Objective: To investigate the precise detection methods for weak partial D type 15 and evaluate their implications for blood transfusion safety, along with the development of corresponding strategies. Methods: A combination of serological methods, including the microplate method, indirect antiglobulin tube method, and microcolumn gel card method, was employed to identify RhD-negative and RhD variant samples. RhD-negative samples were screened for the presence of RHD genes using whole-blood direct PCR amplification. Subsequently, RhD variant samples and RhD-negative samples containing RHD genes underwent full-coding-region sequencing of the RHD gene to confirm their genotypes. The genotyping results were further correlated with the serological test findings for comprehensive analysis. Results: Among 615 549 first-time healthy blood donors, 3 401 samples with an RhD-negative phenotype and 156 samples with RhD variant were identified. Of the 3 401 RhD-negative samples, 1 054 were found to harbor RHD genes. Gene sequencing analysis of the 156 RhD variants and the 1 054 serological negative samples revealed that 89 samples contained the RHD 15 (c. 845G>A) allele. Conclusion: The integration of serological testing methods and genotyping technologies for the precise determination of RhD blood type plays a critical role in ensuring the safety and compatibility of blood transfusions.

Background: In acute and chronic renal inflammatory diseases, the activation of inflammatory cells is involved in the defect of erythropoietin (EPO) production. Ras guanine nucleotide-releasing protein-4 (RasGRP4) promotes renal inflammatory injury in type 2 diabetes mellitus (T2DM). Our study aimed to investigate the role and mechanism of RasGRP4 in the production of renal EPO in diabetes. Methods: The degree of tissue injury was observed by pathological staining. Inflammatory cell infiltration was analyzed by immunohistochemical staining. Serum EPO levels were detected by enzyme-linked immunosorbent assay, and EPO production and renal interstitial fibrosis were analyzed by immunofluorescence. Quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of key inflammatory factors and the activation of signaling pathways. In vitro, the interaction between peripheral blood mononuclear cells (PBMCs) and C3H10T1/2 cells was investigated via cell coculture experiments. Results: RasGRP4 decreased the expression of hypoxia-inducible factor 2-alpha (HIF2A) via the ubiquitination–proteasome degradation pathway and promoted myofibroblastic transformation by activating critical inflammatory pathways, consequently reducing the production of EPO in T2DM mice. Conclusion RasGRP4 participates in the production of renal EPO in diabetic mice by affecting the secretion of proinflammatory cytokines in PBMCs, degrading HIF2A, and promoting the myofibroblastic transformation of C3H10T1/2 cells.

Background: In acute and chronic renal inflammatory diseases, the activation of inflammatory cells is involved in the defect of erythropoietin (EPO) production. Ras guanine nucleotide-releasing protein-4 (RasGRP4) promotes renal inflammatory injury in type 2 diabetes mellitus (T2DM). Our study aimed to investigate the role and mechanism of RasGRP4 in the production of renal EPO in diabetes. Methods: The degree of tissue injury was observed by pathological staining. Inflammatory cell infiltration was analyzed by immunohistochemical staining. Serum EPO levels were detected by enzyme-linked immunosorbent assay, and EPO production and renal interstitial fibrosis were analyzed by immunofluorescence. Quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of key inflammatory factors and the activation of signaling pathways. In vitro, the interaction between peripheral blood mononuclear cells (PBMCs) and C3H10T1/2 cells was investigated via cell coculture experiments. Results: RasGRP4 decreased the expression of hypoxia-inducible factor 2-alpha (HIF2A) via the ubiquitination–proteasome degradation pathway and promoted myofibroblastic transformation by activating critical inflammatory pathways, consequently reducing the production of EPO in T2DM mice. Conclusion RasGRP4 participates in the production of renal EPO in diabetic mice by affecting the secretion of proinflammatory cytokines in PBMCs, degrading HIF2A, and promoting the myofibroblastic transformation of C3H10T1/2 cells.

