ObjectiveTo investigate the mechanism by which Notoginsenoside R1 （NGR1） ameliorates hypoxia/reoxygenation （H/R）-induced injury in AC16 human cardiomyocyte cell lines through the regulation of mitophagy. MethodsCommon genes linked to hypoxia/reoxygenation injury and mitophagy were identified by intersecting data from GeneCards and MitoCarta databases. AC16 cell viability was assessed via CCK-8 assay under varying NGR1 concentrations （0， 6.25， 12.5， 25， 50， 100， 200， 300， 400， 500 μmol/L）. AC16 cells were divided into the following groups： control group （Control）， model group （H/R）， and treatment groups （H/R + NGR1 at 100， 200 and 300 μmol/L）. Mitochondrial membrane potential （ΔΨm） was measured using 5，5'，6，6'-tetrachloro-1，1'，3，3'-tetraethylbenzimidazolylcarbocyanine iodide （JC-1） staining. Transcriptional levels of mitophagy-related genes （Parkin， Pink1， P62） were quantified by reverse transcription-quantitative PCR （RT-qPCR）. Protein expression of mitophagy-related markers （Parkin， Pink1， P62， and LC3BⅡ） was evaluated via Western blot analysis. Mitochondrial ultrastructure was visualized by transmission electron microscopy （TEM）. ResultsCompared to the control group， cell viability in the H/R group significantly decreased （P<0.01）. Treatment with NGR1 at concentrations above 100 μmol/L significantly enhanced the cell viability of AC16 cells compared to the H/R group （P<0.01）. H/R induced a significant decrease in mitochondrial membrane potential （P<0.01）， which was restored by NGR1 treatment （P<0.01）. The mRNA levels of Parkin， Pink1， and P62 in the H/R group were upregulated compared to the control group （P<0.05）， while NGR1 intervention downregulated their expression （P<0.05）. Protein expression levels of Parkin， Pink1， and LC3BⅡ in the H/R group significantly increased， while P62 expression decreased compared to the control group （P<0.01）. In contrast， different doses of NGR1 treatment significantly reduced the expression of Parkin， Pink1， and LC3BⅡ while increasing P62 expression （P<0.05）. TEM revealed that the mitochondrial structure in the H/R group was severely disrupted， with fragmented and disorganized cristae， which was alleviated by NGR1. ConclusionNGR1 ameliorates H/R-induced AC16 cell injury， and its mechanism may be associated with modulating the Pink1/Parkin pathway to suppress excessive mitophagy.

ObjectiveSpinal cord injury (SCI) directly impairs the regulatory function of the autonomic nervous system, induces intestinal dysfunction, and significantly reduces patients’ quality of life. Preclinical studies have shown that electroacupuncture (EA) therapy can regulate the brain-gut axis and is used to treat central nervous system diseases such as major depressive disorder, Alzheimer’s disease and Parkinson’s disease. Recent research has established that fecal microbiota transplantation (FMT) from EA-treated SCI rats restored intestinal motility and colonic morphology. However, it remains unclear whether the regulation of gut microbiota by EA therapy directly contributes to neural repair after SCI. This study aims to explore whether gut microbiota mediates the neuroprotective effect of EA in the treatment of SCI and its possible mechanism. MethodsThe study employed RNA transcriptome analysis of spinal cord tissue to characterize gene expression profiles and to identify key signaling pathways following EA treatment for SCI. Hematoxylin-Eosin (HE) staining and Nissl staining were used to observe the morphological changes in spinal cord tissue. Western blot (WB) and enzyme-linked immunosorbent assay (ELISA) were applied to detect the effects of EA on the expression of proteins related to nucleotide-binding domain leucine-rich repeat and pyrin domain-containing receptor 3 (NLRP3) -dependent pyroptosis. Using 16S rDNA sequencing, the study observed alterations in gut microbiota diversity and community composition in SCI rats. Prior to establishing SCI models, rats were pretreated with an antibiotic cocktail to induce gut dysbiosis, and the effects on intestinal function and spinal cord neural repair were evaluated. FMT was performed to investigate the regulatory effects of post-EA FMT on motor function, general status, liver and spleen indices, and NLRP3-mediated pyroptosis in SCI rats. ResultsEA improved motor function and reduced regulated neuronal cell death in SCI rats. Transcriptomic analysis demonstrated the activation of immune- and inflammation-related pathways post-SCI, including NOD-like receptors, nuclear factor-kappa B(NF-κB), and Toll-like receptor (TLR) pathways. EA primarily influenced intestinal inflammation and autoimmune functions. 16S rDNA sequencing illustrated that EA did not alter the diversity of gut microbiota. However, EA altered the gut microbiota composition in SCI rats, increasing Lactobacillus and Akkermansia genera while rebalancing the Firmicutes/Bacteroidetes ratio. Furthermore, depletion of gut microbiota by antibiotics disrupted the intestinal barrier, reduced the expression of intestinal barrier proteins Zonula Occludens-1 (ZO-1) and Occludin, elevated serum lipopolysaccharide-binding protein (LBP) levels, exacerbated spinal cord tissue damage, and hindered motor function recovery in SCI rats. FMT from donors treated with EA reduced LBP levels in the intestine, blood, and spinal cord of rats, inhibited the TLR4 myeloid differentiation primary response protein 88 (MyD88)-NF‑κB pathway and NLRP3-dependent pyroptosis, and improved motor function. On the other hand, FMT treatment resulted in decreased body weight and food intake, whereas FMT using EA-treated donors effectively alleviated these alterations. ConclusionEA effectively alleviated neuroinflammatory responses in rats with SCI, primarily through regulating the gut microbiota and suppressing the NLRP3-dependent pyroptosis signaling pathway.

