[Androgen-secreting tissue construction with co-cultured rat testes somatic cells by tissue engineering technique].

OBJECTIVE: To explore the feasibility of constructing androgen-secreting tissue of a certain size and shape using co-cultured somatic cells of rat testis. METHODS: Thirty male Wistar rats were castrated. model and implanted rat model were prepared by resecting bilateral testes. The suspension of mixed testes cells was cultured to obtain various somatic cells of testes and Leydig cells were collected by differential anchorage-dependent method. These two kinds of cells were seeded onto biodegradable scaffolds of polyglycolic acid (PGA) fibers and cultured in vitro. The tissue formation of cell-scaffold constructs was observed by optical microscope and electronic microscope and the level of testosterone in the supernatant was detected regularly. After 7-day culture in vitro, the 2 kinds of cell-scaffold constructs, scaffold with purified Leydig cells or co-cultured testis somatic cells (seed cells), were implanted into the gastrocolic omentum or cavity of tunica vaginalis of the castrated rats. The implants were harvested 4, 6, 9, 12, and 24 weeks later to evaluate the tissue formation of cell-scaffold constructs in vivo. The serum testosterone level of the implanted rats was assayed to evaluate the testosterone secreting function of the regenerative tissue. RESULTS: Both the co-cultured testis somatic cells and Leydig cells had fine compatibility with the PGA fibers and adhered to the scaffolds very well. Testosterone was detected at a certain degree in the supernatant of cell-scaffold constructs, indicating the testosterone secreting function of the constructs. Two months after the implantation both kinds of cell-scaffold constructs formed testosterone secreting tissue in both gastrocolic omentum and cavity of tunica vaginalis of the implanted rats. The regenerative tissues were vascularized very well with a certain size and shape. Six weeks after implantation the serum testosterone level of the Leydig cell group was 0.60 ng/ml +/- 0.04 ng/ml, and that of the co-culture group was 0.84 ng/ml +/- 0.03 ng/ml, both significantly higher than that of the control castrated rats (0.56 ng/ml +/- 0.05 ng/ml, both P < 0.01), and the serum testosterone level of the co-cultured testes somatic cell implantation group was significantly higher than that of the Leydig cell implantation group too (P < 0.01). CONCLUSION: It is completely feasible to construct androgen-secreting tissue in vitro and in vivo using tissue engineering technique. Co-cultured testis somatic cells may serve as the better seed cells for androgen-secreting tissue engineering than purified Leydig cells in terms of the quantity and function of cells.

Temporal and spatial CGRP innervation in recombinant human bone morphogenetic protein induced spinal fusion in rabbits.

STUDY DESIGN: Animal experiment using a rabbit posterolateral intertransverse process fusion model. OBJECTIVE: To explore the temporal and spatial distribution of sensory nerve fibers expressing calcitonin-gene related peptide (CGRP) during spinal fusion induced by recombinant human bone morphogenetic protein-4 and the role of the CGRP innervation in ectopic bone formation and remodeling. SUMMARY OF BACKGROUND DATA: Sensory neuropeptide CGRP involved in local bone turnover has been evidenced but its underlying mechanism is poorly understood. Knowledge in the CGRP innervation in ectopic bone induced by bone morphogenetic proteins can help us to understand its role in bone turnover. METHODS: Twenty-seven New Zealand white rabbits underwent single level posterolateral intertransverse process fusion of the lumbar vertebrae with implantation of porous poly-d,l-lactic acid blocks loaded with 1.25 microg recombinant human bone morphogenetic protein-4 solution. Animals were killed and the operated lumbar vertebrae were harvested for histomorphological evaluation at 3 days (n = 3), 1 week (n = 6), 3 weeks (n = 6), 7 weeks (n = 6), and 12 weeks (n = 6) following surgery, respectively. RESULTS: New cartilage presented at 1 week postimplantation adjacent to the implant, reached a peak volume at week 3 followed by a drop till week 12 after its ossification. Trabeculae-like woven bone structure presented at week 3. CGRP-positive nerve fibers regenerated already at 3 days postimplantation, reached its peak density at week 3. The CGRP-positive fibers presented both in fibrous tissues adjacent to proliferating cartilages and in bone marrow of newly formed trabecular bone. CONCLUSIONS: The observed spatial and temporal regeneration of CGRP-positive nerve fibers in ectopic bone formation suggested CGRP innervation is associated with ectopic osteogenesis.