Aim To investigate the molecular mecha-nisms of the Wenyang Huazhuo Tongluo formula in in-hibiting endothelial-to-mesenchymal transition(En-doMT)of pulmonary microvascular endothelial cells and improving pulmonary microvascular injury in sys-temic sclerosis(SSc).Methods Pulmonary micro-vascular endothelial cells were cultured with serum from SSc patients to establish SSc pulmonary microvas-cular endothelial cells.A hypoxia model was estab-lished in SSc pulmonary microvascular endothelial cells using liquid paraffin sealing,and the cells were treated with the Wenyang Huazhuo Tongluo formula or HIF-1a inhibitor KC7F2.Western blot was used to detect the protein expression levels of VE-cadherin,CD31,vimen-tin,HIF-1α,Foxm1,smad3,Tie-1,and vWF.ELISA was used to measure the concentrations of E-selectin and ICAM-1 in cell culture medium.The luciferase re-porter gene system was used to detect the promoter ac-tivity of the Foxm1 gene.Results Compared to the control group,the levels of VE-cadherin,CD31,HIF-1α,Foxm1,smad3,Tie-1,and vWF significantly de-creased under hypoxic condition,while the levels of vi-mentin,E-selectin,and ICAM-1 significantly in-creased.In addition,the cell morphology exhibited a distinct"spindle-like"myoblast morphology.Treat-ment with the Wenyang Huazhuo Tongluo formula or KC7F2 reversed these changes in protein expression levels and cell morphology induced by hypoxia.Con-clusion The Wenyang Huazhuo Tongluo formula im-proves pulmonary microvascular injury in SSc by inhib-iting the HIF-1a/Foxm1/smad3 pathway-mediated En-doMT of pulmonary microvascular endothelial cells.

Aim To investigate the effect of paeonol(PAE)on cardiac hypertrophy induced by Angio-tensinogen Ⅱ(Ang Ⅱ)in rats and its mechanism.Methods Forty Sprague-Dawley rats were divided in-to five groups:control group,Ang Ⅱ model group,low concentration paeonol group,middle concentration pae-onol group,high concentration paeonol group.PAE was administered intragastrically(25,50 and 100 mg·kg-1·d-1)for 28 days.The hypertrophic model was established by adding 1 μmol·L-1 Ang Ⅱ to H9c2 cells for 48 hours.Results In Ang Ⅱ-induced rats,PAE improved echocardiography parameters,the cardi-ac hypertrophy index,the mRNA and protein expres-sion levels of ANP and BNP were decreased,and the cardiac fibrosis was alleviated.In vitro,PAE reduced cardiomyocyte hypertrophy and decreased the mRNA and protein expression levels of ANP and BNP in H9c2 cells induced by Ang Ⅱ,at the same time,the expres-sion of endoplasmic reticulum stress marker proteins p-PERK,GRP78,ATF4 and CHOP were decreased and reversed after treatment with the endoplasmic reticulum stress agonist tunicamycin(TN).Conclusion This study suggests that PAE can improve cardiac hypertro-phy by inhibiting endoplasmic reticulum stress,which may be a new drug to delay the development of cardiac hypertrophy.