ObjectiveSpinal cord injury (SCI) directly impairs the regulatory function of the autonomic nervous system, induces intestinal dysfunction, and significantly reduces patients’ quality of life. Preclinical studies have shown that electroacupuncture (EA) therapy can regulate the brain-gut axis and is used to treat central nervous system diseases such as major depressive disorder, Alzheimer’s disease and Parkinson’s disease. Recent research has established that fecal microbiota transplantation (FMT) from EA-treated SCI rats restored intestinal motility and colonic morphology. However, it remains unclear whether the regulation of gut microbiota by EA therapy directly contributes to neural repair after SCI. This study aims to explore whether gut microbiota mediates the neuroprotective effect of EA in the treatment of SCI and its possible mechanism. MethodsThe study employed RNA transcriptome analysis of spinal cord tissue to characterize gene expression profiles and to identify key signaling pathways following EA treatment for SCI. Hematoxylin-Eosin (HE) staining and Nissl staining were used to observe the morphological changes in spinal cord tissue. Western blot (WB) and enzyme-linked immunosorbent assay (ELISA) were applied to detect the effects of EA on the expression of proteins related to nucleotide-binding domain leucine-rich repeat and pyrin domain-containing receptor 3 (NLRP3) -dependent pyroptosis. Using 16S rDNA sequencing, the study observed alterations in gut microbiota diversity and community composition in SCI rats. Prior to establishing SCI models, rats were pretreated with an antibiotic cocktail to induce gut dysbiosis, and the effects on intestinal function and spinal cord neural repair were evaluated. FMT was performed to investigate the regulatory effects of post-EA FMT on motor function, general status, liver and spleen indices, and NLRP3-mediated pyroptosis in SCI rats. ResultsEA improved motor function and reduced regulated neuronal cell death in SCI rats. Transcriptomic analysis demonstrated the activation of immune- and inflammation-related pathways post-SCI, including NOD-like receptors, nuclear factor-kappa B(NF-κB), and Toll-like receptor (TLR) pathways. EA primarily influenced intestinal inflammation and autoimmune functions. 16S rDNA sequencing illustrated that EA did not alter the diversity of gut microbiota. However, EA altered the gut microbiota composition in SCI rats, increasing Lactobacillus and Akkermansia genera while rebalancing the Firmicutes/Bacteroidetes ratio. Furthermore, depletion of gut microbiota by antibiotics disrupted the intestinal barrier, reduced the expression of intestinal barrier proteins Zonula Occludens-1 (ZO-1) and Occludin, elevated serum lipopolysaccharide-binding protein (LBP) levels, exacerbated spinal cord tissue damage, and hindered motor function recovery in SCI rats. FMT from donors treated with EA reduced LBP levels in the intestine, blood, and spinal cord of rats, inhibited the TLR4 myeloid differentiation primary response protein 88 (MyD88)-NF‑κB pathway and NLRP3-dependent pyroptosis, and improved motor function. On the other hand, FMT treatment resulted in decreased body weight and food intake, whereas FMT using EA-treated donors effectively alleviated these alterations. ConclusionEA effectively alleviated neuroinflammatory responses in rats with SCI, primarily through regulating the gut microbiota and suppressing the NLRP3-dependent pyroptosis signaling pathway.

