Neonatal diabetes mellitus （NDM） is a rare monogenic disorder primarily caused by insufficient insulin secretion resulting from mutations in the KCNJ11 and ABCC8 genes. Sulfonylureas， represented by glibenclamide， have become the standard therapy for this type of NDM by precisely closing the mutated ATP-sensitive potassium channels in pancreatic β cells， thereby restoring insulin secretion. Clinical studies confirm that sulfonylureas enable over 90% of patients to successfully transition from insulin to oral treatment， achieving long-term stable glycemic control and improving neurological outcomes to a certain extent. In terms of safety， severe hypoglycemia induced by sulfonylureas is relatively rare and gastrointestinal reactions are mild； moreover， sulfonylureas show good long-term tolerability， and have no adverse effects on child growth and development. In the future， by further refining the full-chain management pathway of “rapid genetic diagnosis-early intervention-specialized dosage forms-long-term follow-up”， the clinical application of sulfonylureas is expected to provide NDM patients with an optimized treatment regimen and maximize their health benefits.

ObjectiveTo investigate the migration and distribution characteristics of chemical constituents in blood and hippocampal tissues before and after vinegar processing of Citri Reticulatae Pericarpium Viride（CRPV）， and to explore the potential material basis and mechanisms underlying their regulatory effects on emotional disorders by comparing the effects of raw and vinegar-processed products of CRPV. MethodsUltra-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry（UPLC-Q-TOF-MS/MS） was employed to characterize and identify the chemical constituents of raw and vinegar-processed products of CRPV extracts， as well as their migrating components in blood and hippocampal tissues after oral administration. Reference standards， databases， and relevant literature were utilized for compound annotation， with data processing performed using PeakView 1.2 software. Seventy male C57BL/6 mice were randomly divided into seven groups， including the blank group， model group， diazepam group（2.5 mg·kg^-1）， raw CRPV low/high dose groups（0.6， 1.2 g·kg^-1）， and vinegar-processed CRPV low/high dose groups（0.6， 1.2 g·kg^-1）， with 10 mice per group. Except for the blank group， all other groups underwent chronic restraint stress（2 h·d^-1） for 20 d. Each drug-treated group received oral administration at the predetermined dose starting 10 d after modeling， with a total treatment duration of 10 d. Following model-based drug administration， mice underwent open-field， forced swimming， and elevated plus maze tests. After anesthesia with isoflurane， whole brains were collected from each group of mice， and hippocampi were dissected. Reactive oxygen species（ROS） level in hippocampal tissues was quantified by enzyme-linked immunosorbent assay（ELISA）. Hematoxylin-eosin（HE） staining was used to observe hippocampal tissue morphology. Immunofluorescence was performed to detect neuronal nuclei（NeuN） and peroxisome proliferator-activated receptor alpha（PPARα） expressions in hippocampal tissue. Then， pharmacodynamic evaluations were conducted to assess the effects of raw and vinegar-processed CRPV on mood disorders， exploring the potential mechanisms. ResultsVinegar processing caused significant changes in the chemical composition of CRPV， with 18 components showing increased relative content and 35 components showing decreased relative content. The primary changes occurred in flavonoid compounds， including 20 flavonoids， 20 flavonoid glycosides， 3 triterpenes， 3 phenolic acids， 1 alkaloid， and 6 other compounds. Twenty-one components were detected in blood（15 methoxyflavones， 4 flavonoid glycosides， and 2 phenolic acids）， with 17 shared between raw and vinegar-processed CRPV. Seven components reached hippocampal tissues（all common to both forms）. In regulating emotional disorders， Vinegar-processed CRPV exhibited superior antidepressant-like effects compared to raw products. HE staining revealed that both treatments improved hippocampal neuronal morphology， particularly in the damaged CA1 and CA3 regions. Immunofluorescence and ELISA analyses demonstrated that both raw and vinegar-processed CRPV significantly modulated NeuN and PPARα expressions in hippocampal tissue while alleviating oxidative stress induced by excessive ROS（P<0.05）. ConclusionThe chemical composition of CRPV undergoes changes after vinegar processing， but the migrating components in blood and hippocampus are primarily methoxyflavonoids. These components may serve as the potential material basis for activating the PPARα pathway， thereby negatively regulating ROS generation in the hippocampus， reducing oxidative stress， and promoting the development of NeuN-positive neurons. These findings provide experimental evidence for enhancing quality standards， pharmacodynamic material research， and active drug development of raw and vinegar-processed CRPV.

