Isolation of viable rat ventral prostate epithelial and nonepithelial cells

A procedure is described for the dispersion, partial purification, and identification of epithelial and nonepithelial cells from the rat ventral prostate, the latter based in part upon the ability of cells to bind androgen. Since initial efforts to mechanically disrupt prostatic tissue at 2 C lysed many cells, minced rat prostates were exposed to collagenase at 37 C and further dispersed by repetitive pipetting and passage through a tissue sieve. Washed cells in culture medium were centrifuged through an isokinetic Ficoll gradient from which three visible fraction (1, 2, and 4) and a less definite fraction 3 were harvested. Characterically, fraction 1 contained cellular debris; fraction 2 contained round nucleated cells, fewer elliptically shaped cells, red blood cells, and rare free cell nuclei; fraction 3 contained somewhat larger elliptical and round cells; and fraction 4 contained larger round and elliptically shaped cells. These were epithelial cells, as judged by electron microscopy. Isolated prostate cells from rats castrated for 24 h were incubated with [3H]testosterone; 80-90% of the retained radioactivity, the majority of which was dihydrotestosterone, was associated with washed cells from fraction 4. Similar results were obtained after in vivo administration of labeled androgen and subsequent analysis of radioactivity in cells from these fractions. Histochemically and enzymatically demonstrable formalin-insensitive acid phosphatase was increased in bands 3 and 4, which were enriched in epithelial cells. Many cells in fractions 2-4 were viable before and after exposure to Ficoll, as estimated by their ability to exclude trypan blue, incorporate radioactive uridine into RNA, generate cell monolayers during 1-4 weeks of culture, and actively metabolize androgens. Compared to fraction 4, cells from fraction 2, considered to be enriched in nonepithelial cells, actively metabolized but bound much less [3H]testosterone and its metabolites. A number of epithelial cells in fraction 4 isolated from prostates dissociated at 2 C were associated with typical-C-type RNA viruses.     

Proinsulin and C-peptide in diabetes.

The availability of C-peptide measurement continues to provide new and useful information about the state of beta cell secretory function and the natural history of diabetes. Measurement of proinsulin is of value in the diagnosis of insulin-secreting tumors.     

Familial hyperproinsulinemia. Two cohorts secreting indistinguishable type II intermediates of proinsulin conversion.

Familial hyperproinsulinemia, a hereditary syndrome in which individuals secrete high amounts of 9,000-mol wt proinsulin-like material, has been identified in two unrelated cohorts. Separate analysis of the material from each of the two cohorts had suggested that the proinsulin-like peptide was a conversion intermediate in which the C-peptide remained attached to the insulin B-chain in one case, whereas it was a conversion intermediate in which the C-peptide remained attached to the insulin A-chain in the other. To reinvestigate this apparent discrepancy, we have now used chemical, biochemical, immunochemical, and physical techniques to compare in parallel the structures of the immunoaffinity chromatography-purified, proinsulin-like peptides isolated from the serum of members of both families. Our results show that affected individuals in both cohorts secrete two-chained intermediates of proinsulin conversion in which the COOH-terminus of the C-peptide is extended by the insulin A-chain and from which the insulin B-chain is released by oxidative sulfitolysis. Analysis of the conversion intermediates by reverse-phase high-performance liquid chromatography using two different buffer systems showed that the proinsulin-related peptides from both families elute at a single position very near that of the normal intermediate des-Arg31, Arg32-proinsulin. Further, treatment of these peptides with acetic anhydride prevented trypsin-catalyzed cleavage of the C-peptide from the insulin A-chain, a result demonstrating the presence of Lys64 and the absence of Arg65 in both abnormal forms. We conclude that individuals from both cohorts with familial hyperproinsulinemia secret very similar or identical intermediates of proinsulin conversion in which the C-peptide remains attached to the insulin A chain and in which Arg65 has been replaced by another amino acid residue.     

Novel everting urologic access sheath: decreased axial forces during insertion

BACKGROUND: Advancement of urologic instruments through the genitourinary tract is associated with significant axial forces that likely contribute to patient discomfort, even after injection of a local anesthetic, and may lead to mucosal trauma, postprocedural dysuria and hematuria, and increased susceptibility to infection and strictures. Placing an everting urethral sheath prior to instrumentation may decrease these problems. Materials and METHODS: Two 7-cm-long, 5-mm diameter urethral luminal models were created, one with and one without an artificial stricture. We measured the forces generated during advancement of a novel everting access sheath (Cystoglide; Percutaneous Systems, Mountain View, CA) through the models in comparison with a representative cystoscope and a urologic dilator simulating a traditional access sheath. RESULTS: The mean force generated during advancement of the everting sheath was significantly less than that of both the representative cystoscope (P<0.01) and the traditional access sheath (P<0.01). This held true for the urethral models both with and without an artificial stricture (P<0.01) and with and without lubrication (P<0.01). CONCLUSIONS: This novel introduction sheath markedly decreased the axial forces applied to an artificial urethral luminal wall. It is possible that the clinical use of this technology will decrease the discomfort and potential complications associated with lower urinary-tract endoscopy.     

Rekindling student interest in generalist careers

Despite changes in the structure of the U.S. health care system, patients continue to need and seek out generalist physicians. However, the proportion of U.S. graduates of medical schools who choose to enter generalist residency training decreased from 50% in 1998 to less than 40% in the 2004 match. Unless we act now to reverse this trend, we may face a shortage of primary care physicians to care for the complex medical needs of an aging population. This article reviews the history of and trends in career choice and proposes 4 evidence-based recommendations to rekindle student interest in generalist careers: 1) We must improve satisfaction and enthusiasm among generalist physician role models. 2) Schools of medicine should redouble their efforts to produce primary care physicians. 3) We must facilitate the pathway from medical school to generalist residency. 4) The U.S. government should increase funding for primary care research and research training. In the absence of a major overhaul of economic incentives in favor of generalist careers, we will need to work at these multiple levels to restore balance to the generalist physician workforce and align with the desires and expectations of patients for continuing healing relationships with generalist physicians.