Nonobstructive azoospermia (NOA), one of the most severe types of male infertility, etiology often remains unclear in most cases. Therefore, this study aimed to detect four biallelic detrimental variants (0.5%) in the minichromosome maintenance domain containing 2 ( MCMDC2 ) genes in 768 NOA patients by whole-exome sequencing (WES). Hematoxylin and eosin (H&E) demonstrated that MCMDC2 deleterious variants caused meiotic arrest in three patients (c.1360G>T, c.1956G>T, and c.685C>T) and hypospermatogenesis in one patient (c.94G>T), as further confirmed through immunofluorescence (IF) staining. The single-cell RNA sequencing data indicated that MCMDC2 was substantially expressed during spermatogenesis. The variants were confirmed as deleterious and responsible for patient infertility through bioinformatics and in vitro experimental analyses. The results revealed four MCMDC2 variants related to NOA, which contributes to the current perception of the function of MCMDC2 in male fertility and presents new perspectives on the genetic etiology of NOA.
Humans ; Male ; Azoospermia/genetics* ; Meiosis/genetics* ; Spermatogenesis/genetics* ; Adult ; Exome Sequencing ; Microtubule-Associated Proteins/genetics* ; Alleles ; Infertility, Male/genetics*

Fifty whitespotted bamboo sharks (Chiloscyllium playgiosum) of both sexes were used to establish a large capacity variable domain of the new antigen receptor (VNAR) library with a total capacity of over 10⁹ colony-forming units (CFU). It was applied to screen VNARs against human serum albumin (HSA) and human transcription factor EB (TFEB), respectively. Meanwhile, VNAR libraries specific to HSA and TFEB with capacities above 10⁸ CFU were obtained following conventional immunization. These two approaches were systematically studied in terms of VNAR yield and composition. By comparing the VNAR sequences obtained from naïve and antigen-immunized libraries, we found that the complementary-determining region 3 (CDR3) of the former differs in composition from that of the latter. It shares a higher degree of homology with the naïve library. Meanwhile, the binding efficiency assessed by ELISA is also different between the naïve and antigen-immunized libraries. The binding of VNARs from the TFEB-immunized library appeared to surpass that observed with the naïve libraries, whereas the performance of VNARs from the HSA-immunized library indicated that both the immunized and naïve libraries for HSA had positive binding responses in polyclonal and monoclonal ELISA. The results are useful to develop novel diagnostic and therapeutic products based on shark VNARs.

The regulatory processes in developmental biology research are significantly influenced by long non-coding RNAs (lncRNAs). However, the dynamics of lncRNA expression during human tooth development remain poorly understood. In this research, we examined the lncRNAs present in the dental epithelium (DE) and dental mesenchyme (DM) at the late bud, cap, and early bell stages of human fetal tooth development through bulk RNA sequencing. Developmental regulators co-expressed with neighboring lncRNAs were significantly enriched in odontogenesis. Specific lncRNAs expressed in the DE and DM, such as PANCR, MIR205HG, DLX6-AS1, and DNM3OS, were identified through a combination of bulk RNA sequencing and single-cell analysis. Further subcluster analysis revealed lncRNAs specifically expressed in important regions of the tooth germ, such as the inner enamel epithelium and coronal dental papilla (CDP). Functionally, we demonstrated that CDP-specific DLX6-AS1 enhanced odontoblastic differentiation in human tooth germ mesenchymal cells and dental pulp stem cells. These findings suggest that lncRNAs could serve as valuable cell markers for tooth development and potential therapeutic targets for tooth regeneration.
Humans ; RNA, Long Noncoding/metabolism* ; Odontogenesis/genetics* ; Tooth Germ/embryology* ; Cell Differentiation ; Gene Expression Regulation, Developmental ; Mesoderm/metabolism* ; Tooth/embryology* ; Gene Expression Profiling ; Sequence Analysis, RNA ; Dental Pulp/cytology*