Objective To explore the feasibility and reference value of allogeneic lung transplantation and postoperative monitoring in miniature pigs for lung transplantation research. Methods Two miniature pigs (R1 and R2) underwent left lung allogeneic transplantation. Complement-dependent cytotoxicity tests and blood cross-matching were performed before surgery. The main operative times and partial pressure of arterial oxygen (PaO₂) after opening the pulmonary artery were recorded during surgery. Postoperatively, routine blood tests, biochemical blood indicators and inflammatory factors were detected, and pathological examinations of multiple organs were conducted. Results The complement-dependent cytotoxicity test showed that the survival rate of lymphocytes between donors and recipients was 42.5%-47.3%, and no agglutination reaction occurred in the cross-matching. The first warm ischemia times of D1 and D2 were 17 min and 10 min, respectively, and the cold ischemia times were 246 min and 216 min, respectively. Ultimately, R1 and R2 survived for 1.5 h and 104 h, respectively. Postoperatively, in R1, albumin (ALB) and globulin (GLB) decreased, and alanine aminotransferase increased; in R2, ALB, GLB and aspartate aminotransferase all increased. Urea nitrogen and serum creatinine increased in both recipients. Pathological results showed that in R1, the transplanted lung had partial consolidation with inflammatory cell infiltration, and multiple organs were congested and damaged. In R2, the transplanted lung had severe necrosis with fibrosis, and multiple organs had mild to moderate damage. The expression levels of interleukin-1β and interleukin-6 increased in the transplanted lungs. Conclusions The allogeneic lung transplantation model in miniature pigs may systematically evaluate immunological compatibility, intraoperative function and postoperative organ damage. The data obtained may provide technical references for subsequent lung transplantation research.

Objective:To evaluate the levels and source of cadmium exposure by dietary pathway among middle-aged and elderly people ≥40 in cadmium-contaminated areas of China.Methods:A total of 7 193 people aged 40-89 years from four typical cadmium-contaminated areas in China were selected as the study subjects. Food Frequency Questionnaire (FFQ), Total Diet Study (TDS) and a 3-day-24-hour dietary recall survey were conducted. Dietary cadmium intake and food sources through dietary pathways were assessed based on cadmium content in foods, consumption amounts and intake frequencies.Results:The mean age of the participants was 63.39±12.21 years, with 50.05% being males. The average monthly dietary cadmium intake was 7.39 μg/(kg·BW). Staple foods and vegetables were the primary sources of dietary cadmium intake, accounting for 57.51% and 32.48%, respectively. The monthly dietary cadmium intake in all surveyed regions did not exceed the Provisional Tolerable Monthly Intake (PTMI) recommended by the Joint FAO/WHO Expert Committee on Food Additives (JECFA).Conclusion:The monthly dietary cadmium intake among middle-aged and elderly people in cadmium-contaminated areas of China is relatively low, with the risk remaining at an acceptable level. Staple foods and vegetables are the most significant contributors to dietary cadmium intake.

Turning to critical illness is a common stage of various diseases and injuries before death. Patients usually have complex health conditions, while the treatment process involves a wide range of content, along with high requirements for doctor′s professionalism and multi-specialty teamwork, as well as a great demand for time-sensitive treatments. However, this is not matched with critical care professionals and the current state of medical care in China. Telemedicine, which shortens the distance of medical professionals and the gap of disease diagnosis and treatments in various regions through electronic information, can effectively solve the current problem. Therefore, there is an urgent need to develop a standardized, high-quality visualization telemedicine round system .Therefore, experts have been organized to search domestic and foreign literature on telemedicine round for critically ill patients and to form this consensus based on clinical experiences so as to further improve the level of critical care treatments in regions.