Objective:To establish a new type of small intestinal organoids with injury-related regenerative capacity, and to simulate the process of intestinal injury, regeneration, and repair in vitro. Methods:The crypt structures of ileal mucosa from 6 to 8 weeks old, 18 to 24 g specific pathogen-free C57BL/6 mice were isolated. The ENR, ENR+ tumor necrosis factor-α(TNF-α) and 8C culture systems were designed to establish small intestinal organoids under conditions of intestinal homeostasis, inflammatory injury and injury-related regeneration, and the morphology of intestinal organoids were observed. The cell types and spatial arrangements of intestinal organoids, and the expression of genes clusterin( Clu), annexin A1( Anxa1), stem cell antigen-1( Sca1) and regenerating islet-derived protein 3-beta( Reg3 b) at protein levels were detected by immunofluorescence staining. The expression of genes Clu, Anxa1, Sca1 and Reg3 b at mRNA levels were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Independent sample- t test was used for statistical analysis. Results:In ENR and 8C culture system, both intestinal organoids contained intestinal stem cells, goblet cells, Paneth cells and intestinal endocrine cells, and the spatial arrangement of cells was similar to the intestinal epithelium. In the 8C culture system, the amplification capacity of the new small intestinal organoids was significantly enhanced, the growth rate was faster, and the structure was larger and more complex than those of small intestinal organoids in ENR and ENR+ TNF-α culture systems. The results of qRT-PCR showed that, the relative mRNA expression levels of novel small intestinal organoid regeneration genes Clu, Anxa1, and Sca1 in the 8C culture system were higher than those in the ENR and ENR+ TNF-α culture systems (0.68±0.31 vs.0.20±0.07 and 0.36±0.19, 0.48±0.13 vs. 0.07±0.02 and 0.18±0.11, 0.56±0.20 vs. 0.02±0.01 and 0.08±0.04), and the differences were statistically significant ( t=4.82 and 2.77, 8.62 and 4.89, and 8.58 and 7.50; all P<0.05). The results of immunofluorescence staining indicated that, the expression levels of novel small intestinal organoid regeneration genes Clu, Anxa1, Sca1 and Reg3 b at protein level in the 8C culture system were higher than those in the ENR and ENR+ TNF-α culture systems (31.62±1.69 vs. 9.73±2.39 and 15.11±2.16, 42.65±1.85 vs. 19.70±1.18 and 24.97±2.82, 63.80±2.73 vs. 37.10±1.59 and 43.27±2.53, 53.26±1.84 vs. 27.75±3.78 and 33.16±3.50), and the differences were statistically significant( t=12.95 and 10.41, 18.13 and 9.09, 14.63 and 9.56, and 10.51 and 8.80; all P<0.001). Conclusion:The small intestinal organoids established in the novel culture system have the characteristics of injury-related regeneration, and provide a novel in vitro model for studying the regeneration of epithelial tissues and organs.

Glioblastoma(GBM)is a lethal cancer with limited therapeutic options.Dendritic cell(DC)-based cancer vaccines provide a promising approach for GBM treatment.Clinical studies suggest that other immu-notherapeutic agents may be combined with DC vaccines to further enhance antitumor activity.Here,we report a GBM case with combination immunotherapy consisting of DC vaccines,anti-programmed death-1(anti-PD-1)and poly I:C as well as the chemotherapeutic agent cyclophosphamide that was integrated with standard chemoradiation therapy,and the patient remained disease-free for 69 months.The patient received DC vaccines loaded with multiple forms of tumor antigens,including mRNA-tumor associated antigens(TAA),mRNA-neoantigens,and hypochlorous acid(HOCl)-oxidized tumor lysates.Furthermore,mRNA-TAAAs were modified with a novel TriVac technology that fuses TAAs with a destabilization domain and inserts TAAs into full-length lysosomal associated membrane protein-1 to enhance major histo-compatibility complex(MHC)class Ⅰ and Ⅱ antigen presentation.The treatment consisted of 42 DC cancer vaccine infusions,26 anti-PD-1 antibody nivolumab administrations and 126 poly I:C injections for DC infusions.The patient also received 28 doses of cyclophosphamide for depletion of regulatory T cells.No immunotherapy-related adverse events were observed during the treatment.Robust antitumor CD4+and CD8+T-cell responses were detected.The patient remains free of disease progression.This is the first case report on the combination of the above three agents to treat glioblastoma patients.Our results suggest that integrated combination immunotherapy is safe and feasible for long-term treatment in this patient.A large-scale trial to validate these findings is warranted.

