Use of Nonautologous Microencapsulated Fibroblasts in Growth Hormone Gene Therapy to Improve Growth of Midget Swine

ABSTRACTThe aim of the present study was to investigate the expression activity, both in vitroand in vivo, of the porcine growth hormone complementary DNA (pGH cDNA) in porcine fetal fibroblast (PFF) cells. The pGH gene had been constructed inside the bicistronic retroviral vector PSN and subsequently transfected into PFF cells further encapsulated with immunoprotective microcapsules. This would provide a way to evaluate the improvement in growth performance of Tao-Yuan swine by the use of nonautologous microencapsulated fibroblasts carrying the pGH cDNA via the technique of somatic gene therapy. Results from Southern blot analysis confirmed that the full length of the pGH cDNA was completely integrated into the genome of the PFF cells after they had been infected one to four times using a PSN retroviral vector. Moreover, Northern blot analysis showed that high transcription activity was present in clones infected twice, and exogenous pGH secretion was found when the pGH-infected PFF had been further cultured for 48 hr in vitroand subjected to immunoblot assay. Encapsulation of the pGHPFF with an alginatepoly-l-lysinealginate membrane did not show any deterioration in their proliferation and survival both in vitroand in vivo. The pGH gene in encapsulated recombinant fibroblasts was fully expressed after it had been transplanted into the peritoneal cavity of the Tao-Yuan swine, and reverse transcription-polymerase chain reaction (RT-PCR) analysis was performed on the microcapsules retrieved 1 month later. The feasibility of pGH gene therapy to improve midget Tao-Yuan swine growth enhancement is further supported by the fact that transplantation of the encapsulated recombinant fibroblast cells resulted in a much more significant increase in weight gain than in those swine in either the age-matched untreated control group or in those that had been transplanted with uncapsulated recombinant PFF cells (10.56 1.01 kg versus 6.95 0.94 and 5.27 1.30 kg; p< 0.05). These experimental data suggest that growth hormone gene therapy did provide an alternative approach for growth improvement in midget Tao-Yuan swine.