ObjectiveThe detection of RNA single nucleotide polymorphism (SNP) is of great importance due to their association with protein expression related to various diseases and drug responses. At present, splintR ligase-assisted methods are important approaches for RNA direct detection, but its specificity will be limited when the fidelity of ligases is not ideal. The aim of this study was to create a method to improve the specificity of splintR ligase for RNA detection. MethodsIn this study, a dual-competitive-padlock-probe (DCPLP) assay without the need for additional enzymes or reactions is proposed to improve specificity of splintR ligase ligation. To verify the method, we employed dual competitive padlock probe-mediated rolling circle amplification (DCPLP-RCA) to genotype the CYP2C9 gene. ResultsThe specificity was well improved through the competition and strand displacement of dual padlock probe, with an 83.26% reduction in nonspecific signal. By detecting synthetic RNA samples, the method demonstrated a dynamic detection range of 10 pmol/L-1 nmol/L. Furthermore, clinical samples were applied to the method to evaluate its performance, and the genotyping results were consistent with those obtained using the qPCR method. ConclusionThis study has successfully established a highly specific direct RNA SNP detection method, and provided a novel avenue for accurate identification of various types of RNAs.

Objective To study the quality comparability of human coagulation factor Ⅷ(FⅧ)produced before and after the change of factory site.Methods A comparative study was carried out on quality quantitative indexes,related im-purities and stability data of FⅧ produced before and after the change of factory site.Results The FⅧ quantitative quality before and after the change of factory site all met the quality standard,and the related impurities including aluminum resi-due,tributyl phosphate residue,polysorbate 80 residue and PEG residue all met the quality standard.Other impurities in-cluding human fibrinogen,fibronectin,plasminogen,IgA,IgM and IgG were extremely low in content and equivalent in quality.The content of VWF(von Willebrand factor)had no obvious change before and after the change of factory site,but was significantly higher than that of other domestic manufacturers'commercial products.The results of accelerated stability and long-term stability tests showed that the titer of FⅧ fluctuated within the methodological error range,and the results all met the quality standard.Conclusion The change of factory site of FⅧ has no effect on the quality.

Chimeric antigen receptor (CAR)-T cell therapy has achieved remarkable success in the treatment of hematological malignancies. Based on the immunomodulatory capability of CAR-T cells, efforts have turned toward exploring their potential in treating autoimmune diseases. Bibliometric analysis of 210 records from 128 academic journals published by 372 institutions in 40 countries/regions indicates a growing number of publications on CAR-T therapy for autoimmune diseases, covering a range of subtypes such as systemic lupus erythematosus, multiple sclerosis, among others. CAR-T therapy holds promise in mitigating several shortcomings, including the indiscriminate suppression of the immune system by traditional immunosuppressants, and non-sustaining therapeutic levels of monoclonal antibodies due to inherent pharmacokinetic constraints. By persisting and proliferating in vivo, CAR-T cells can offer a tailored and precise therapeutics. This paper reviewed preclinical experiments and clinical trials involving CAR-T and CAR-related therapies in various autoimmune diseases, incorporating innovations well-studied in the field of hematological tumors, aiming to explore a safe and effective therapeutic option for relapsed/refractory autoimmune diseases.

Objective:To study the expression of the Homeobox C6(HOXC6)gene in the homeobox family in PCa,its effect on the biological behavior of PCa cells and its action mechanism.Methods:Based on the studies of HOXC6 retrieved from the data-base of Gene Expression Profiling Interactive Analysis(GEPIA),we analyzed the expression of HOXC6 in PCa and the relationship of its expression level with the survival prognosis of the patients.We detected the expression of the HOXC6 protein in PCa tissues and cells by Western blot,stably interfered with the expression of the HOXC6 gene in human PCa DU145 and PC-3 cells and normal prosta-tic epithelial RWPE-1 cells using the siRNA plasmid,and determined the effects of HOXC6 on the proliferation,migration and inva-siveness of PCa cells by CCK8,plate cloning and scratch healing and Transwell invasion assays.Using the GEPIA database,we ana-lyzed the correlation of the Wnt tumor inhibitory factor-secreted frizzled-related protein 1(SFRP1)gene with HOXC6,and detected the expressions of HOXC6,SFRP1,Wnt and β-catenin in PC-3 cells after siRNA-HOXC6 transfection by Western blot.Results:The expression of HOXC6 was dramatically higher in the PCa than in the normal prostate tissue(P<0.01),and in the PCa cells than in the normal prostatic epithelial cells(P<0.01).Bioinformatics analysis indicated a lower survival rate of the PCa patients with a high than those with a low HOXC6 expression(P=0.011).The relative expression of the HOXC6 protein,absorbance value,number of clones formed and number of invaded cells were significantly lower in the siRNA group than in the negative controls(P<0.05).Ac-cording to the GEPIA database,highly expressed SFRP1 was associated with a good prognosis of PCa,and the protein expressions of Wnt and β-catenin were markedly increased while that of SFRP1 decreased in the PCa PC-3 cell line(P<0.05).The expressions of the Wnt and β-catenin proteins were decreased and that of SFRP1 increased significantly in the siRNA-HOXC6 transfection group com-pared with those in the siRNA negative control and PCa PC-3 groups(P<0.05).Conclusion:HOXC6 is highly expressed in PCa tissues and related to the proliferation,migration and invasiveness of PCa cells.HOXC6 promotes the growth of DU145 and PC-3 cells in PCa by inhibiting the SFRP1/Wnt/β-catenin signaling pathway,and may be a potential target for clinical treatment of PCa.

