Objective To explore the application value of lymphocyte subsets combined with various cytokines in the disease progression of elderly patients with corona virus disease 2019(COVID-19).Methods From December 2022 to January 2023,146 elderly patients with COVID-19 diagnosed in the emergency ward of the Eighth Medical Center of PLA General Hospital were selected and divided into two groups according to the prognosis:127 cases in the COVID-19 survival group,19 cases in the COVID-19 death group.In addition,51 osteoporosis patients in geriatric medicine department were collected as control group.The proportion and absolute count of lymphocyte subsets(including T,B and NK cells),and 12 cytokines in plasma(including IL-1β,IL-2,IL-4,IL-5,IL-6,IL-8,IL-10,IL-12p70,IL-17,TNF-α and IFN-γ)were compared between the control group and COVID-19 group,survival group and death group.The receiver operating characteristic(ROC)curve was used to evaluate its prognostic value in elderly patients with COVID-19 infection.Results Compared with the control group:① The proportion of NK cells in COVID-19 group was decreased,while the proportion of B cells was increased,and the differences were statistically significant(Z=-3.386,-4.140,all P<0.01).There was no significant difference in the proportion of T,CD8+T and CD4+T cells,and the differences were not statistically significant(Z=-1.244,-1.770,-0.951,all P>0.05).② The absolute numbers of T,CD8+T,CD4+T,NK and B cells in COVID-19 group were decreased,and the differences were statistically significant(Z=-9.418～-6.539,all P<0.01).③ The concentrations of IL-2,IL-6,IL-1β,IFN-γ,IL-8,IL-17,IL-12P70 and IL-10 in COVID-19 group were all increased,and the differences were statistically significant(Z=-8.851～-1.986,all P<0.05).There was no significant difference in the concentrations of IL-5,IFN-α,TNF-α and IL-4,and the differences were not statistically significant(Z=-0.460～-0.217,all P>0.05).Compared with the survival group:① There was no significant difference in the proportion of T,CD8+T,CD4+T,NK and B cells in the death group(Z=-1.873～-0.422,all P>0.05).② The absolute numbers of T,CD8+T and CD4+T cells in the death group were all decreased,and the differences were statistically significant(Z=-2.667,-2.287,-2.556,all P<0.05),while there was no significant difference in absolute numbers of NK and B cellsm and the differences were not statistically significant(Z=-1.934,-0.532,all P>0.05).③ The concentrations of IL-6,IFN-γ,IL-8,IL-17 and IL-10 in the death group were all increased,and the differences were not statistically significant(Z=-4.211～-2.655,all P<0.05),and there was no significant difference in the concentrations of IL-5,IFN-α,IL-2,IL-1β,IL-12p70,TNF-α and IL-4 the differences were not statistically significant(Z=-1.329～-0.279,all P>0.05).ROC curve analysis for the prognostic value of lymphocyte subsets combined with cytokines in elderly patients with COVID-19 showed that:the areas of total T cells,B cells and NK cells under ROC curve for predicting the prognosis of COVID-19 infection were 0.94,0.80 and 0.93,respectively.The areas of CD4+T cells and CD8+T cells under ROC curve for predicting the prognosis of COVID-19 infection were 0.93 and 0.90,respectively.The areas of IL-6,IFN-γ,IL-8,IL-17 and IL-10 in cytokines under the ROC curve for predicting the prognosis of COVID-19 infection were 0.91,0.71,0.87,0.74 and 0.90,respectively.However,the area of combined lymphocyte subsets and cytokines under ROC curve for predicting the prognosis of COVID-19 infection reached 0.99.Conclusion The immune status of elderly patients with COVID-19 was generally low.Evaluation of immune status has important clinical guidance significance in disease diagnosis,disease observation and prognosis.

