The primary aim of this research proposal is to purify and characterize both biochemically and physiologicaly a prostatic growth factor activity from the rat anterior prostate (coagulating gland) and from human benign prostatic hyperplastic tissue (BPH). The principal hypothesis is that the coagulating gland and perhaps human BPH are expressing a PDGF-like growth factor important for both normal and abnormal growth and differentiation. Preliminary studies reveal mitogen activity that shares some properties of PDGF; the activity is heat and acid stable, has a MW by Sephadex G-200 gel filtration chromotography of approximatey 30,000, and shows """"""""competence"""""""" rather than """"""""progression"""""""" activity in the Balb/c 3T3 fibroblast mitogen assay system. To further characterize this activity, a sufficient quantity will be purified from rat coagulating gland using the following approach. After initial homogenization of the tissue, the material will be run on a carboxymethyl-Sepharose column followed by gel filtration on Sephacryl-200. The activity will be further purified by to heparin-Sepharose chromotography, phenyl-Sepharose chromatography and isoelectric focusing followed by purification by SDS-polyacrylamide gel electrophoresis and HPLC. Biochemical properties to be elucidated will be molecular weight, isoelectric point, stability, carbohydrate content and amino acid sequence. The hormonal dependence of the mitogen will be assessed as will its activity in vitro on primary culture of rat coagulating gland. To specifically compare with native PDGF, hybridization studies with visis oncogene probe will be performed and receptor competition experiments with (125I)PDGF will be attempted. More limited but similar studies with BPH tissue will be carried out to compare the two activities. Long range goals will be to raise an antibody to the mitogen for immunocytochemistry, for purposes of developing an RIA and to construct an affinity column to enhance purification. Other tissues will be screened for activity and in vitro experiments will be performed to study its regulation, its relationship to other known growth factors and sex steroids and its role in growth and differentiation of the normal and abnormal prostrate. As the coagulating gland is homologous embryologically to the region of the human prostate involved in BPH, identifying the nature of the growth factor activity may well provide insight into the pathophysiology of BPH.
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