Background: In acute and chronic renal inflammatory diseases, the activation of inflammatory cells is involved in the defect of erythropoietin (EPO) production. Ras guanine nucleotide-releasing protein-4 (RasGRP4) promotes renal inflammatory injury in type 2 diabetes mellitus (T2DM). Our study aimed to investigate the role and mechanism of RasGRP4 in the production of renal EPO in diabetes. Methods: The degree of tissue injury was observed by pathological staining. Inflammatory cell infiltration was analyzed by immunohistochemical staining. Serum EPO levels were detected by enzyme-linked immunosorbent assay, and EPO production and renal interstitial fibrosis were analyzed by immunofluorescence. Quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of key inflammatory factors and the activation of signaling pathways. In vitro, the interaction between peripheral blood mononuclear cells (PBMCs) and C3H10T1/2 cells was investigated via cell coculture experiments. Results: RasGRP4 decreased the expression of hypoxia-inducible factor 2-alpha (HIF2A) via the ubiquitination–proteasome degradation pathway and promoted myofibroblastic transformation by activating critical inflammatory pathways, consequently reducing the production of EPO in T2DM mice. Conclusion RasGRP4 participates in the production of renal EPO in diabetic mice by affecting the secretion of proinflammatory cytokines in PBMCs, degrading HIF2A, and promoting the myofibroblastic transformation of C3H10T1/2 cells.

Background: In acute and chronic renal inflammatory diseases, the activation of inflammatory cells is involved in the defect of erythropoietin (EPO) production. Ras guanine nucleotide-releasing protein-4 (RasGRP4) promotes renal inflammatory injury in type 2 diabetes mellitus (T2DM). Our study aimed to investigate the role and mechanism of RasGRP4 in the production of renal EPO in diabetes. Methods: The degree of tissue injury was observed by pathological staining. Inflammatory cell infiltration was analyzed by immunohistochemical staining. Serum EPO levels were detected by enzyme-linked immunosorbent assay, and EPO production and renal interstitial fibrosis were analyzed by immunofluorescence. Quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of key inflammatory factors and the activation of signaling pathways. In vitro, the interaction between peripheral blood mononuclear cells (PBMCs) and C3H10T1/2 cells was investigated via cell coculture experiments. Results: RasGRP4 decreased the expression of hypoxia-inducible factor 2-alpha (HIF2A) via the ubiquitination–proteasome degradation pathway and promoted myofibroblastic transformation by activating critical inflammatory pathways, consequently reducing the production of EPO in T2DM mice. Conclusion RasGRP4 participates in the production of renal EPO in diabetic mice by affecting the secretion of proinflammatory cytokines in PBMCs, degrading HIF2A, and promoting the myofibroblastic transformation of C3H10T1/2 cells.

Objective:This study aimed to assess the infection status of cytomegalovirus (CMV) and Epstein-Barr virus (EBV) in healthy populations in China over the past decade and analyze the differences in CMV and EBV infection and related risk factors in healthy populations before and after the lifting of coronavirus disease 2019 (COVID-19) pandemic control measures.Methods:This study retrospectively analyzes the CMV and EBV infection status of 8 827 healthy donors who underwent prehematopoietic stem cell transplantation screening at Peking University People's Hospital from January 2014 to December 2023. Logistic regression analysis was conducted to determine the risk factors for CMV and EBV infection.Results:The CMV and EBV IgG positivity rates were 94.52% and 95.40% among the healthy donors, respectively, with no significant differences before and after the lifting of pandemic control measures (all P value>0.05). However, IgG antibody titers increased [CMV: (100.44±36.50) U/ml vs (109.98±36.31) U/ml, P<0.001; EBV: (281.57±226.79) U/ml vs (361.08±268.58) U/ml, P<0.001] after lifting the COVID-19 restrictions. However, the CMV IgM positivity rate remained unchanged. The EBV IgM positivity rate significantly increased after lifting measures (2.77% vs 6.29%, P<0.001), reaching 8.10% within 3 months. Further analysis of the factors affecting EBV IgM positivity revealed that gender ( OR=1.479, 95% CI 1.169-1.872, P=0.001), age[compared with the group younger than 18 years, the 18-50-year age group ( OR=0.584, 95% CI 0.421-0.820, P=0.002), the >50-year age group ( OR=0.389, 95% CI 0.248-0.610, P<0.001) ], and the lifting of COVID-19 restrictions ( OR=2.360, 95% CI 1.287-3.047, P<0.001) were independent factors influencing EBV IgM positivity in the general population. The EBV IgM positivity rate in individuals under 18 years old was not affected by gender or the lifting of COVID-19 restrictions when stratified by age group. Both genders ( OR=1.499, 95% CI 1.138 - 1.975, P=0.004) and the lifting of COVID-19 restrictions ( OR=2.608, 95% CI 1.940-3.507, P<0.001) were independent factors affecting EBV IgM positivity in the 18-50-year age group. The lifting of COVID-19 restrictions ( OR=2.222, 95% CI 1.101-4.484, P=0.026) was the sole independent factor affecting EBV IgM positivity in individuals over 50 years old. Conclusions:Previous infection rates of CMV and EBV are high in healthy populations in China, which increase with age. COVID-19 infection may increase EBV reactivation rates in healthy individuals, with a more pronounced effect on those aged >18 years.