Objective To compare the clinical efficacy of the posterior hemipelvectomy of the greater trochanter approach with the standard Kocher-Langenbeck(K-L)approach in the treatment of posterior acetabulum wall fractures and to explore a more optimal approach for the treatment of posterior acetabulum wall fractures.Methods Total of 26 patients with posterior ac-etabulum wall fractures were retrospectively analysed and divided into two groups:the posterior hemipelvectomy of the greater trochanter group(test group)and the standard K-L approach group(control group).In the test group,there were 24 patients including 16 males and 8 females with an average age of(42.00±4.52)years old,the time of injury to surgery was(6.75±1.15)d.In the control group,there were 23 patients including 16 males and 7 females with an average age of(41.00±5.82)years old,the time of injury to surgery was(7.09±1.20)days.The total hospital stay,length of incision,operation time,intraoperative bleeding,postoperative drainage,discharge,fracture reduction quality(Matta criteria),hip abduction muscle strength,hip func-tion(Merle d'Aubigne-Postel score),postoperative complications and the incidence of ectopic ossification were compared.Results All cases were followed up for 6 months.There was no significant difference in incision length,intraoperative bleeding and postoperative drainage between two groups(P＞0.05).However,the operation time of the test group was shorter than that of the control group(P＜0.05).There was no statistically significant difference in fracture reduction and hip function between two groups(P＞0.05).The hip abduction muscle strength of test group was better than that of control group(P＜0.05).In addition,there was no significant difference in the incidence of postoperative complications and heterotopic ossification between two groups(P＞0.05).Conclusion Compared with the standard K-L approach,the posterior hemipelvectomy of the greater trochanter approach can shorten the operative time,has better recovery of the postoperative hip abduction muscle strength,ex-poses the view of the fracture involving the more comminuted posterior acetabulum wall or the fracture of the roof of the socket,improved the rate of fracture anatomical repositioning,provides a new idea for the clinical treatment of posterior acetabulum wall fractures,and allows patients to perform functional exercises at an early stage.

Hepatosplenic candidiasis (HSC) is a rare type of candidiasis that can occur in patients with hematologic malignancies, hematopoietic stem cell transplantation. At present, there is still a lack of studies on HSC in patients with hematologic disorders. Based on The Chinese Guidelines for the Diagnosis and Treatment of Invasive Fungal Disease in Patients with Hematological Disorders and Cancers (the 6th revision), We retrospectively analyzed the clinical characteristics and prognosis of patients with HSC treated in Peking University Institute of Hematology from 2008 to 2022. Finally, eighteen patients were included, with 1 (5.6%) proven, 2 (11.1%) probable, and 15 (83.3%) possible HSC. Among them, 3 (16.7%) patients occurred after haploid hematopoietic stem cell transplantation and 15 (83.3%) patients occurred after chemotherapy. 6 (33.3%) patients had positive blood cultures, including 4 cases of Candida tropicalis and 2 cases of Candida albicans. At 4 weeks of antifungal therapy, 10 (58.8%) patients achieved partial response (PR), At 8 weeks, 1 (6.3%) patients achieved complete response and 10 (62.5%) patients achieved PR. At 6 months after diagnosis, 3 (16.7%) patients died of hematopoietic recurrence, and none of them died of HSC. As a rare fungal infection disease, HSC has a low positive rate of microbiological and histological examinations, a persistent treat cycle, and has difficulty in remission, reminding us of the need for vigilance in patients with hematopoietic disorders and persistent fever.