ObjectiveTo investigate the inhibitory effect of gingerol， an active component of ginger， on Staphylococcus aureus （S. aureus） infection， and to preliminarily explore its mechanism related to extracellular vesicles （EVs）. MethodsS.aureus infection models were established in human umbilical vein endothelial cell （HUVECs）， Vero E6 cells， and C57BL/6 mice. Experimental groups included control， infection， and gingerol-treated groups. Bacterial load and VE-cadherin protein expression were detected using immunofluorescence and Western blot. EVs were isolated by size exclusion chromatography and characterized by transmission electron microscopy and nanoparticle tracking analysis. miRNA sequencing was performed on EVs. ResultsGingerol treatment significantly reduced the bacterial load in both in vitro and in vivo infection models and upregulated VE-cadherin expression. miRNA sequencing of EVs revealed that S. aureus infection upregulated the expression of hsa-miR-320c， while gingerol treatment reversed this abnormal expression. Bioinformatic analysis further predicted that the target genes of hsa-miR-320c were significantly enriched in cell junction-related pathways. ConclusionGingerol exhibits clear antibacterial and host-protective effects， by regulating hsa-miR-320c in EVs to maintain endothelial barrier integrity.

In recent years, cancer treatment methods and means are becoming more and more diversified, and single treatment methods often have limited efficacy, while the synergistic effect of immunity combined with chemotherapy can inhibit tumor growth more effectively. Based on this, we constructed a sodium alginate hydrogel composite system loaded with chemotherapeutic agents and tumor vaccines (named SA-DOX-NA) with a view to the combined use of chemotherapeutic agents and tumor vaccines. Firstly, the tumor vaccine (named NA) degradable under acidic conditions was constructed by in situ polymerization using chicken ovalbumin (OVA), acrylamide (AAM) and 2-(dimethylamino) ethyl methacrylate (DMAEMA) monomer. Then a hydrogel composite system SA-DOX-NA co-loaded with chemotherapeutic drug doxorubicin (DOX) and NA was prepared using sodium alginate as a matrix. The results showed that SA-DOX-NA had good in situ gel-forming ability and formed a mesh structure that could realize the co-loading of DOX and NA as well as the slow drug release. The electron microscopy results showed that SA-DOX-NA had good in situ gel-forming ability with good internal connectivity, and the three-dimensional mesh structure could realize the co-loading of DOX and NA as well as the slow drug release. The antitumor and immunomodulatory results showed that SA-DOX-NA both effectively inhibited the growth of tumor cells and efficiently promoted the proliferation and activation of DC2.4 dendritic cells without additional adjuvant. In summary, SA-DOX-NA exerts the dual efficacy of chemotherapy and immunotherapy for tumor treatment, and has a good application prospect in local tumor treatment.

Objective: To design and establish a new method for flow cytometry-based detection of commonly observed highly expressed antigens on red blood cells, and to further evaluate the differences and distribution characteristics of antigen expression levels between ABO blood type homozygotes and heterozygotes in healthy individuals. Methods: Residual blood samples after donor blood type identification by Shanghai Blood Center in April 2024 were collected. Among them, samples of 19 homozygous and 19 heterozygous individuals of type A and type B were selected. Then the expression level of ABO antigen on red blood cells were detected using the new method established in this study and the traditional aldehyde fixed red blood cell method. Both methods were tested independently three times and the results were compared. Results: The mean values of the three detection results of the new method was (×10 /RBC): AA homozygous 3.3±0.5, AO heterozygous 2.8±0.3, BB homozygous 3.6±0.3, BO heterozygous 3.1±2.8. The mean values of the three detection results of the aldehyde fixation method were AA homozygous 5.9±0.9, AO heterozygous 5.0±1.4, BB homozygous 3.8±0.6, and BO heterozygous 3.3±0.4. The average antigen distribution of each genotype followed a normal distribution. Comparing the average antigen expression levels of homozygotes and heterozygotes, both methods showed that A/B homozygotes had higher antigen levels than heterozygotes, with AA being 1.17 to 1.18 times that of AO and BB being 1.15 to 1.16 times that of BO. Comparing the inter batch differences in the three test results of two methods, the new method showed no significant difference in the three test results for four genotypes (P>0.05). The aldehyde fixation method showed significant differences in the test results for all three genotypes (P<0.01) except for BB homozygotes (P>0.05). The reliability and reproducibility of the new method were better than those of the traditional aldehyde fixation method. Conclusion: The antigen expression level of ABO homozygotes is higher than that of heterozygotes, and the difference in antigen level between type A homozygotes and heterozygotes is slightly higher than that of type B. The new method is superior to traditional aldolization fixation methods.

