The bioethics and utility of selling kidneys for renal transplantation.

In the 53 years since kidney transplantation was first performed, this procedure has evolved from a highly speculative biomedical endeavor to a medically viable and often standard course of therapy. Long-term survival is markedly improved among patients who receive a kidney compared with patients who remain on the waiting list for such an organ. As outcomes have improved and more clinical indications have emerged, the number of people awaiting transplantation has grown significantly. In stark contrast to the robust expansion of the waiting list, the number of available deceased donors has remained relatively constant over the last several years. The current mechanism for procuring kidneys relies on voluntary donations by the general public, with the primary motivation being altruism. However, in light of the ever-increasing waiting list, it is the researchers' belief that the current system needs to be revised if supply is ever going to meet demand. In response to this critical organ shortage, different programs have been developed in an attempt to increase organ donation. At present, however, no solution to the problem has emerged. This report begins by outlining the scope of the problem and current legislation governing the procurement of transplantable organs/tissues in the United States. It continues with an overview of different proposals to increase supply. It concludes by exploring some of the controversy surrounding the proposal to increase donation using financial incentives. Though the following discussion certainly has implications for other transplantable organs, this report focuses on kidney transplantation because the waiting list for kidneys is by far the longest of all waiting lists for solid organs; and, as kidney transplant carries the smallest risk to living donors, it is the least ethically problematic.

Simvastatin protection against acute immune-mediated glomerulonephritis in mice.

In addition to cholesterol lowering, 3-hydroxy-3-nethylglutaryl coenzyme A (HMG-CoA) reductase inhibitors limit inflammatory changes associated with atherosclerosis. There is also support for their use as inhibitors of progression in chronic renal disease, irrespective of cause. In this study, their capacity to limit acute renal inflammation was evaluated. For this purpose, mice were treated with Simvastatin either prior to, at the time of, or shortly after induction of nephrotoxic nephritis. The severity of disease was determined by evaluation of blood urea nitrogen (BUN), proteinuria, and renal histologic changes. The reversibility of benefit was evaluated by the administration of mevalonic acid along with nephrotoxic serum (NTS) and Simvastatin The severity of the acute nephritis, including proteinuria, elevated BUN, and histologic changes, was ameliorated in a dose-dependent manner, when Simvastatin was administered either prior to NTS injection or at the time of NTS injection. By contrast, Simvastatin did not alter the course of established nephritis. Coadministration of mevalonic acid, the immediate substrate following HMG-CoA reductase, abolished Simvastatin's renoprotective effect, indicating that the benefit is, at least in part, due to interference with HMG-CoA reductase and biosynthetic substrates downstream from the enzyme. These findings provide the rationale for the evaluation of the efficacy of HMG-CoA reductase inhibitors in patients with recurrent forms of renal inflammation, to limit the severity of acute exacerbations of disease, prevent renal scarring and slow the rate of progression.

Cdc42 interacts with the exocyst and regulates polarized secretion.

Polarized delivery and incorporation of proteins and lipids to specific domains of the plasma membrane is fundamental to a wide range of biological processes such as neuronal synaptogenesis and epithelial cell polarization. The exocyst complex is specifically localized to sites of active exocytosis and plays essential roles in secretory vesicle targeting and docking at the plasma membrane. Sec3p, a component of the exocyst, is thought to be a spatial landmark for polarized exocytosis. In a search for proteins that regulate the localization of the exocyst in the budding yeast Saccharomyces cerevisiae, we found that certain cdc42 mutants affect the polarized localization of the exocyst proteins. In addition, we found that these mutant cells have a randomized protein secretion pattern on the cell surface. Biochemical experiments indicated that Sec3p directly interacts with Cdc42 in its GTP-bound form. Genetic studies demonstrated synthetically lethal interactions between cdc42 and several exocyst mutants. These results have revealed a role for Cdc42 in exocytosis. We propose that Cdc42 coordinates the vesicle docking machinery and the actin cytoskeleton for polarized secretion.

Localization of GFP-tagged concentrative nucleoside transporters in a renal polarized epithelial cell line.

Many nucleosides undergo active reabsorption within the kidney, probably via nucleoside transporters. To date, two concentrative nucleoside transporters have been cloned, the sodium-dependent purine-selective nucleoside transporter (SPNT) and concentrative nucleoside transporter 1 (CNT1). We report the stable expression of green fluorescence protein (GFP)-tagged SPNT and CNT1 in Madin-Darby canine kidney (MDCK) cells, a polarized renal epithelial line. We demonstrate that the GFP tag does not alter the substrate selectivity and only modestly affects the kinetic activity of the transporters. By using confocal microscopy and functional studies, both SPNT and CNT1 are localized primarily to the apical membrane of MDCK and LLC-PK(1) cells. Apical localization of these transporters suggests a role in renal nucleoside reabsorption and regulation of tubular function via the adenosine pathway.

Exocyst is involved in cystogenesis and tubulogenesis and acts by modulating synthesis and delivery of basolateral plasma membrane and secretory proteins.