Objective:To screen the candidate genes in a patient with Kabuki syndrome (KS), providing basis for genetic counseling, prenatal screening, prenatal diagnosis and facilitating early treatment.Methods:This study included a 16-year-old female KS patient born of non-consanguineous Chinese parents who presented to Department of Orthognathic & Cleft Lip and Palate Plastic Surgery, School and Hospital of Stomatology, Wuhan University. Genomic DNA was extracted from the peripheral blood of the subjects and analyzed by whole-exome sequencing (WES). Sanger sequencing was performed to validate the mutation in the candidate gene. The conformational and physicochemical changes of the mutant were analyzed by Alphafold2, Antheprot and DOG.2.0.1, respectively. Distribution of KMT2D mutations in patients with KS was analyzed based on the Human Gene Mutation DatabaseResults:The proband manifested a typical KS facial gestalt, congenital cleft palate, fifth finger deformity, hypodontia, renal hypoplasia and hydronephrosis. Two de novo mutations c.[1166A>C; 1167dupC] (NM_003482) in cis on the same allele in the KMT2D gene were identified by WES and confirmed by allele-specific PCR. Bioinformatics analysis showed that three more α-helixes were added, and a (β-) turn and a (β-) sheet were reduced in KMT2D p. Y389S, p.V390Rfs*26 compared with the wild type. Meanwhile, the interceptive mutant-KMT2D protein p.V390Rfs*26 lost all four domains (FYRN domain, FYRC domain, SET domain, and PostSET domain), which may cause functional disabilities. Conclusions:Our study is the first to identify two novel and de novo KMT2D mutations in cis on the same allele in a KS patient and extends the KMT2D mutation spectrum of KS, providing evidence for genetic susceptibility counseling, prenatal screening and diagnosis, and early treatment of KS.

OBJECTIVES: To study the clinical features and prognosis of children and their family members with family clusters of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant infection under the admission mode of parent-child ward. METHODS: A retrospective analysis was performed on the medical data of 190 children and 190 family members with SARS-CoV-2 Omicron variant infection who were admitted to Shanghai Sixth People's Hospital, the designated hospital for coronavirus disease 2019 (COVID-19), April 8 to May 10, 2022. RESULTS: Both the child and adult groups were mainly mild COVID-19, and the proportion of mild cases in the child group was higher than that in the adult group (P<0.05). Respiratory symptoms were the main clinical manifestations in both groups. Compared with the adult group, the child group had higher incidence rates of fever, abdominal pain, diarrhea, and wheezing (P<0.05) and lower incidence rates of nasal obstruction, runny nose, cough, dry throat, throat itching, and throat pain (P<0.05). Compared with the child group, the adult group had higher rates of use of Chinese patent drugs, traditional Chinese medicine decoction, recombinant interferon spray, cough-relieving and phlegm-eliminating drugs, and nirmatrelvir/ritonavir tablets (P<0.05). Compared with the adult group, the child group had a lower vaccination rate of SARS-CoV-2 vaccine (30.5% vs 71.1%, P<0.001) and a shorter duration of positive SARS-CoV-2 nucleic acid (P<0.05). The patients with mild COVID-19 had a shorter duration of positive SARS-CoV-2 nucleic acid than those with common COVID-19 in both groups (P<0.05). The patients with underlying diseases had a longer duration of positive SARS-CoV-2 nucleic acid than those without such diseases in both groups (P<0.05). CONCLUSIONS Both children and adults with family clusters of SARS-CoV-2 Omicron variant infection manifest mainly mild COVID-19. Despite lower vaccination rate of SARS-CoV-2 vaccine in children, they have rapid disease recovery, with a shorter duration of positive SARS-CoV-2 nucleic acid than adults, under the admission mode of parent-child ward.
Adult ; Humans ; COVID-19/epidemiology* ; SARS-CoV-2 ; Cough ; Retrospective Studies ; COVID-19 Vaccines ; China/epidemiology* ; Family ; Nucleic Acids