AIM: To delineate the specific mutation responsible for retinitis pigmentosa(RP)in a Yi pedigree, and to analyze the correlation of RHO gene mutation with clinical phenotype.METHODS:A comprehensive clinical evaluation was conducted on the proband diagnosed with RP and other familial members, complemented by a thorough ophthalmic examination. Peripheral blood samples were obtained from the proband and familial members, from which genomic DNA was extracte. Subsequent whole exome sequencing(WES)was employed to identify the variant genes in the proband. The identified variant gene was validated through Sanger sequencing, then an in-depth analysis of the mutation genes was carried out using genetic databases to ascertain the pathogenic mutation sites. Furthermore, an exhaustive analysis was performed to delineate the genotype and phenotype characteristics.RESULTS:The RP pedigree encompasses 5 generations with 42 members, including 19 males and 23 females. A total of 13 cases of RP were identified, consisting of 4 males and 9 females, which conforms to the autosomal dominant inheritance pattern. The clinical features of this family include an early onset age, rapid progression, and a more severe condition. The patients were found to have night blindness around 6 years old, representing the earliest reported case of night blindness in RP families. The retina was manifested by progressive osteocytoid pigmentation of the fundus, a reduced visual field, and significantly decreased or even vanished a and b amplitudes of ERG. The combined results of WES and Sanger sequencing indicated that the proband had a heterozygous missense mutation of the RHO gene c.1040C>T:p.P347L, where the 1 040 base C of cDNA was replaced by T, causing codon 347 to encode leucine instead of proline. Interestingly, this mutation has not been reported in the Chinese population.CONCLUSION:This study confirmed that the mutant gene of RP in a Yi nationality pedigree was RHO(c.1040C>T). This variant leads to the change of codon 347 from encoding proline to encoding leucine, resulting in a severe clinical phenotype among family members. This study provides a certain molecular, clinical, and genetic basis for genetic counseling and gene diagnosis of RHO.

AIM: To delineate the specific mutation responsible for retinitis pigmentosa(RP)in a Yi pedigree, and to analyze the correlation of RHO gene mutation with clinical phenotype.METHODS:A comprehensive clinical evaluation was conducted on the proband diagnosed with RP and other familial members, complemented by a thorough ophthalmic examination. Peripheral blood samples were obtained from the proband and familial members, from which genomic DNA was extracte. Subsequent whole exome sequencing(WES)was employed to identify the variant genes in the proband. The identified variant gene was validated through Sanger sequencing, then an in-depth analysis of the mutation genes was carried out using genetic databases to ascertain the pathogenic mutation sites. Furthermore, an exhaustive analysis was performed to delineate the genotype and phenotype characteristics.RESULTS:The RP pedigree encompasses 5 generations with 42 members, including 19 males and 23 females. A total of 13 cases of RP were identified, consisting of 4 males and 9 females, which conforms to the autosomal dominant inheritance pattern. The clinical features of this family include an early onset age, rapid progression, and a more severe condition. The patients were found to have night blindness around 6 years old, representing the earliest reported case of night blindness in RP families. The retina was manifested by progressive osteocytoid pigmentation of the fundus, a reduced visual field, and significantly decreased or even vanished a and b amplitudes of ERG. The combined results of WES and Sanger sequencing indicated that the proband had a heterozygous missense mutation of the RHO gene c.1040C>T:p.P347L, where the 1 040 base C of cDNA was replaced by T, causing codon 347 to encode leucine instead of proline. Interestingly, this mutation has not been reported in the Chinese population.CONCLUSION:This study confirmed that the mutant gene of RP in a Yi nationality pedigree was RHO(c.1040C>T). This variant leads to the change of codon 347 from encoding proline to encoding leucine, resulting in a severe clinical phenotype among family members. This study provides a certain molecular, clinical, and genetic basis for genetic counseling and gene diagnosis of RHO.