To investigate the distribution and development of common allergens in children in Hangzhou area, and to provide an epidemiological basis for the prevention, diagnosis, and treatment of allergic diseases in this area. This study is a retrospective study, selecting 3 524 children who underwent allergen screening at DiAn Medical Laboratory Center (Hangzhou) Co., Ltd. from January 2022 to January 2024 as the research subjects, including 2 012 males and 1 512 females. Among them, there were 1 098 infants (0-1 year olds), 1 673 toddlers (1-3 year olds), and 753 children (3-14 year olds). Immunoblotting was used to detect 21 allergen-specific immunoglobulin E (sIgE) antibodies. The positive rates of various allergens were calculated according to gender, age, and other factors, and the epidemiological characteristics and trends were analyzed. The results showed that the overall positive rate of sIgE was 60.33% (2 126/3 524). The main inhaled allergens were Dermatophagoides pteronyssinus/Dermatophagoides farinae (992, 28.15%), dog dander (295, 8.37%) and cat dander (181, 5.13%). The main food allergens were milk (696, 19.75%), egg white (541, 15.36%) and shrimp (205, 5.83%). Boys had significantly higher sIgE positivity rates than girls for Dermatophagoides pteronyssinus/Dermatophagoides farinae(642, 31.90%, χ2=10.10, P=0.001), house dust(61, 3.02%, χ2=5.12, P=0.024), cat dander(124, 6.16%, χ2=4.06, P=0.044), mold combinations(103, 5.14%, χ2=7.51, P=0.006), and tree pollen combinations(42, 2.07%, χ2=5.44, P=0.020) (all P<0.05); With age, there was a significant increase in positivity rates for house dust mite/dust mite, house dust, mold combinations, milk, and beef (all P<0.05), and a significant decrease in positivity rates for cockroach consumption, egg whites, shrimp, crab, cod, lobster/scallop, and soybeans (all P<0.05), the positive rates of only ingestive allergens decreased significantly ( P<0.001), and the positive rates of only inhalant and mixed allergens increased significantly ( P<0.001). In conclusion,in the Hangzhou area, dust mites are the most common inhalant allergens among children with allergic diseases, while milk is the most common food allergen. Boys are more sensitive to inhalant allergens, and as children grow older, the positive rates for different allergens undergo significant changes. Dynamic monitoring of changes in specific IgE antibodies to various allergens can assist in the diagnosis, prevention, and treatment of allergic diseases.

Objective:To establish tumor specific protein (SP70) targeted tumor cell enrichment technology and to assess applicational value of next-generation sequencing (NGS) analysis for enriched circulating tumor cell (CTC) in precision medicines of non-small cell lung cancer (NSCLC).Methods:The monoclonal antibody NJ001 was covalently coupled to the surface of magnetic beads to build targeted magnetic bead enrichment technology based on SP70. The limit of detection, coincidence rate, interference experiment, recovery test and clinical performance were evaluated. From March 2016 to August 2017, NGS analysis with or without pre-treatment of targeted enrichment for serous fluids of 43 NSCLC in the First Affiliated Hospital with Nanjing Medical University were compared (Kappa or Fisher exact test).Results:The CTC enrichment technology based on SP70 targeted immunomagnetic beads can specifically enrich tumor cells. The limit of detection was 10 4 SPC-A1 cells/L, and the coincidence rate, sensitivity and specificity were 100% (3/3). The endogenous interfering substances such as red blood cells, hemoglobin, white blood cells, epithelial cells and triglycerides had no interfering effects, as well as the exogenous interfering substances such as EDTA-K2, cefoxitin, carboplatin and paclitaxel. The recovery rate was 56.0% (56 000/100 000). A total of 30 gene mutations including 65 loci were found in 43 NSCLC under SP70 targeted enrichment, with a higher detection rate compared with unenrichment method [95.0% (19/20) vs 65.0% (13/20), χ 2=5.625, P=0.044]. Conclusion:In this study, SP70-targeted enriched CTC liquid biopsy method was established, with higher sensitivity and specificity of NGS detection than unenrichment method.

Combined with elastic network model(ENM),the perturbation response scanning(PRS)has emerged as a robust technique for pinpointing allosteric interactions within proteins.Here,we proposed the PRS analysis of drug-target networks(DTNs),which could provide a promising avenue in network medicine.We demonstrated the utility of the method by introducing a deep learning and network perturbation-based framework,for drug repurposing of multiple sclerosis(MS).First,the MS comorbidity network was constructed by performing a random walk with restart algorithm based on shared genes between MS and other diseases as seed nodes.Then,based on topological analysis and functional annotation,the neurotransmission module was identified as the"therapeutic module"of MS.Further,perturbation scores of drugs on the module were calculated by constructing the DTN and introducing the PRS analysis,giving a list of repurposable drugs for MS.Mechanism of action analysis both at pathway and structural levels screened dihydroergocristine as a candidate drug of MS by targeting a serotonin receptor of se-rotonin 2B receptor(HTR2B).Finally,we established a cuprizone-induced chronic mouse model to evaluate the alteration of HTR2B in mouse brain regions and observed that HTR2B was significantly reduced in the cuprizone-induced mouse cortex.These findings proved that the network perturbation modeling is a promising avenue for drug repurposing of MS.As a useful systematic method,our approach can also be used to discover the new molecular mechanism and provide effective candidate drugs for other complex diseases.

