Prokineticin-1 modulates proliferation and differentiation of enteric neural crest cells.

Prokineticins (Prok-1 and Prok-2) belong to a newly identified AVIT protein family. They are involved in variety of activities in various tissues, including smooth muscle contraction of the gastrointestinal tract and promoting proliferation of endothelial cells derived from adrenal gland. Importantly, they also act as the survival factors to modulate growth and survival of neurons and hematopoietic stem cells. In this study we demonstrated that Prok-1 (but not Prok-2) protein is expressed in the mucosa and mesenchyme of the mouse embryonic gut during enteric nervous system development. Its receptor, PK-R1 is expressed in the enteric neural crest cells (NCCs). To elucidate the physiological role(s) of Prok-1 in NCCs, we isolated the NCCs from the mouse embryonic gut (E11.5) and cultured them in the form of neurospheres. In an in vitro NCC culture, Prok-1 was able to activate both Akt and MAPK pathways and induce the proliferation and differentiation (but not migration) of NCCs via PK-R1. Knock-down of PK-R1 using siRNA resulted in a complete abolishment of Prok-1 induced proliferation. Taken together, it is the first report demonstrating that Prok-1 acts as a gut mucosa/mesenchyme-derived factor and maintains proliferation and differentiation of enteric NCCs.

Mammary gland-specific secretion of biologically active immunosuppressive agent cytotoxic-T-lymphocyte antigen 4 human immunoglobulin fusion protein (CTLA4Ig) in milk by transgenesis.

A major challenge in the field of transplantation is to prevent graft rejection and prolong graft survival. Tolerance induction is a promising way to achieve long-term graft survival without the need for potent immunosuppression and its associated side effects. The recent success of co-stimulatory blockade by the chimeric protein CTLA4Ig in the modulation of the recipient's immune system and the prolongation of graft survival in animal models suggests a possible application of CTLA4Ig in clinical transplantation. To produce sufficient amounts of CTLA4Ig for future clinical application, we sought to use the mammary gland as a bioreactor and produce CTLA4Ig in the milk of transgenic farm animals. Prior to the generation of transgenic farm animals, we tested our strategy in mice. Using the promoter of the sheep beta-lactoglobulin gene, we expressed our CTLA4Ig chimeric gene in the mammary gland of transgenic mice. The yield of CTLA4Ig was fivefold higher in transgenic milk than that from transfected cells. Purified milk-derived CTLA4Ig is biologically active and suppresses T cell activation. We showed that the production of CTLA4Ig in the milk has no adverse immunosuppression effect on the transgenic animals and the offsprings that were fed with the transgenic milk. The findings suggest that the approach to produce CTLA4Ig in milk by transgenesis is feasible; further studies involving farm animals are warranted.

Association of the CD134/CD134L costimulatory pathway with acute rejection of small bowel allograft.

BACKGROUND: The CD134/CD134L interaction provides a strong costimulatory signal that is CD28-independent. The CD134/CD134L pathway has been studied in inflammatory, autoimmune diseases, and graft-versus-host disease, but no information exists on the involvement of CD134/CD134L interactions in solid organ transplantation. METHODS: CD134/CD134L costimulation was studied in a rat model of small bowel transplantation. Immunohistochemistry and flow cytometry were used to analyze the expression and localization of CD134/CD134L. Mixed lymphocyte culture and quantitative RT-PCR were used to measure the effect on T proliferation and T helper cell cytokine transcripts of single or combined CD134 and CD28 costimulatory blockade. RESULTS: CD134 and CD134L molecules were strongly expressed in acutely rejected small bowel allografts. This expression was significantly suppressed by FK506. Interruption of the CD134 and CD28 costimulatory pathways resulted in an pronounced increase in expression of interleukin-10 transcripts. CONCLUSION: These results present the first evidence that the CD134/CD134L interaction is associated with acute allograft rejection. Combined CD134/CD134L blockade may be an effective treatment to both prevent acute allograft rejection and promote the induction of cells with regulatory capacity.