Objective To investigate the molecular characteristics of immune response signaling molecules induced by transfection of coxsackievirus B2 ( CVB2 ) structural proteins into epithelial cells. Methods Recombinant eukaryotic expression plasmids containing the coding regions of CVB2 structural proteins VP1-VP4 were constructed and then transfected into 16HBE cells. Culture supernatants and cell ly-sates of the transfected 16HBE cells were collected. Expression of signaling molecules involved in innate im-mune responses in transfected 16HBE cells at mRNA level was detected by RT-Q-PCR. The proliferation of T cells co-cultured with culture supernatants and cell lysates of the transfected 16HBE cells was analyzed by ELISPOT. Results Expression of innate immunity-related signaling molecules such as TGF-β-activated ki-nase ( TAK) , NF-κB-inducing kinase ( NIK) , IκB kinase α ( IKKα) and IFN-β at mRNA level was up-regulated in 16HBE cells transfected with CVB2 structural proteins VP1-VP4. Both culture supernatants and cell lysates of the transfected 16HBE cells enhanced the proliferation of T cells. Conclusions CVB2 struc-tural proteins VP1-VP4 could enhance the expression of innate immunity-related signaling molecules to var-ying degrees and promote the activation of adaptive immunity.

Objective:To study the characteristics of intestinal microflora in patients with vitiligo, and to analyze the relationship between the changes of intestinal microflora and the incidence of vitiligo, so as to provide new ideas for clinical treatment.Methods:Fecal specimens from 30 patients with vitiligo and 30 healthy adults were collected and analyzed qualitatively by Roche/45 high-throughput sequencing platform. At the same time, macrogenomics was used to analyze the feces of 5 patients with vitiligo and 5 healthy adults to identify the potential regulatory pathways.Results:The bacterial species in the feces of patients with vitiligo were similar to those of healthy people, but the intestinal microbial diversity of patients with vitiligo was significantly reduced ( P<0.01); the abundance of Proteus and Clostridium was significantly reduced at phylum level; at genus level, 7 of them were Bacteroides, Escherichia coli Shigella, Rochella, Bacillus anthracis, Clostridium clostridium, Jordani bacteria. The abundance of RF9 and Prunella-7 decreased significantly ( P<0.01), while the abundance of 4 genera (Rumen Coccus-1, Rumen Coccus UCG, Trichomonas and Streptococcus) increased significantly ( P<0.01). The expression of Streptococcus and Phase Anthraceae in vitiligo patients was significantly different: the former increased by 10.8 times, the latter decreased by 6.517 times, and an intestinal microorganism based on 11 vitiligo-related genera was constructed. The random forest model of bacterial flora showed that AUC of the discriminant model was 0.89 in ROC, and macrogenomic analysis showed that the disorders of vitiligo-related bacterial flora were mainly related to immune-related pathways (such as WNT pathway, Notch pathway), energy metabolism, mitochondrial function and amino acid metabolism (such as phenylalanine metabolism). Conclusions:The diversity of bacterial community in intestinal microecological environment of vitiligo patients is significantly different from those in normal people. The imbalance of bacterial community may be involved in the pathogenesis and development of vitiligo. Supplementation of probiotics may be beneficial to the treatment of vitiligo.

Objective To compare the discrepancies among results of three commonly used laboratory direct test methods for maximal oxygen uptake(VO2max),explore their linear regression relationships,mutual predictability and comparability.Methods Using a quasi-experimental design of cluster sampling and within-group interaction design,20 male cross-country skiers were tested for VO2max using the Bruce protocol(Method 1),90-second incremental load exercise on power bicycle(Method 2),and 1-minute incremental load exercise on treadmill(Method 3),with an interval of one week.The indepen-dent and dependent variable were the three VO2max test methods and the VO2max,respectively.Results Significant differences were found in the average VO2max of the three test results,with the value mea-sured by Method 1 ranking the first,followed by that assessed by Method 3 and Method 2(P<0.05).Moreover,the frequency of individual differences in the results of the three methods showed that the VO2max of Method 1 was about 6 and 3 ml/min·kg higher than that measured by Method 2 and 3.However,at the same treadmill speed,the average blood lactate evaluated using Method 3 was higher than Method 1,and the speed reached aerobic and anaerobic thresholds about one speed unit(1 km/h)lower than Method 1.Meanwhile,linear regression analyses of the test results between Method 1 and 2,as well as Method 1 and 3 showed that both the regression models and coefficients were statis-tically significant(P<0.001),with the R-squared values of 9.25 and 9.05,respectively.Conclusion The Bruce protocol performs best in assessing the maximum value of the athlete's VO2max phase,whose results have linear regression relationships with the other two methods,and can be used for pre-dicting their results.Moreover,athletes of different events and levels can choose different VO2max test methods accordingly.Lastly,the speed and heart rate ranges corresponding to the aerobic and anaero-bic thresholds can serve as an effective and convenient method to control the training intensity.

Auditory neuropathy spectrum disorder (ANSD) represents a variety of sensorineural deafness conditions characterized by abnormal inner hair cells and/or auditory nerve function, but with the preservation of outer hair cell function. ANSD represents up to 15% of individuals with hearing impairments. Through mutation screening, bioinformatic analysis and expression studies, we have previously identified several apoptosis-inducing factor (AIF) mitochondria-associated 1 (AIFM1) variants in ANSD families and in some other sporadic cases. Here, to elucidate the pathogenic mechanisms underlying each AIFM1 variant, we generated AIF-null cells using the clustered regularly interspersed short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system and constructed AIF-wild type (WT) and AIF-mutant (mut) (p.‍T260A, p.‍R422W, and p.‍R451Q) stable transfection cell lines. We then analyzed AIF structure, coenzyme-binding affinity, apoptosis, and other aspects. Results revealed that these variants resulted in impaired dimerization, compromising AIF function. The reduction reaction of AIF variants had proceeded slower than that of AIF-WT. The average levels of AIF dimerization in AIF variant cells were only 34.5%‍‒‍49.7% of that of AIF-WT cells, resulting in caspase-independent apoptosis. The average percentage of apoptotic cells in the variants was 12.3%‍‒‍17.9%, which was significantly higher than that (6.9%‍‒‍7.4%) in controls. However, nicotinamide adenine dinucleotide (NADH) treatment promoted the reduction of apoptosis by rescuing AIF dimerization in AIF variant cells. Our findings show that the impairment of AIF dimerization by AIFM1 variants causes apoptosis contributing to ANSD, and introduce NADH as a potential drug for ANSD treatment. Our results help elucidate the mechanisms of ANSD and may lead to the provision of novel therapies.
Humans ; Apoptosis Inducing Factor/metabolism* ; NAD/metabolism* ; Dimerization ; Apoptosis