BACKGROUND:The small-diameter blood vessel prosthesis faces the problem of lumen stenosis or even occlusion due to thrombogenesis and intimal hyperplasia after transplantation.The application of stem cells as seed cells to achieve endothelialization of blood vessel prosthesis helps to improve the long-term patency rate after vascular graft transplantation. OBJECTIVE:To summarize the research progress of the application of stem cells in the endothelialization of small-diameter blood vessel prosthesis. METHODS:The relevant articles published on PubMed and WanFang databases from 2013 to 2023 were retrieved by the first author.Chinese and English search terms included"vascular graft,tissue-engineered blood vessel/vascular tissue engineering,endothelialization,stem cells,endothelial progenitor cells,mesenchymal stem cells,induced pluripotent stem cells,embryonic stem cells."The relative articles in the domestic and overseas about the application of stem cells in the endothelialization of small-diameter blood vessel prosthesis in the past 10 years were retrieved.A total of 552 articles were initially found and we finally selected 81 articles to review according to inclusion and exclusion criteria. RESULTS AND CONCLUSION:(1)The low long-term patency rate restricts the application of small-diameter blood vessel prosthesis in clinic.The main causes of low long-term patency rate are thrombogenesis and intimal hyperplasia.The endothelium of native vessels has the function of anti-thrombogenesis and intimal hyperplasia.Endothelialization can simulate the function of native vessels,which is an effective way to improve long-term patency rate.(2)The small-diameter blood vessel prosthesis will undergo in vivo endothelialization after being implanted,but it is difficult to form complete endothelium.Stem cells have the potential to differentiate into endothelial cells.Recruiting stem cells in vivo or seeding them on the inner surface of blood vessel prosthesis in vitro is research strategy to achieve endothelialization.(3)The long-term patency rate of small-diameter blood vessel prosthesis has been improved to a certain extent through seeding endothelial progenitor cells,mesenchymal stem cells,induced pluripotent stem cells,and embryonic stem cells.Each has its own advantages.Endothelial progenitor cells are easy to obtain and can be directly used for seeding.Mesenchymal stem cells come from a wide range of sources and have the function of paracrine and immunological regulation.Induced pluripotent stem cells are rich in sources and the immunogenicity can be eliminated.Embryonic stem cells have a strong proliferative ability and can differentiate into many cells.(4)The application of stem cells in blood vessel prosthesis has not yet been transformed into clinic.Further researches are needed to promote clinical translation.

OBJECTIVES: To analyze the efficacy and safety of concomitant left atrial appendage clipping during heart valve surgery for valvular heart disease patients with atrial fibrillation. METHODS: Fifty-eight patients who underwent concomitant left atrial appendage clipping during cardiac valve surgery in the Second Affiliated Hospital of Nanchang University from January 2017 to June 2023 were included in the analysis, including 1 case who underwent aortic valve replacement, 49 cases who underwent mitral valve replace-ment (or valvuloplasty)+tricuspid valvuloplasty, and 8 cases who underwent double valve replacement+tricuspid valvuloplasty (3 cases combined with coronary artery bypass grafting). The patients were followed up for 3-36 months [(16.69±6.61) months] after operation, and the changes of cardiac function and the occurrence of serious adverse complications were evaluated. RESULTS: The cardiopulmonary bypass time ranged from 75 to 145 min [(102.50±21.03) min], and the aortic cross-clamp time ranged from 35 to 80 min [(58.02±14.63) min]. The length of postoperative intensive care unit stay was 1 to 5 days [(2.47±0.82) d], and the length of postoperative hospital stay was 7 to 22 days [(10.84±2.69) d]. Cardiac ultrasound indicated complete closure of the left atrial appendage in all cases. During the follow-up, New York Heart Association (NYHA) functional classifications were improved in 54 patients. No left atrial appendage-related bleeding events or other perioperative complications were observed; and no cerebral infarction, limb embolism events, or mortality cases occurred during the follow-up. CONCLUSIONS For valvular heart disease patients with atrial fibrillation, concomitant left atrial appendage clipping during cardiac valve surgery demonstrates efficacy and safety, with no severe adverse events during a medium-term follow-up.
Humans ; Atrial Appendage/surgery* ; Atrial Fibrillation/complications* ; Male ; Female ; Heart Valve Diseases/complications* ; Aged ; Middle Aged ; Heart Valve Prosthesis Implantation/methods* ; Treatment Outcome ; Cardiac Surgical Procedures/methods* ; Mitral Valve/surgery*