AIM: To study the relationship between ApoE polymorphism and lipid metabolism of patients with cerebral infarction. METHODS: ApoE phenotype was determined in 110 patients with cerebral infarction and 60 normal controls by isoelectric focusing and immunoblotting. The TC, TG and HDL-C levels in serum of these subjects were measured with enzymes methods, ApoA I and ApoB levels with rocket immunoelectrophoresis methods, ApoE and Lp(a) levels with ELISA methods. RESULTS: The differences of the ApoE polymorphism distribution and ApoE allele frequencies ( P

AIM: To construct recombinant adenovirus vector containing brain derived neurotrophic factor, (BDNF) gene using bacterial homogenous recombination, and investigate the expression in expanded rat mesenchymal stem cells (rMSC) in vitro. METHODS: BDNF gene and proBDNF gene were subcloned into adenovirus shuttle plasmid pAdTrack-CMV containing enhanced green fluorescent protein gene (EGFP) expression cassette, forming shuttle vector of pAdTrack-BDNF, and pAdTrack-proBDNF, and co-transformed into BJ5183 bacterial cells with adenovirus backbone vector pAdEasy-1 using chemical transformation. After the recombinant adenovirus vector was obtained, the identified recombinant adenovirus plasmid DNA was digested with Pac I and transfected to 293 cells to package recombinant adenovirus particles. rMSC were infected by recombinant adenovirus and EGFP expression was detected using fluorescent microscope. Infection efficiency was assessed by flow cytometrics. Western blotting identified expression of Ad -proBDNF and Ad-BDNF in rMSC. rMSC infected with Ad -proBDNF and Ad-BDNF were induced to differentiate into neuron-like cells. rMSC infected with Ad -proBDNF and Ad-BDNF were injected into nude mice and assessd in vivo. RESULTS: We successfully constructed the recombinant adenovirus Ad -proBDNF and Ad-BDNF that expressed in expanded rMSC in vitro.CONCLUSION: Recombinant adenovirus high-effectively mediates Ad -proBDNF and Ad-BDNF expression in expanded rMSC in vitro and in vivo.

AIM: To study the effects of cotransplantation of donor-derived bone marrow mesenchymal stem cells on graft versus host disease in a rat allogeneic bone marrow transplantation model. METHODS: Fisher 344 rat bone marrow MSCs were isolated and cultured to the fifth passage (P5) in vitro . The recipient Wistar rats were conditioned with lethal total body irradiation and transplanted with F344 rat bone marrow cells and spleen cells in the presence or absence of (P5) MSCs. The onset time of graft versus host disease (GVHD), incidence of GVHD and survival time were monitored. RESULTS: Cotransplantation of MSCs deferred the onset time of GVHD[(19.1?1.7) d vs (15.6?1.5) d, P

AIM: To investigate the feasibility and infection efficiency of MSCs with replication-deficient adenovirus containing delivered gene, and whether enhanced green fluorescence protein (EGFP) gene track the change during rMSCs differentiating neuron-like cells. METHODS: Rat marrow mesenchymal stem cells (rMSCs) were expanded in low density in vitro . Under the control of CMV promoter, pAd-EGFP-Vector was constructed by homologous recombination in E.coil BJ 5183, and the recombinant virus was produced in HEK 293 packaging cell line. rMSCs infected with Ad-EGFP were observed and analyzed with fluorescence microscope. Infection efficiency was assessed by microscopical scoring and flow cytometrics. After withdrawing serum and exposure to ?-mercaptoethanol medium, rMSCs infected with Ad-EGFP was induced to differentiate into neuron-like cells. As a control, the plasmid of pTrack-EGFP also was transfected into rMSCs to evaluate transfection efficiency.RESULTS: The results showed that Adenovirus vector (AdVec) delivered EGFP gene with high efficiency to marrow mesenchymal stem cells. Gene expression analysis showed that 36%?2 % of rMSCs infected with recombinant adenovirus expressed the transgene of EGFP at high levels. However, the transfection of plasmid pTrack-EGFP using routine method of lipofectamin mixed with plasmid DNA (pTrack-EGFP) was not easily successful and the transfection efficiency was much lower. rMSCs infected with Ad-EGFP in different passage could differentiate into typical morphology alike neural cells after withdrawing serum and exposure to ?-mercaptoethanol medium. Immuno-staining with neuron-specific enolase (NSE), a neuronal marker, was strong positive, which suggested that rMSCs infected with Ad-EGFP had the potential to differentiate into neurons or neuron-like cells. CONCLUSION: The AdVec system can deliver target gene into MSCs and EGFP gene carried by AdVec can track the change during rMSCs differentiating into neuron-like cells.