Objective:To investigate the collateral circulation compensation model in patients with favorable prognosis of basilar artery occlusion/severe stenosis treated with drugs or endovascular therapy.Methods:Clinical data of patients with basilar artery occlusion/severe stenosis and good clinical outcome were retrospectively collected in the Department of Neurology, Sixth Medical Center of PLA General Hospital from January 2019 to January 2020. They were divided into intensive drug therapy group and combined endovascular therapy group. The number and ways of collateral compensation pathway described by digital substraction angiography (DSA) were analyzed, and the characteristics of the collateral compensation model were summarized. SPSS22.0 software was used for statistical analysis, and the constituent ratio (%) was used for statistical description of the enumeration data.Results:A total of 32 eligible patients were included, including 27 males and 5 females, with an average age 45-76 (59±10) years. The compensation model included posterior communicating artery-posterior cerebral artery (13 cases, 40.6%), posterior communicating artery-posterior cerebral artery-basilar artery (10 cases, 31.2%), cerebellar artery-anastomotic branches of superior cerebellar artery (8 cases, 25.0%), anterior choroid artery-anastomotic branches of posterior choroid artery (2 cases, 6.2%), collateral circulation not established (11 cases, 34.4%).In drug treatment group, collateral compensation was found in the majority (14/15), with mainly posterior communicating artery (10/14).Most patients in combined treatment group did not develop collateral compensation (10/17), anastomotic branches of PICA-SCA were the main routes (6/7).Conclusion:In patients with basilar artery occlusion/severe stenosis, favorable clinical outcome can be achieved in both groups of patients treated with intensive drug therapy or endovascular therapy.

OBJECTIVE: To evaluate the incidence and clinical characteristics of metabolic syndrome (MS) within one year after hematopoietic stem cell transplantation (HSCT) in order to screen the risk factors for HSCT-MS, provide early intervention and improve the long-term quality of survival of patients. METHODS: The clinical follow-up data of 64 HSCT patients (survival time > 1 year) who received HSCT in our center from January 2007 to August 2018 were collected. Among them, 50 cases were allogeneic hematopoietic stem cell transplantation (allo-HSCT) and 14 cases were autologous hematopoietic stem cell transplantation (auto-HSCT). The changes of MS-related indexes and clinical characteristics before and 1, 3, 6 and 12 months after HSCT were analyzed retrospectively. RESULTS: In allo-HSCT group, 14 cases were diagnosed as MS before operation, including high-density lipoprotein cholesterol (hypo-HDL-C)> hyper triglycerides（hyper-TG）> hyper fasting glucose（hyper-FBG）> abdominal obesity (AO) > hypertension. The preoperative diagnosis of MS in the auto-HSCT group was 5 cases, in the order of hyper-FBG> hyper-TG> AO> hypo-HDL-C> hypertension. Incidence of MS at 1, 3, 6 and 12 months after transplantation: 19, 26, 24 and 20 cases in the allo-HSCT group, respectively; auto-HSCT group were 7, 7, 6 and 6 cases, respectively. Hyper-TG and hypo-HDL-C were prominent in both groups. CONCLUSION The incidence of HSCT-MS is significantly higher within 1 year after HSCT. Regardless of allo-HSCT and auto-HSCT, the prevention and control of HSCT-MS is emphasized as an important guarantee to improve the long-term survival quality of HSCT patients.
Hematopoietic Stem Cell Transplantation ; Hematopoietic Stem Cells ; Humans ; Metabolic Syndrome ; Retrospective Studies ; Transplantation, Homologous

