Objective:To investigate the clinical phenotype and genetic characteristics of patients with Fabry disease caused by a GLA variant, IVS4+919G>A.Methods:It was a prospective study. Fabry disease screening was conducted among high-risk population in Ninghai from October 2021 to August 2023. Those children with decreased α-galactosidase enzyme activity<2.40 μmol/(L·h) or elavated Lyso-GL-3 level>1.10 μg/L in dried blood spot (DBS) method underwent GLA genetic testing for diagnosis confirmation. Meanwhile, family screening was carried out. A proband and his family members diagnosed with Fabry disease were research subjects. The clinical and genetic characteristics of patients with Fabry disease caused by the GLA variant (IVS4+919G>A) were analyzed.Results:The female proband aged 9.8 years with pain in both lower limbs as the initial symptom was found to have a heterozygous GLA variant IVS4+919G>A among 102 patients. In family screening, there were 4 family members (proband's father, elder sister, elder male cousin and elder female cousin) with Fabry disease and a family member (proband's fifth aunt) with a GLA variant. Among these 4 diagnosed family members, the elder male cousin of the proband, a boy aged 13.2 years had a heterozygous GLA variant, IVS4+919G>A with intermittent pain in both lower limbs as the initial symptom. The proband′s father had knee joint pain. The proband′s elder sister had decreased vision and his elder female cousin had no obvious symptoms. The proband′s fifth aunt with a GLA variant had decreased vision.Conclusions:High-risk screening in children and family screening are helpful for early diagnosis and treatment of Fabry disease. Neuropathic pain may be a early symptom in children with Fabry disease caused by the GLA variant, IVS4+919G>A.

With various diseases ravaging internationally, the demands for recombinant adenoviral vector (Adv) vaccines have increased dramatically. To meet the demand for Adv vaccine, development of a new cell culture process is an effective strategy. Applying hyperosmotic stress in cells before virus infection could increase the yield of Adv in batch culture mode. Emerging perfusion culture can significantly increase the yield of Adv as well. Therefore, combining the hyperosmotic stress process with perfusion culture is expected to improve the yield of Adv at high cell density. In this study, a shake flask combined with a semi-perfusion culture was used as a scaled-down model for bioreactor perfusion culture. Media with osmotic pressure ranging from 300 to 405 mOsm were used to study the effect of hyperosmotic stress on cell growth and Adv production. The results showed that using a perfusion culture process with a hyperosmotic pressure medium (370 mOsm) during the cell growth phase and an isosmotic pressure medium (300 mOsm) during the virus production phase effectively increased the yield of Adv. This might be due to the increased expression of HSP70 protein during the late phases of virus replication. The Adv titer in a bioreactor with such a process reached 3.2×10¹⁰ IFU/mL, three times higher than that of the traditional perfusion culture process. More importantly, this is the first time that a strategy of combining the hyperosmotic stress process with perfusion culture is applied to the production of Adv in HEK 293 cells. It also reveals the reason why the hyperosmotic stress process increased the yield of Adv, which may facilitate the process optimization of for producing other Adv in HEK 293 cells.
Humans ; HEK293 Cells ; Genetic Vectors/genetics* ; Batch Cell Culture Techniques ; Bioreactors ; Perfusion

PURPOSE: Allergic rhinitis (AR) is an inflammatory disorder of the upper airway. Exosomes or extracellular vesicles are nanosized vesicles of endosomal origin released from inflammatory and epithelial cells that have been implicated in allergic diseases. In this study, we characterized the microRNA (miRNA) content of exosomes in AR. METHODS: Extracellular vesicles were isolated from nasal mucus from healthy control subjects (n=10) and patients with severe AR (n=10). Vesicle RNA was analyzed by using a TaqMan microRNA assays Human Panel-Early Access kit (Applied Biosystems, Foster City, CA, USA) containing probes for 366 human miRNAs, and selected findings were validated with quantitative RT-PCR. Target prediction and pathway analysis for the differentially expressed miRNAs were performed using DIANA-mirPath. RESULTS: Twenty-one vesicle miRNAs were up-regulated and 14 miRNAs were under-regulated significantly (P<0.05) in nasal mucus from AR patients when compared to healthy controls. Bioinformatic analysis by DIANA-mirPath demonstrated that 32 KEGG biological processes were significantly enriched (P<0.05, FDR corrected) among differentially expressed vesicle miRNA signatures. Among them, the B-cell receptor signaling pathway (P=3.709E-09), the natural killer cell-mediated cytotoxicity (P=8.466E-05), the T-cell receptor signaling pathway (P=0.00075), the RIG-I-like receptor signaling pathway (P=0.00127), the Wnt signaling pathway (P=0.00130), endocytosis (P=0.00440), and salivary secretion (P=0.04660) were the most prominent pathways enriched in quantiles with differential vesicle miRNA patterns. Furthermore, miR-30-5p, miR-199b-3p, miR-874, miR-28-3p, miR-203, and miR-875-5p, involved in B-cell receptor and salivary secretion signaling pathways, were selected for validation using independent samples from 44 AR patients and 20 healthy controls. MiR-30-5p and miR-199b-3p were significantly increased in extracellular vesicles from nasal mucus when compared to healthy controls, while miR-874 and miR-28-3p were significantly down-regulated. In addition, miRNA-203 was significantly increased in AR patients, while miRNA-875-5p was found to be significantly decreased in AR patients. CONCLUSIONS: This study demonstrated that vesicle miRNA may be a regulator for the development of AR.
B-Lymphocytes ; Biological Processes ; Endocytosis ; Epithelial Cells ; Exosomes ; Humans ; MicroRNAs* ; Mucus* ; Receptors, Antigen, T-Cell ; Rhinitis* ; RNA ; Wnt Signaling Pathway

