ABSTRACT
In order to construct a recombinant replication deficient human type 5 adenovirus (Ad5) expressing a foot-and-mouth disease virus (FMDV) capsid protein, specific primers for P12A and 3B3C genes of FMDV-OZK93 were synthesized. The P12A and 3B3C genes were then amplified and connected by fusion PCR, and a recombinant shuttle plasmid pDC316-mCMV-EGFP-P12A3B3C expressing the FMDV-OZK93 capsid protein precursor P12A and 3B3C protease were obtained by inserting the P12A3B3C gene into the pDC316-mCMV-EGFP plasmid. The recombinant adenovirus rAdv-P12A3B3C-OZK93 was subsequently packaged, characterized and amplified using AdMaxTM adenovirus packaging system, and the expression was verified by infecting human embryonic kidney cell HEK-293. The humoral and cellular immunity levels of well-expressed and purified recombinant adenovirus immunized mice were evaluated. The results showed that rAdv-P12A3B3C-OZK93 could be stably passaged and the maximum virus titer reached 1×109.1 TCID50/mL. Western blotting and indirect immunofluorescence showed that rAdv-P12A3B3C-OZK93 expressed the FMDV-specific proteins P12A and VP1 in HEK-293 cells. In addition, the PK cell infection experiment confirmed that rAdv-P12A3B3C-OZK93 could infect porcine cells, which is essential for vaccination in pigs. Comparing with the inactivated vaccine group, the recombinant adenovirus could induce higher FMDV-specific IgG antibodies, γ-IFN and IL-10. This indicates that the recombinant adenovirus has good immunity for animal, which is very important for the subsequent development of foot-and-mouth disease vaccine.
Subject(s)
Animals , Humans , Mice , Adenoviridae/genetics , Adenoviruses, Human/genetics , Antibodies, Viral , Capsid/metabolism , Capsid Proteins , Foot-and-Mouth Disease/prevention & control , Foot-and-Mouth Disease Virus/genetics , HEK293 Cells , Recombinant Proteins/genetics , Serogroup , Swine , Viral Proteins , Viral Vaccines/geneticsABSTRACT
Objective To analysis the DNA sequence of HEV ORF3 cDNA and express in HepG2 cell .Methods Recombinant vector carrying HEV ORF3 cDNA was constructed and identified by DNA sequencing .Expression of recombinant vector in HepG2 cell was determined by immunofluorescence technique .Results The recombinant vector was constructed and expressed in HepG2 cell successfully .Conclusion The research laid experimental foundation for exploring the biological function of ORF3 protein in the pathogenesis of viral hepatitis E .
ABSTRACT
BACKGROUND: Muscle-derived stem cells (MDSCs) have been accepted as seeding cells in tissue engineered artificial nerves. Tacrolimus exhibits anti-immunologic rejection and promotes nerve regeneration and recovery. Whether can combine these factors with acellular nerve to form a new bridge that can inhibit immunologic rejection and promote nerve regeneration is uncertain. OBJECTIVE: Using the freeze-thawing combined optimized acellular nerve hypotonic-chemical detergent to prepare acellular nerveallograft scaffold. To explore the effect of co-application of MDSCs and FK-506 on nerve regeneration and function recovery following acellular nerve ailograft implantation.METHODS: The sciatic nerve derived from SD rats was prepared nerve bridge after acallular disposal. Gel containing FK-506 and MDSCs was injected into acellular nerveallograft scaffold with 100 μL microsyringe to repair defects. A total of 32 SD rats were randomly divided into 4 groups, with 8 animals in each group. Agap of 10 mm of left sciatic nerve was removed. And then the defects were repaired by extracted nerve graft containing FK-506+MDSCs, MDSCs and FK-506, respectively. In the control group, only hyaluronic acid gel was injected. Sciatic nerve function index (SFI) and electrophysiological exam were performed at weeks 8 and 12 after operation. Gross observation, neurohistological and ultrastructure were observed at week 12 after operation.RESULTS AND CONCLUSION: Compared with the same time point, the SFI, recovery rate of the motor nerve conduction velocities (MNCV), and myelinated nerve fibers in grafting part and in its distal part in the FK-506+MDSCs group were superior to other groups (P < 0.05). The nerve grafts were in normal size with considerabie blood vessels and slightly connected to peripheral Ussues. Compared to other groups, the regenerated nerve fiber in the FK-506+MDSCs group was more density with well-arranged order. A great quantity of Schwann cells proliferated in grafting. The density end diameter of myelinated fiber in the middle and distal part of the grafting were all greater than that of MDSCs and FK-506 groups, and there were few connective tissues between microfascicles, which was close to normal level. The co-application of MDSCs and FK-506 enhances peripheral nerve regeneration and functional recovery in acelluler nerve allograft graft. Therefore, MDSCs and FK-506 are synergistic factors in peripheral nerve injury repair.
ABSTRACT
Autogenous nerve transplantation is the best means for treating peripheral nerve injury. However, it would detriments function of donor area. Therefore, homologous nerve transplantation is adopted in clinic. The key of successful nerve allograft transplantation is to reduce nerve allograft antigenicity. Compared to physics method, chemistry method achieved a better result in reducing antigenicity of allograft nerve. Here, we introduce immune response and characteristics of grafts following homologous nerve transplantation, discuss the pretreatment method which can reduce the antigenicity. And propose that the combination of one or many kinds of methods would decrease the antigenicity of homogenous nerve in future.
ABSTRACT
Objective To introduce operative skill of liver resection by blocking hilum of liver under normal temperature.Method Retrospective analysis of experience treating 86 cases of liver cancer with the operation.Result Among 86 cases of hepatic malignant tumor,there were not serious complications such as death,massive haemorrhage etc.during operation;postoperative liver failure occurred in 7 cases,among them,cure 5 cases and 2 cases of death.The incision infection occurred in 5 patients;The thorax dropsy occurred in 19 cases, which was absorbed by oneself after treating in 14 cases,cured by puncture occurred in 5 cases.Conclusion Operative skill of blocking hilum of liver under normal temperature is more ideal means in treating the liver cancer.