Objective To establish a quality control mechanism based on the Poisson distribution for monitoring con-tamination in nucleic acid testing(NAT)laboratories.Methods The study collected NAT single-reactivity rates and dis-criminatory test reactivity rates from the Grifols Panther NAT system from 2022 to 2023.The Poisson distribution probability method was used to calculate the daily probability of nucleic acid single reactivity.Sensitivity and specificity of the quality control model were further validated using single-reactivity rates,discriminatory reactivity rates and ROC curve analysis,fol-lowed by proposing corresponding multi-rule quality control strategies.Results Using P=0.05 as the threshold for out-of-control,the Poisson distribution probability quality control model identified 40 out-of-control points with P<0.05.After man-ual verification using discriminatory reactivity rates,20 out-of-control points were confirmed.The sensitivity of determining out-of-control was 60.0%,and the specificity was 95.6%.Due to the high number of out-of-control points,it is recommen-ded to combine discriminatory reactivity rates and employ multi-rule quality control methods for quality monitoring.Under the multi-rule quality control,there were 18 warning points and 2 out-of-control points for NAT single reactivity from 2022 to 2023.Conclusion The multi-rule NAT single-reactivity monitoring mechanism based on the Poisson distribution probability is more suitable for nucleic acid laboratories to monitor and warn of laboratory contamination,thereby enhancing the manage-ment capabilities and detection quality of nucleic acid laboratories.

Objective To investigate the serological characteristics and molecular mechanism of an individual with Am phenotype.Methods The sample with ABO blood group discrepancy was confirmed by serological techniques.The full cod-ing and flanking regions of the ABO gene including intron 1 transcription factor binding site were identified through direct se-quencing of PCR-amplified products.PCR products of exon 6-7 were validated to isolate the ABO gene haplotypes by clo-ning and sequencing individual colonies.Bioinformatics software was used to analyze the structure of the mutant protein.Re-sults The serologic characteristics of ABO blood typing showed the rare Am phenotype.The c.467C/T and c.912C/A heter-ozygous sites in exon 7 were identified by direct sequencing analysis.Further TA cloning and sequencing revealed that the patient carried an ABO*O.01.01 allele and a novel ABO*A allele.The new allele sequence had one nucleotide alteration(C＞A)at position 912 on the background of the ABO*A1.02 allele.The new allele sequence has been included in the Gen-Bank database with the entry number JX489776.The c.912C＞A mutation was predicted to be"probably damaging"and"deleterious"by PolyPhen2 and PROVEAN algorithms,respectively.The free energy change(ΔΔG)value predicted it to have a destabilizing effect on the GTA protein.Meanwhile,modeling of the 3D structure predicted that the p.S304R amino acid substitution may alter the hydrogen bond of the GTA protein.Conclusion The p.S304R substitution of α-1,3-N-acetylgalactosaminyltransferase gene may reduce the antigen expression owing to a greatly destabilizing effect on the structure and function of the GTA protein.

Objective To investigate the effects of doublecortin-like kinase 1(DCLK1)on the malignant biological behaviors,such as proliferation,migration,and invasion,of A549 cell line and their corresponding mechanisms.Methods DCLK1-overexpressing A549 cell lines were established through lentiviral infection,and DCLK1 expression was validated by using RT-PCR and Western blot analysis.Proliferation ability was assessed with CCK-8 and plate cloning assays,and migration and invasion abilities were examined with Transwell assays.The pathway regulated by DCLK1 in lung adenocarcinoma was analyzed on the basis of the TCGA lung adenocarcinoma cohort with pathway enrichment analysis and verified through Western blot analysis.Results DCLK1 overexpression in A549 cells promoted cell proliferation,migration,and invasion.The inhibition of the FAK/PI3K/AKT/mTOR signaling pathway impaired the DCLK1-mediated malignant behavior of A549 cells.Conclusion DCLK1 promotes the malignant behavior of A549 cells through the activation of the FAK/PI3K/AKT/mTOR signaling pathway.