Objective To discuss the feasibility of biodegradable covered magnesium alloys stent for a rabbit model of lateral aneurysm in the common carotid artery (CCA). Methods The left CCA was ligated in 20 conventional New Zealand rabbits. MRA of the neck was performed 1 month after ligation. The rabbits with thickening of the right CCA and non-or slight thickening of the bilateral vertebral arteries were selected for lateral aneurysm model making. The venous pouch and the right CCA wall by discontinuous extroversion sutures, to form a lateral aneurysm model. The biodegradable covered magnesium alloys stents or Willis covered stents were inserted two weeks after model making. Angiography was performed prior to the procedure, 3, 6 and 12 months after stent implantation to evaluate the disappearance of the aneurysms and patency of the right CCA. The degradation behaviour is invastagated with molybdenum target 2 weeks, 1, 2, 4, 6,9 and 12 months after stent placement. Results The left CCAs were successful ligated in all rabbits. MRA 1 month after ligation showed thickening of the right CCA and non-thickening of the bilateral vertebral arteries in 17 of 20 rabbits. In these animals, the CCA lateral aneurysm model was deemed successful, and biodegradable covered magnesium alloys stents and Willis covered stents were implanted in 9 and 8 aneurysms, respectively. DSA after biodegradable covered magnesium alloys stent placement displayed disappearance of the aneurysms and patency of the CCA in all 9 rabbits during follow-ups. DSA 3 months after Willis stent placement displayed patency of the CCA in 7 rabbits and occlusion of the artery in one. No occlusion of the right CCA was observed on angiography at 6 and 12 months. The degradation of the biodegradable covered magnesium alloys stent was investigated with molybdenum target during follow-ups, and no changes was observed in Willis covered stent. Conclusion Biodegradable covered magnesium alloys stent is a feasible approach for the treatment of a lateral aneurysm in the right CCA.

This study was aimed to determine effect of Bu-Shen Kang-Shuai (BSKS) Tablet on HO-1 mRNA and its associated oxidative stress levels among atherosclerotic rabbits. A total of 56 rabbits were randomly divided into the normal group (8 rabbits) and the experimental group (48 rabbits). Normal diet was given to the normal group. Atherosclerotic rabbits models were established in the experimental group. At the eighth week, rabblits in the experi-mental group were randomly divided into the model group, BSKS Tablet group and simvastatin group. Blood samples were collected before medication, 8-, 12-, 16-week after medication from rabbits of each group. Rabbits were sacri-ficed under aseptic conditions at the last blood collection. Expressions of aortic HO-1 mRNA and PPARα mRNA were measured by Q-PCR method. The level of MMP-9 was measured by immunohistochemical assay. Serum HbCO, COX-2 activity and cGMP level were measured by ELISA assay. The results showed that after the intervention of BSKS Tablet, serum HbCO level decreasd, cGMP was obviously increased. However, there was no obvious change on the COX-2 activity. The immunohistochemical assay showed that BSKS Tablet obviously reduced MMP-9 level of rabbits. There was only small amount of aortic HO-1 mRNA expression in the normal group. However, the expres-sion of aortic HO-1 mRNA in the atherosclerosis group was increased. After intervention of BSKS Tablet, the ex-pression of HO-1 mRNA was increased with statistical significance (P < 0.05). Simvastatin had similar antioxidant effect. It was concluded that the compound preparation of traditional Chinese medicine (TCM) BSKS Tablets had an important antioxidant effect in treatment of atherosclerosis. Its protective mechanism may be through the regulation of HO-1 mRNA gene expression and effects of HO-1/CO-cGMP pathway activities of related enzymes while peroxida-tion stability of atherosclerotic plaque.

