, purified, and identified by Western blotting. The virulence and diffusivity of each strain in cells were evaluated by immune-plaque assay. The replication dynamics curves were used to compare the replication differences between the strains, and the weight loss was observed by intranasal route in the mouse model to compare the virulence levels of the viruses.Results:In this study, Western blotting result proved that the amplified and purified vaccinia virus strains were correct. Immunophagocytosis and replication kinetics showed that the replication capacity of the three NTV modified strains in CEF was similar to that of NTV. The diffusion ability and replication ability between Vero cells were improved, but the replication multiple was less than 100 times. The replication level of MRC-5 was significantly enhanced compared with that of NTV, and the replication ratio of NTV-C7L was more than 20000 times. The virulence in mice showed that the body weight of the three NTV modified strains had no statistical significance compared with that of NTV.Conclusions:The three NTV modified strains recovered their replication ability in human MRC-5 cells, but their virulence in mice was similar to that of NTV, which provided the preliminary conditions for being candidate strains of smallpox/monkeypox vaccine replacement products.