Objective: To evaluate the long-term prognosis of coronary artery disease (CAD) patients undergoing percutaneous coronary intervention (PCI) by risk stratification with American College of Cardiology (ACC)/American Heart Association (AHA) classification of coronary lesions. Methods: Data used in this study derived from the I-LOVE-IT 2 trial. I-LOVE-IT 2 trial was a prospective, multicenter, randomized, assessor-blinded, noninferiority study. A total of 1 255 patients in I-LOVE-IT 2 trial with only one lesion and underwent biodegradable polymer drug-eluting stent implantation were included and grouped according to ACC/AHA classification of coronary lesions, namely type A/B1 lesion group (n=184), type B2 lesion group (n=457) and type C lesion group (n=614). The primary endpoint was 48-month patient-oriented composite endpoint (PoCE), a composite of all-cause mortality, all myocardial infarction, stroke, and/or any revascularization. The secondary endpoints were target lesion failure (TLF), components of PoCE, major bleeding (bleeding academic research consortium(BARC) type 3-5) and definite/probable stent thrombosis within 48 months. The incidences of endpoint events were compared in the three groups. The multivariable Cox hazard ratio model was used to analyze the independent predictors of PoCE and TLF at 48 months. Results: Incidences of PoCE at 48 months were significantly higher in patients with type C lesion compared with patients with type A/B1 (24.43%(150/614) vs. 14.13%(26/184), P<0.05) or B2 lesion (24.43%(150/614) vs. 15.97%(73/457), P<0.05). The multivariable Cox hazard ratio model showed that the type C lesion were the independent predictors of 48-month PoCE (HR=1.59, 95%CI 1.21-2.08, P<0.001) and TLF (HR=2.31, 95%CI 1.53-3.49, P<0.001). After multivariable adjustment, the HRs of PoCE for patients with type C lesion versus type A/B1 and type B2 were 1.91 (95%CI 1.25-2.92, P=0.003) and 1.64 (95%CI 1.23-2.20, P<0.001), respectively. Meanwhile, the HRs of TLF for patients with type C lesion versus type A/B1 and type B2 were 2.45 (95%CI 1.29-4.64, P=0.006) and 2.55 (95%CI 1.62-4.02, P=0.001), respectively. Conclusions: The ACC/AHA classification of coronary lesions has good discrimination with long-term outcomes for CAD patients undergoing PCI. The type C lesion is associated with a worse prognosis, enough attention should be paid in these patients during routine clinical management.
Cardiovascular Agents ; Coronary Artery Disease ; Drug-Eluting Stents ; Humans ; Percutaneous Coronary Intervention ; Prognosis ; Prospective Studies ; Risk Assessment ; Risk Factors ; Sirolimus ; Treatment Outcome

Objective: To probe into the mechanism and interventional effects of silybin-phospholipid complex on amiodarone-induced steatosis in mice. Methods: Eight-week-old male C57BL/6 mice were divided into three groups (5 mice in each group): a control group (WT) with normal diet, a model group with amiodarone 150mg/kg/d by oral gavage (AM), and an intervention group on amiodarone 150mg/kg/d combined with silybin-phospholipid complex(AM+SILIPHOS. All mice were fed their assigned diet for one week. Then, one week later, serum alanine aminotransferase, aspartate aminotransferase, triglyceride, total cholesterol and high-density lipoprotein were detected of each group. A liver pathological change was observed by oil red O and H&E staining. Ultrastructural pathological changes of hepatocytes were observed to evaluate the intervention effect by transmission electron microscopy. RT-q PCR was used to detect the expression of peroxisome proliferator-activated receptor alpha and its regulated lipid metabolism genes CPTI, CPTII, Acot1, Acot2, ACOX, Cyp4a10 and Cyp4a14 in liver tissues. Intra-group comparison was done by paired t-test. One-way ANOVA was used for comparison between groups and semi-quantitative data were tested using Mann-Whitney U test. Results: Oil Red O and H&E staining results of liver tissue in the intervention group showed that intrahepatic steatosis was significantly reduced when compared to model group. Transmission electron microscopy showed that the model group had pyknotic nuclei, mitochondrial swelling, structural damage, and lysosomal degradation whereas the intervention group had hepatic nucleus without pyknosis, reduced mitochondrial swelling and slight structural damage than that of model group. RT-q PCR results showed that the expression of peroxisome proliferator-activated receptor alpha, CPTI, CPTII, Acot1, Acot2, ACOX, Cyp4a10 and Cyp4a14 were increased in the model group but the expression of CPTI, Cyp4a14, Acot1 and peroxisome proliferator-activated receptor alpha were decreased in the intervention group (P < 0.05). Conclusion Silybin-phospholipid complex can alleviate amiodarone-induced steatosis, and its mechanism may play a role in protecting mitochondrial function and regulating fatty acid metabolism. Thus, silybin-phospholipid complex has potential intervention effect on amiodarone-induced fatty liver.