Objective To investigate the expression of connective tissue growth factor(CTGF),coll agen I and collagen Ⅲ in sacroiliac joint(SIJ)of patients with spondyloarthropathy(SpA).Methods Thirty patients with SpA,including 17 patients with grade Ⅱ saeroiliitis and 13 patients with grade Ⅰ sacroiliitis,were performed on CT guided needie biopsy of SIJ.After sacroiliitis were confirmed by staining with hematoxylin and eosin in sacroiliac joint tissue sample,immunohistochemical assay was performed to determine the expression of CTGF,collagen Ⅰ and collagen Ⅲ in sacroiliac ioint tissue.Univariate Chi-square test was used for data comparison between multiple groups and t-test was used for two group data comparison.Results Contrast to healthy controls,CTGF were found upexpressed on the cytoplasm of inflammatory cells in pannus and bone marrow of sacroiliac tissue samples of patients with SpA,while collagen I and collagen Ⅲ were found up-expressed in bone,cartilage and ligament tissue[(57.9±42.4)/HP vs(2.7±2.5)/HP P<0.05,0.298±0.080 vs 0.044±0.024 and 28.254±41.165 vs 0.105±0.054.P<0.05 respectively].Conclusion CTGF,collagen Ⅰ and collagen Ⅲ are up-expressed in SIJ of SpA patients.CTGF may play an important role in articular cartilage fibrosis and ossification of SpA.

Objective To investigate the expression of transforming growth factor beta (TGF-β) and Smad signaling in ankylosing spondylitis (AS) and to explore their roles in the pathogenesis. Methods Fiftythree patients with AS were included in the study. In these 53 cases, 30 patients were performed computed tomography-guided needle biopsy in sacroiliac joint. Serum TGF-β_1 was determined by enzyme-linked immunosorbent assay (ELISA). Immunohistological studies were performed with the streptavidin-peroxidase conjugated methods to assess the expression of TGF-β_1, p-smad3 and Smad7 in sacroiliac joint tissue sample.One-way ANOVA, two independent samples t test and kolmogoorov-Simimov test were used to do statistical analysis. Results In 53 cases patients with AS, 20 cases were with high level Erythro-cyte sedimentation rate(ESR) and C-reactive protein (CRP), while those of the other 33 cases were normal. Serum average TGF-β_1level [ (15.9±5.6) ng/ml ], in patients with high level ESR/CRP [(5.4±5.8) ng/ml ] was significantly increased as compared to the controls and patients with normal ESR/CRP [(4.1±3.6) ng/ml] (P<0.05). There was no expression of TGF-β_1 could be detected in the pannus and bone marrow in SI joints tissue of 30 cases with AS, while decreased level of smad7 expression was detected. In addition, p-smad3 expression was found in the nuclear. Conclusion TGF-β_1 signaling may play an important role in the inflammatory erosion and cartilage fibrosis of sacrojlitis in AS.

Objective: To investigate the distribution features of somatosensory cortical evoked potential map with augmented blocking of the sensation transmission along meridians.Method: The EEG-4400 electro-encephalogram (EEG) and ND-1 brain electrical activity mapping were adopted on 11 volunteers with remarkable sensation transmission along meridians, showing that the sensation can transmit to head and face after stimulating the points below the knee joints. Also, special observation was made on accurate location of somatosensory cortical evoked potential map in 10 people without sensation transmission.Result: Observation on 11 volunteers with remarkable transmission along the Three Foot-yang Meridians showed that they presented with concurrent high potential reactions in somatosensory cortical lower limbs and face without blocking the augmented sensation transmission along the meridians; however, when mechanical pressure was exerted to block the sensation transmission,only one reaction in the lower limbs occurred in the somatosensory cortical evoked map and the other one in the face disappeared. Conclusion: Peripheral tissue evoking is the decisive factor for transmission along the meridians.

0.05)and could be induced significantly after 12 h (P

Objective:To observe whether the Simiao Yongan Decoction regulate the nuclear factor-?B(NF-?B) expression and related factors in atherosclerosis(AS)rabbit model,thus restrain the occurrence of atherosclerosis and plaque formation.Methods:56 male Japanese rabbits were randomly divided into normal group,model group,Simvastatin group and Simiao Yongan Decoction group,in addition to the normal group,other groups were established aortic atherosclerotic plaque model.From the1st day of experiment,the corresponding drug intervention began,10 weeks later,the rabbits were killed over the weekend after the anesthesia line access aortic NF-?B immunohistochemical staining,and the experimental dynamic 0,3,6,10 weeks in serum TNF-?,IL-1 and MCP-1 content.Results:The expression of NF-?B p65 subunit in model group was rich,and compared with it,the expression of NF-?B p65 subunit in treatment groups decreased,and the Simiao Yongan Decoction can obviously inhibit the expression of NF-?B p65 subunit.In model group,the expressions of IL-1,TNF-?and MCP-1 in serum increased significantly at different time points(P