Endo-beta-N-acetylglucosaminidase (ENGase) is widely distributed in various organisms. The first reported ENGase activity was detected in Diplococcus pneumoniae in 1971. The protein (Endo D) was purified and its peptide sequence was determined in 1974. Three ENGases (Endo F1-F3) were discovered in Flavobacterium meningosepticum from 1982 to 1993. After that, the activity was detected from different species of bacteria, yeast, fungal, plant, mice, human, etc. Multiple ENGases were detected in some species, such as Arabidopsis thaliana and Trichoderma atroviride. The first preliminary crystallographic analysis of ENGase was conducted in 1994. But to date, only a few ENGases structures have been obtained, and the structure of human ENGase is still missing. The currently identified ENGases were distributed in the GH18 or GH85 families in Carbohydrate-Active enZyme (CAZy) database. GH18 ENGase only has hydrolytic activity, but GH85 ENGase has both hydrolytic and transglycosylation activity. Although ENGases of the two families have similar (β/α)8-TIM barrel structures, the active sites are slightly different. ENGase is an effective tool for glycan detection andglycan editing. Biochemically, ENGase can specifically hydrolyze β‑1,4 glycosidic bond between the twoN-acetylglucosamines (GlcNAc) on core pentasaccharide presented on glycopeptides and/or glycoproteins. Different ENGases may have different substrate specificity. The hydrolysis products are oligosaccharide chains and a GlcNAc or glycopeptides or glycoproteins with a GlcNAc. Conditionally, it can use the two products to produce a new glycopeptides or glycoprotein. Although ENGase is a common presentation in cell, its biological function remains unclear. Accumulated evidences demonstrated that ENGase is a none essential gene for living and a key regulator for differentiation. No ENGase gene was detected in the genomes of Saccharomyces cerevisiae and three other yeast species. Its expression was extremely low in lung. As glycoproteins are not produced by prokaryotic cells, a role for nutrition and/or microbial-host interaction was predicted for bacterium produced enzymes. In the embryonic lethality phenotype of the Ngly1-deficient mice can be partially rescued by Engase knockout, suggesting down regulation of Engase might be a solution for stress induced adaptation. Potential impacts of ENGase regulation on health and disease were presented. Rabeprazole, a drug used for stomach pain as a proton inhibitor, was identified as an inhibitor for ENGase. ENGases have been applied in vitro to produce antibodies with a designated glycan. The two step reactions were achieved by a pair of ENGase dominated for hydrolysis of substrate glycoprotein and synthesis of new glycoprotein with a free glycan of designed structure, respectively. In addition, ENGase was also been used in cell surface glycan editing. New application scenarios and new detection methods for glycobiological engineering are quickly opened up by the two functions of ENGase, especially in antibody remodeling and antibody drug conjugates. The discovery, distribution, structure property, enzymatic characteristics and recent researches in topical model organisms of ENGase were reviewed in this paper. Possible biological functions and mechanisms of ENGase, including differentiation, digestion of glycoproteins for nutrition and stress responding were hypothesised. In addition, the role of ENGase in glycan editing and synthetic biology was discussed. We hope this paper may provide insights for ENGase research and lay a solid foundation for applied and translational glycomics.

Objective:To evaluate the association between preoperative serum β 2-microglobulin (β 2MG) concentrations and postoperative delirium (POD) in elderly patients. Methods:The study selected patients who underwent knee or hip arthroplasty under spinal-epidural anesthesia on an elective basis at Qingdao Municipal Hospital from May 2021 to November 2022. The patients were divided into a POD group and a non-POD group based on the occurrence of POD. The study was conducted as part of the Perioperative Neurocognitive Impairment and Biomarkers Lifestyle Cohort, which was a nested case-control study. The study collected baseline data from two groups of patients and analyzed the differences between them. Logistic regression was used to identify the risk factors for POD. The stability of the regression model was tested using sensitivity analysis. The mediation model was used to examine whether cerebrospinal fluid (CSF) biomarkers mediated the relationship between β 2MG and POD. The receiver operating characteristic curve was drawn and the area under the curve was calculated to evaluate the accuracy of preoperative β 2MG concentrations and CSF biomarker concentration in predicting POD. Results:There were 57 cases in POD group and 449 cases in non-POD group. The results of logistic regression analysis showed that the increased β 2MG and CSF total tau protein (t-tau) concentrations were risk factors for POD, and the increased CSF β-amyloid 42 concentration was a protective factor for POD after adjustment for multiple confounders such as age, gender, education, Mini-Mental State Examination, history of hypertension and infusion volume ( P<0.05). The results of mediation analysis showed that the serum β 2MG′s effect on POD was partly mediated by t-tau (18.1%). The results of the receiver operating characteristic curve showed that the area under the curve of the β 2MG concentration combined with the CSF biomarker concentration was 0.742. Conclusions:Elevated preoperative serum β 2MG concentration is a risk factor for POD in elderly patients, and the relationship may be partly mediated by CSF t-tau.

