Adult ; Female ; Gases ; Humans ; Intestine, Small ; metabolism ; microbiology ; physiology ; Liver Cirrhosis ; metabolism ; microbiology ; Male ; Middle Aged

OBJECTIVETo investigate the effects of transfection of agrin gene on the recovery of muscle function after a free neurovascular muscle transfer.

METHODSThe electrical gene transfection was performed when the gracilis muscle of the SD rat was completed free neurovascular transfer. The experimental group was treated with pCS2+ -agrin, the group with plasmid pCS2+ as the negative control and the group with normal saline as the frank control. The muscle function, expression of neural agrin and the junctional nAChR number was measured after the operation.

RESULTSAt 4, 5 and 10 weeks postoperatively, the pCS2+ -agrin group was significantly better than the control groups in muscle function (P < 0.05 ). The immunohistochemical staining showed an increasing deposition of the agrin protein near the endplate at 1 and 5 weeks after the operation, but decreasing remarkably to the level of control groups at 10 weeks postoperatively. The pCS2+ -agrin group was significantly more than the control groups in junctional nAChR number at every points of the time postoperatively.

CONCLUSIONSTransfection of agrin gene in the transferred muscle may increase the early recovery of muscle function.

Agrin ; genetics ; Animals ; Female ; Genetic Therapy ; Genetic Vectors ; Muscle Proteins ; genetics ; Muscle, Skeletal ; transplantation ; Rats ; Rats, Sprague-Dawley ; Recovery of Function ; Transfection

OBJECTIVETo develop a novel single nucleotide polymorphism (SNP)-PCR based method for quantitative detection of chimerism after allogeneic haemopoietic stem cell transplantation (allo-HSCT), and to explore its feasibility, accuracy and superiority.

METHODS18 SNP loci were sereened to identify informative markers for detecting chimerism in each donor/recipient pair before transplantation. Then the chimerism rate of each informative marker was analyzed by real-time quantitative PCR (RQ-PCR). The accuracy and sensitivity were verified by multiple proportion dilution and analogy chimerism compared with quantitative detection of short tandem repeat (STR)-PCR, fluorescence in situ hybridization (FISH) and fusion gene.

RESULTS(1) The average slope of the 17 time amplications of the internal control plasmid was -3.39, the average intercept was 39.97, correlation coefficients were more than 0.995, which was close to the theoretical level. The intra- and interassay variability was 0.50% and 1.1%, respectively, which were both in the allowed ranges. A linear correlation with artificial mixed chimerism is above 0.99 and a sensitivity of 0.01% proved reproducible. (2) At least one informative marker could be found in over 95% of 40 donor/recipient pairs. The results of the chimerisms derived from SNP-PCR were consistent with that from STR-PCR (96.7%), FISH and fusion gene analasis (P > 0.05); the quantitative results of special fusion gene transcripts were negtive in complete chimerism samples, and positive in mixed chimerism samples.

CONCLUSIONSThis new assay which overcome the PCR competition and plateau biases of STR-PCR provides an accurate, reliable and rapid quantitative assessment of mixed chimerism after allo-transplantation. It is highly promising for of clinical application and may take the place of STR-PCR in the conventional chimerisim assessment.

Chimerism ; Hematopoietic Stem Cell Transplantation ; Humans ; In Situ Hybridization, Fluorescence ; Polymorphism, Single Nucleotide ; Stem Cell Transplantation ; Transplantation Chimera ; Transplantation, Homologous