OBJECTIVE: To explore the clinical and genetic characteristics of a Chinese pedigree affected with Hypertrophic cardiomyopathy (HCM) due to a truncating variant of ALPK3 gene. METHODS: A 44-year-old male admitted to Taizhou Hospital of Zhejiang Province on December 29, 2018 was selected as the study subject. Whole-exome sequencing (WES) was carried out, and candidate variant was interpreted by following the guidelines from the American College of Medical Genetics and Genomics (ACMG). For ALPK3 was considered an autosomal recessive gene, the WES results was considered insufficient to explain his phenotype. In April 2023, the proband's WES data were re-analyzed using updated annotation pipelines, and peripheral blood samples were collected from his first-degree relatives (mother and brother) for Sanger sequencing validation. Conservation analysis and protein structural modeling were performed to assess the impact of the variant. Clinical evaluation and genetic counseling were provided to the proband's family members. Relevant literature on ALPK3tv-induced HCM patients were searched in Wanfang Data Knowledge Service Platform, CNKI, and PubMed database using "ALPK3" and "hypertrophic cardiomyopathy" as keywords. Clinical characteristics of HCM patients with heterozygous ALPK3tv variants were summarized and compared with the clinical characteristics of HCM patients with positive sarcomere-associated gene variants (SARC+). This study was approved by the Medical Ethics Committee of Taizhou Hospital of Zhejiang Province Affiliated to Wenzhou Medical University (Ethics No.: K20230314). RESULTS: The proband was a 44-year-old male who was transferred to our institution on December 29, 2018 due to "chest tightness and pain for 6 months, exacerbated for 2 days". Emergency coronary angiography was performed, which led to a preliminary diagnosis of "acute coronary syndrome", and the patient was admitted to the Cardiology Department for treatment. Based on electrocardiogram and echocardiogram findings, the diagnosis was revised as HCM. The patient's condition has stabilized post-coronary angiography, and he was discharged with improved condition. On January 2019, WES was conducted to determine the etiology of the proband's HCM. WES results identified a novel heterozygous c.2156dupC (p.Pro720ThrfsTer53) truncating variant in the ALPK3 gene. At that time, the inheritance pattern could not explain the phenotype. In 2022, a literature indicated that heterozygous ALPK3tv could lead to autosomal dominant HCM. Consequently, in April 2023, the proband's whole-exome data were re-annotated, revealing changes in the transcript and protein versions, with the updated site annotated as ALPK3 (NM_020778.5): c.1550dupC (p.Pro518ThrfsTer53). Sanger sequencing confirmed that the proband's mother and brother also carried this variant. The mother exhibited obstructive HCM, while the brother showed no related phenotype. Bioinformatics analysis demonstrated conservation of this site across multiple species, and the variant has resulted in the loss of a protein domain. Based on ACMG guidelines, the variant was classified as likely pathogenic. Literature review and Bayesian calculation further elevated the pathogenicity rating, indicating that this variant was the cause of HCM in the patient. Literature study revealed distinctions between HCM caused by this variant type and SARC+ HCM. The age of onset among heterozygous ALPK3tv patients was delayed by approximately 10 years compared to SARC+ patients. Both forms of HCM exhibited a male predominance, which was particularly marked in individuals with ALPK3tv. Electrocardiographic left ventricular hypertrophy was more prevalent in heterozygous ALPK3tv patients than in SARC+ patients. The incidence of apical or concentric hypertrophy patterns was higher in heterozygous ALPK3tv patients compared to asymmetric septal hypertrophy, which predominated in SARC+ patients. ALPK3tv patients exhibited lower penetrance and later onset compared to SARC+ patients. A positive correlation between left ventricular wall thickness and age was noted in female patients only. CONCLUSION In this pedigree, the proband has presented with HCM, characterized by echocardiographic evidence of apical left ventricular hypertrophy without significant outflow tract obstruction or extracardiac phenotypes. Although his mother and brother had carried the same heterozygous ALPK3 (NM_020778.5) c.1550dupC (p.Pro518ThrfsTer53), the mother exhibited severe obstructive HCM, while the brother was asymptomatic, suggesting incomplete or age-dependent penetrance within the family. This study has enriched the evidence for the pathogenicity of ALPK3tv among Chinese HCM pedigrees and underscored the importance of periodic literature reviews and genetic re-analysis for unresolved genetic testing results.
Humans ; Male ; Pedigree ; Adult ; Cardiomyopathy, Hypertrophic/genetics* ; Heterozygote ; Asian People/genetics* ; Exome Sequencing ; Mutation ; China ; Female ; East Asian People