Objective:To construct and prepare recombinant virus strains chimeric with human metapneumovirus (HMPV) antigenic epitopes.Methods:Recombinant influenza virus vectors which chimeric with different HMPV antigenic epitopes were rescued by reverse genetics using eight-plasmid system. The recombinant influenza virus strain used the internal genes of A/PR/8/34 (PB1, PB2, PA, NP, NS, M, HA, and NA) as a backbone, with concomitant genetic modifications to insert the B-cell epitopes of HMPV into the HA gene, and the CTL+ Th cell epitopes of HMPV into the NA gene. Preparation of recombinant influenza virus strains using reverse genetics in a " 7+ 1" model. The recombinant virus strains were evaluated by measuring hemagglutinin (HA) titers, half tissue culture infectious dose (TCID 50) and growth curves. Sequencing analysis was conducted to verify whether the rescued viruses carried the chimeric HMPV epitopes. Results:The epitopes of HMPV were inserted into the influenza virus genome and two recombinant influenza virus strains were rescued successfully, named as FLU/HMPV/B and FLU/HMPV/CTL+ Th. HA titers of the recombinant strains were both 2 7, their TCID 50 were 10 5.2/ml and 10 5.0/ml, respectively. After cultured for three passages in chick embryo, these two recombinant strains could proliferate steadily. Whole genome sequencing verified that the FLU/HMPV/B carried the B-cell epitopes of HMPV, the FLU/HMPV/CTL+ Th carried the CTL and Th cell epitopes of HMPV. Growth curve tests also verified that the recombinant strains could proliferate steadily in eggs. Conclusions:Two recombinant influenza virus vector strains carrying the B cell, CTL and Th epitopes of HMPV were rescued successfully. The result of the recombinant virus strains in terms of growth characteristics as well as genetic stability indicate that they meet the requirements for proceeding to the next step of animal experiments. The immunogenicity and immunoprotective effect will be further evaluated by mouse experiments. Ultimately new ideas for the realization of " one vaccine for two uses" or " one vaccine formultiple uses", as well as a new strategy for the development of HMPV vaccine will be proposed.

Objective:To identify the risk factors and to construct a predictive model for early postnatal mortality (with the first 7 days of life) in extremely preterm infants.Methods:This retrospective study involved 244 extremely preterm infants with a gestational age of 22 to 27 weeks and 6 days, born at the Affiliated Hospital of Jining Medical College from January 2017 to December 2022. They were divided into an early survival group ( n=140) and an early mortality group ( n=84), based on survival for ≥7 days after birth. LASSO and logistic regression were used to select risk factors for early mortality. A nomogram predictive model was constructed using the R software program. The goodness-of-fit tests, area under the curve (AUC), calibration curves, and decision curves were used to evaluate its performance and clinical usefulness. Results:LASSO regression and multivariate logistic regression analyses showed that breech delivery ( OR=3.055, 95% CI: 1.125-8.296), intubation in the delivery room ( OR=4.320, 95% CI: 1.328-14.053), diagnosis of grade Ⅲ-Ⅳ neonatal respiratory distress syndrome within 6 h after birth ( OR=11.552, 95% CI: 3.056-43.677), and use of adrenaline in the delivery room ( OR=10.706, 95% CI: 1.454-78.816) were risk factors for early mortality in extremely preterm infants. Conversely, large gestation age ( OR=0.234, 95% CI: 0.125-0.436), antenatal administration of corticosteroids to promote fetal lung maturity ( OR=0.046, 95% CI: 0.014-0.145), and the use of pulmonary surfactant within 6 h after birth ( OR=0.021, 95% CI: 0.004-0.122) were protective factors against mortality. The goodness of fit test of the early death risk nomogram prediction model for extremely preterm infants indicates a good fit ( P=0.702). The AUC of the model was 0.963 (95% CI: 0.943-0.983), with a sensitivity of 0.904 (95% CI: 0.806-0.949), specificity of 0.892 (95% CI: 0.829-0.938), and accuracy of 0.880. Decision curve analysis indicated that a threshold probability>2% would yield a net benefit. Conclusions:Breech delivery, intubation in the delivery room, use of adrenaline in the delivery room, and the diagnosis of grade Ⅲ-Ⅳ neonatal respiratory distress syndrome within 6 h post-birth are independent risk factors for early mortality in extremely preterm infants. Large gestational age, antenatal administration of corticosteroids to promote fetal lung maturity and use of pulmonary surfactant within 6 h after birth are protective factors. The constructed prediction model based on the aforementioned factors can quantitatively, conveniently, and intuitively assess the risk of early mortality in extremely preterm infants.

