Gene therapy of hypoparathyroidism with TheraCyte-encapsulated stem cells

Abstract

The parathyroid hormone (PTH) (1-34) gene was inserted into a pcDNA3 promoter and E. coli competent cells were used to amplify the cDNA. C3H/10T1/2 stem cells were transfected with PTH (1-34) cDNA using Lipofectamine reagents. After G418 treatment live cells at a density of 4x107 were loaded onto a TheraCyte unit. After parathyroidectomy, rats were either the implanted with 4x107 TheraCyte-encapsulated cells (group A), subcutaneously injected with 4x107 live cells containing PTH (1-34) cDNA (group B) or injected with nothing (group C).Serum levels of calcium, phosphorus and PTH (1-34) were measured at baseline, 1 month, 2 months, 3 months and 4 months after therapy. Immunohistochemical staining and RT-PCR were performed to find PTH (1-34)-positive cells and to detect PTH (1-34) mRNA.Serum calcium and PTH (1-34) levels were significantly higher in group A than in group B or C. PTH (1-34)-positive cells were found in the TheraCyte group 4 months after implantation. PTH (1-34) mRNA was detected in stem cells 48 hr after transfection and also in stem cells after transfection and 72 hr after G418 treatment.Implantation of the TheraCyte-encapsulated stem cells, which were tranfected with PTH (1-34) cDNA can treat hypoparathyroidism.