Objective To explore the construction of α-1,3-galactosyltransferase (GGTA1) gene-knockout (GTKO) Diannan miniature pigs and the kidney xenotransplantation from pigs to rhesus macaques, and to assess the effectiveness of GTKO pigs. Methods The GTKO Diannan miniature pigs were constructed using the CRISPR/Cas9 gene-editing system and somatic cell cloning technology. The phenotype of GTKO pigs was verified through polymerase chain reaction, Sanger sequencing and immunofluorescence staining. Flow cytometry was used to detect antigen-antibody (IgM) binding and complement-dependent cytotoxicity. Kidney xenotransplantation was performed from GTKO pigs to rhesus macaques. The humoral immunity, cellular immunity, coagulation and physiological indicators of the recipient monkeys were monitored. The function and pathological changes of the transplanted kidneys were analyzed using ultrasonography, hematoxylin-eosin staining, immunohistochemical staining and immunofluorescence staining. Results Single-guide RNA (sgRNA) targeting exon 4 of the GGTA1 gene in Diannan miniature pigs was designed. The pGL3-GGTA1-sgRNA1-GFP vector was transfected into fetal fibroblasts of Diannan miniature pigs. After puromycin selection, two cell clones, C59# and C89#, were identified as GGTA1 gene-knockout clones. These clones were expanded to form cell lines, which were used as donor cells for somatic cell nuclear transfer. The reconstructed embryos were transferred into the oviducts of trihybrid surrogate sows, resulting in 13 fetal pigs. Among them, fetuses F04 and F11 exhibited biallelic mutations in the GGTA1 gene, and F04 had a normal karyotype. Using this GTKO fetal pig for recloning and transferring the reconstructed embryos into the oviducts of trihybrid surrogate sows, seven surviving piglets were obtained, all of which did not express α-Gal epitope. The binding of IgM from the serum of rhesus monkey 20# to GTKO pig PBMC was reduced, and the survival rate of GTKO pig PBMC in the complement-dependent cytotoxicity assay was higher than that of wild-type pig. GTKO pig kidneys were harvested and perfused until completely white. After the left kidney of the recipient monkey was removed, the pig kidney was heterotopically transplanted. Following vascular anastomosis and blood flow restoration, the pig kidney rapidly turned pink without hyperacute rejection (HAR). Urine appeared in the ureter 6 minutes later, indicating successful kidney transplantation. The right kidney of the recipient was then removed. Seven days after transplantation, the transplanted kidney had good blood flow, the recipient monkey's serum creatinine level was stable, and serum potassium and cystatin C levels were effectively controlled, although they increased 10 days after transplantation. Seven days after transplantation, the levels of white blood cells, lymphocytes, monocytes and eosinophils in the recipient monkey increased, while platelet count and fibrinogen levels decreased. The activated partial thromboplastin time, thrombin time and prothrombin time remained relatively stable but later showed an upward trend. The recipient monkey survived for 10 days. At autopsy, the transplanted kidney was found to be congested, swollen and necrotic, with a small amount of IgG deposition in the renal tissue, and a large amount of IgM, complement C3c and C4d deposition, as well as CD68⁺ macrophage infiltration. Conclusions The kidneys of GTKO Diannan miniature pigs may maintain normal renal function for a certain period in rhesus macaques and effectively overcome HAR, confirming the effectiveness of GTKO pigs for xenotransplantation.