Objective To explore the current status and potential subtypes of frailty among family caregivers of patients with dementia,and to analyze the related influencing factors of different subtypes.Methods Dementia patients and their family caregivers in 8 community health service centers in Shanghai from June to October 2023 were recruited by convenience sampling.General information questionnaire,Tilburg Frailty Indicator(TFI),Pittsburgh Sleep Quality Index(PSQI),Self-Rating Depression Scale(SDS),Zarit Caregiver Burden Interview(ZBI),and Connor-Davidson Resilience Scale(CD-RISC)were conducted for investigation.Latent profile analysis was used to explore the potential subtypes of frailty among family caregivers of patients with dementia.The influencing factors associated with the potential subtypes were identified by univariate analysis and multivariate Logistic regression analysis.Results A total of 470 family caregivers of patients with dementia were surveyed,and 46.17％of them suffered from frailty.Frailty among family caregivers of patients with dementia can be classified into 3 potential subtypes:comprehensive-low frailty subtype(70.64％),psychosocial-medium frailty subtype(19.57％),and physical-high frailty subtype(9.79％).Family caregivers of patients with dementia who had poor sleep quality and suffered from 2 or more chronic diseases were more likely to be classified into the physical-high frailty subtype(P＜0.05).Family caregivers of patients with dementia who had higher levels of depression,lower mastery levels of caregiving knowledge and skills and spousal caregivers were more likely to be classified into the psychosocial-medium frailty subtype(P＜0.05).Family caregivers of patients with dementia who had higher levels of resilience were more likely to be classified into the comprehensive-low frailty subtype(P＜0.05).Conclusion The incidence of frailty among family caregivers of patients with dementia is at a high level with significant heterogeneity.It is suggested that medical staff should pay attention to the frailty of family caregivers,with a focus on family caregivers in the psychosocial-medium frailty subtype or physical-high frailty subtype,and take timely and targeted interventions according to the characteristics and influencing factors of different subtypes,so as to prevent or delay the occurrence and development of frailty.

Objective:To evaluate the relationship between physical activity (PA) and postoperative delirium (POD) in elderly patients undergoing knee or hip arthroplasty.Methods:The study was conducted as part of the Perioperative Neurocognitive Impairment and Biomarkers Lifestyle Cohort, which was a nested case-control study. Medical records from elderly patients undergoing elective knee or hip arthroplasty under spinal-epidural anesthesia at Qingdao Municipal Hospital from August 2022 to August 2023 were collected. The patients were divided into a POD group ( n=89) and a non-POD group ( n=221) based on the occurrence of POD. Peripheral blood samples were collected before surgery, and the cerebrospinal fluid (CSF) 2 ml was extracted after successful puncture under spinal-epidural anesthesia for determination of the concentrations of amyloid-β 42 (Aβ 42), total tau protein (t-tau), and phosphorylated tau protein (p-tau) by enzyme-linked immunosorbent assay. Logistic regression was used to analyze the influencing factors of POD, and the mediation analysis was conducted to examine the mediating role of CSF biomarker in the relationship between PA and POD. Results:Logistic regression analysis showed that the increased concentration of CSF biomarkers Aβ 42 ( OR=0.997, P=0.006), elevated ratio of Aβ 42/t-tau ( OR=0.642, P=0.003), elevated ratio of Aβ 42/p-tau ratio ( OR=0.872, P=0.001) and PA ( OR=0.374, P=0.001) were protective factors for POD, while the elevated concentrations of t-tau ( OR=1.006, P=0.001) and p-tau ( OR=1.030, P=0.011) were risk factors for POD after adjusting for multi-confounders such as hypertension, diabetes, history of drinking, years of education and Mini-Mental State Examination score. The results of the mediation analysis showed that Aβ 42 (20%), t-tau (16%), Aβ 42/t-tau (23%) and Aβ 42/p-tau (28%) played mediating roles in the relationship between PA and POD. Conclusions:PA is a protective factor for POD in elderly patients undergoing knee or hip arthroplasty and CSF biomarkers may play a mediating role in the relationship between PA and POD.