AIM: To construct the plasmid vectors containing different regions of human eNOS promoter coupled to a red fluorescent protein reporter gene, which may express in mammalian cells. METHODS: Different regions of human eNOS promoter were subcloned respectively into a red fluorescent protein vector, pDsRed1-1. These recombinant vectors, pDsF1033Red, pDsF494Red and pDsF166Red, were then transfected into NIH3T3 cell lines, followed by the observation under a fluorescent microscope. RESULTS: After identified to be right by double restriction enzyme digestion, PCR and sequencing, the vectors might be effectively expressed in NIH3T3 cells. 95 % of the red fluorescent emitted by a red fluorescent protein dispersed all over the cells, appearing at 48-60 h after transfection, reaching peak at 96-144 h, becoming the strongest in light at 144 h, gradually disappearing after 168 h and remaining little red fluorescent in 21 days. The quantity and intensity in expressions of red fluorescent protein drived by different regions of human eNOS promoter were clearly lower than by a strong promoter, p CMVIE . CONCLUSION: The red fluorescent protein reporter gene vectors containing different regions of human eNOS promoter are successfully constructed and may efficaciously express in mammalian cells, appearing not strong transcriptional activities, which provide practical and feasible tools to study functions of different regions of human eNOS promoter and roles of cis-elements in it. [

BACKGROUND:At present,open reduction and internal fixation and minimally invasive needle aspiration are commonly used in patients with Sanders types Ⅱ and Ⅲ calcaneal fractures.However,there is little comparison between the clinical efficacy of the two methods and high-level clinical evidence is still available. OBJECTIVE:To compare the curative effect of Sanders types Ⅱ and Ⅲ calcaneal fractures treated by three-dimensional digital model-assisted minimally invasive needle penetration and tarsal sinus incision and manual reduction and internal fixation with steel plate. METHODS:From January 2021 to October 2022,80 patients with Sanders types Ⅱ and Ⅲ calcaneal fractures who were treated in the Department of Orthopedics,Cangzhou Hospital of Integrated Traditional Chinese and Western Medicine in Hebei Province were randomly divided into control group(40 cases)and observation group(40 cases).The control group was treated with manual reduction and internal fixation with steel plate through the traditional tarsal sinus incision,while the observation group was treated with a three-dimensional digital model assisted with minimally invasive needle penetration fixation.The operation time,blood loss,hospitalization time and fracture healing time of the two groups were recorded.The changes in Maryland score,AO-FAS score,pain visual analog scale score,quality of life score(SF-36 score),and imaging parameters(B?hler angle,Gissane angle,calcaneal length,width and height)were observed before and 12 months after operation in the two groups.The complications during the follow-up were recorded. RESULTS AND CONCLUSION:(1)Operation time,blood loss,hospitalization time and fracture healing time in the observation group were lower than those in the control group(all P<0.05).(2)The Maryland score,AO-FAS score,SF-36 score,B?hler angle,Gissane angle,calcaneal length and height of the two groups after treatment were significantly higher than those before treatment(all P<0.05).Visual analog scale score and calcaneal width were significantly lower than those before treatment(all P<0.05).(3)After 12 months of follow-up,the incidence of complications in the observation group was lower than that in the control group(all P<0.05).(4)In conclusion,the treatment of Sanders types Ⅱ and Ⅲ calcaneal fractures with three-dimensional digital model-assisted minimally invasive needle penetration fixation can significantly improve the operation time,bleeding volume and other perioperative indicators,and can reduce the occurrence of multiple complications.The recovery of ankle function,relief of pain symptoms,and improvement of quality of life are equivalent to traditional therapy.