Objective:To evaluate the relationship between the preoperative sleep quality and postoperative delirium (POD) in elderly patients.Methods:Three hundred and eighty-nine patients of either sex, aged 65-90 yr, with American Society of Anesthesiologists Physical Status classification Ⅰ or Ⅱ, who underwent elective knee/hip arthroplasty under combined spinal-epidural anesthesia in Qingdao Municipal Hospital from May 2022 to October 2023, were selected. The Pittsburgh Sleep Quality Index scale was applied at 1 day before surgery to assess the sleep quality within 1 month before surgery. The cerebrospinal fluid (CSF) specimens 2 ml were withdrawn from the subarachnoid space before anesthesia for determination of the concentrations of biomarkers (β-amyloid), total tau protein [t-tau], and phosphorylated tau protein [p-tau]) by enzyme-linked immunosorbent assay. Patients were divided into POD group and non-POD group based on whether delirium occurred within 7 days after surgery. Logistic regression was used to identify the risk factors for POD, and the sensitivity analysis was used to test the stability of the regression models. The mediation model was used to examine whether CSF biomarkers mediated the link between sleep quality and POD. The accuracy of preoperative sleep quality and CSF biomarker concentrations in predicting the occurrence of POD was evaluated by plotting the receiver operating characteristic curve and calculating the area under the curve.Results:Two hundred and seventy-nine patients were finally included, with 33 patients in POD group and 246 patients in non-POD group. The results of logistic regression analysis showed that shorter sleep duration, use of hypnotics, daytime dysfunction within 1 month before surgery, elevated t-tau concentrations in CSF, and elevated p-tau concentrations in CSF were risk factors for POD after adjusting for multiple confounding factors such as age, sex, body mass index, years of education, Mini-Mental State Examination score, and VAS score. The results of mediating effect showed that the effect of use of hypnotics and daytime dysfunction within 1 month before surgery on POD was partially mediated by p-tau concentrations in CSF (>10%). The area under the curve of combination of daytime dysfunction, sleep duration, use of hypnotics within 1 month before surgery, and CSF biomarker concentrations in predicting POD was 0.979.Conclusions:Shortened sleep duration, use of hypnotic, and daytime dysfunction with 1 month before surgery are risk factors for POD in elderly patients, and the association between use of hypnotics and daytime dysfunction and POD is partially mediated by p-tau in CSF.

Objective:To investigate the correlation between clinical phenotypes and genetic characteristics of children with ring chromosomes (RCs).Methods:Case series study.The clinical data of 11 434 children who received treatment and peripheral blood chromosome karyotype detection in Henan Children′s Hospital from October 2008 to October 2023 due to growth retardation, intellectual impairment or congenital malformation were analyzed retrospectively.A total of 25 children with RCs were selected.Their age at diagnosis, karyotype distribution, clinical manifestations, and genetic detection results were analyzed.Results:RCs were detected in 25 out of 11 434 children, with a detection rate of 0.21%.The genome-wide copy number variation (CNV) analysis was performed on 7 RCs cases, and it found that pathogenic variation existed in all of them.Among the 25 RC cases (11 males and 14 females of social gender), the age at diagnosis ranged from 2 months to 14 years; there were 20 autosomal rings and 5 sex chromosome rings; 13 cases had chimeric karyotypes, and 12 cases had non-chimeric karyotypes.Most of the 25 children showed clinical manifestations of mental or developmental retardation, and some also presented with specific clinical manifestations, such as short stature, congenital malformation, and epilepsy.Conclusions:The pathogenesis of RCs is complex.The clinical manifestations are determined by both RCs syndrome and specific phenotypes caused by the dose effect and exhibit high heterogeneity, so it is easy to miss or misdiagnose.The combined application of cellular and molecular genetic detection technology can facilitate early diagnosis and treatment of RCs, and the correlation analysis of phenotypes and genetic characteristics can provide guidance for genetic counseling.

Objective:To investigate the epidemiological and genetic characteristics of human metapneumovirus (hMPV) in Tianjin during two influenza epidemic seasons from October 2020 to March 2021 and from October 2021 to March 2022, and enrich the whole genome database of hMPV in China.Methods:A total of 1 040 pharyngeal swab samples collected from patients with influenza-like illness (ILI) were analyzed using microfluidic chip fluorescence quantitative PCR. RT-PCR was used to amplify the whole genome in hMPV-positive samples, and the second-generation sequencing was performed for complete genome sequencing. Bioinformatics software including CLC, DNAStar, and MEGA was used for sequence assembly, nucleotide and amino acid homology analysis, and phylogenetic tree mapping.Results:Among the 1 040 samples, 25 were positive for hMPV with a positive rate of 2.40%. The highest positive rate was observed in the age group of 3 to 5 years, reaching 3.71% (16/431). During the influenza epidemic seasons, the detection rate of hMPV peaked in December, reaching 6.67% (12/180). Twelve strains were successfully sequenced, and there were seven of type B2, four of type A2b, and one of type B1. More variations were detected in the G gene, with 111nt-dup sequence repeats observed in the G gene of three A2b strains.Conclusions:The prevalence of hMPV peaks in December during the influenza epidemic seasons in Tianjin, with Type B2 being the predominant type. Except for the G gene with more mutations, other genes remain stable.