Epithelial cyst and tubule formation are critical processes that involve transient, highly choreographed changes in cell polarity. Factors controlling these changes in polarity are largely unknown. One candidate factor is the highly conserved eight-member protein complex called the exocyst. We show that during tubulogenesis in an in vitro model system the exocyst relocalized along growing tubules consistent with changes in cell polarity. In yeast, the exocyst subunit Sec10p is a crucial component linking polarized exocytic vesicles with the rest of the exocyst complex and, ultimately, the plasma membrane. When the exocyst subunit human Sec10 was exogenously expressed in epithelial Madin-Darby canine kidney cells, there was a selective increase in the synthesis and delivery of apical and basolateral secretory proteins and a basolateral plasma membrane protein, but not an apical plasma membrane protein. Overexpression of human Sec10 resulted in more efficient and rapid cyst formation and increased tubule formation upon stimulation with hepatocyte growth factor. We conclude that the exocyst plays a central role in the development of epithelial cysts and tubules.

Clonality of urogenital organs as determined by analysis of chimeric mice.

Though the first mammalian chimera was reported in 1961, suitable markers for different animal strains which are easily detectable in histological sections of all or most organs have not existed. Chimeric mice were produced having an excellent histological marker, the C3H antigen, which is strain-specific and fulfills all the criteria for an ideal strain-specific histological marker. Using male and female C3H-Balb/c chimeric mice we examined epithelial cells of urogenital organs and their morphological or functional units, such as the glomerulus, to determine whether individual organs and their morphological subunits were monoclonal or polyclonal in origin. We found that the epithelial parenchyma of most male and female urogenital organs (the prostate, seminal vesicle, epididymis, ovaries, vagina, kidney, ureter and bladder) and their morphological subdivisions were derived from cells of both input strains, indicating a polyclonal origin for each organ and/or organ component. A notable exception was the uterus in which all individual uterine glands examined (n = 403) were found to be either entirely Balb/c or entirely C3H, indicating a monoclonal origin. The clonality of urogenital structures is discussed in terms of the morphogenesis of the urogenital system.

To clone or not to clone--a Jewish perspective.

Many new reproductive methods such as artificial insemination, in vitro fertilisation, freezing of human embryos, and surrogate motherhood were at first widely condemned but are now seen in Western society as not just ethically and morally acceptable, but beneficial in that they allow otherwise infertile couples to have children. The idea of human cloning was also quickly condemned but debate is now emerging. This article examines cloning from a Jewish perspective and finds evidence to support the view that there is nothing inherently wrong with the idea of human cloning. A hypothesis is also advanced suggesting that even if a body was cloned, the brain, which is the essence of humanity, would remain unique. This author suggests that the debate should be changed from "Is cloning wrong?" to "When is cloning wrong?".

Hepatocyte growth factor and the kidney: it is not just for the liver.

Mesenchymal-epithelial interactions are important for many biological processes in epithelial organs such as the kidney. Hepatocyte growth factor (HGF) is a mesenchymally derived polypeptide cytokine that acts through its tyrosine kinase c-met receptor and is an important mediator of these interactions. This article reviews data showing the in vitro actions of HGF on renal epithelial cells that result in such diverse responses as mitogenesis, motogenesis, and morphogenesis. It also examines the in vivo evidence linking HGF and the c-met receptor to kidney development, regeneration following injury, and renal disease. Elucidating cellular mechanisms underlying the coordinated control of diverse HGF-induced phenotypic changes in renal epithelia in vitro should contribute to a clearer understanding of complex biological processes such as organogenesis, regeneration, and carcinogenesis in epithelial organs such as the kidney.

Expression of beta-catenin and gamma-catenin in epithelial tumor cell lines and characterization of a unique cell line.

In addition to its structural role, beta-catenin has recently been identified as an oncogene, while its homologue gamma-catenin (plakoglobin) seems to suppress tumorigenicity. Twenty-five epithelial tumor cell lines were screened; 18 expressed both beta- and gamma-catenin, two expressed neither protein, four showed beta- but not gamma-catenin expression, while only one cell line showed gamma- but not beta-catenin expression. As per literature search, the cell line expressing gamma- but not beta-catenin appeared to be unique. This cell line, SKBR-3, is a human breast cancer cell line which does not express beta-catenin or E-cadherin protein. There is, however, expression of beta-catenin, but not E-cadherin, mRNA. In order to determine the mechanism for this unique expression pattern, SKBR-3 cells were transfected with E-cadherin which resulted in expression of beta-catenin protein. Immunofluorescent staining of the E-cadherin transfected SKBR-3 cells revealed beta-catenin in the adherens junctions while transfection with just an epitope tagged (VSV) beta-catenin showed expression only in the nucleus. Double transfection with E-cadherin and VSV beta-catenin showed the beta-catenin in the adherens junction of the E-cadherin transfected cells. These results indicate that the mechanism for the lack of beta-catenin expression in the SKBR-3 cell line is possibly post-translational degradation and that when E-cadherin is transfected into these cells, the beta-catenin is stabilized in the adherens junction and not degraded. This cell line should be of interest to those studying the role of the homologues, beta- and gamma-catenin, in cancer pathogenesis.

Molecular development of the kidney: a review of the results of gene disruption studies.

The kidney has been used for the last 50 years as a model system for the study of tissue inductions and vertebrate organogenesis. While much is known about the morphologic development of the kidney, it is only in the last few years that the molecular mechanisms involved in these processes have begun to be identified. This is largely a result of the identification of genes expressed during kidney development and the application of techniques for single gene disruption. Mammalian kidney development is described, and the methodology for single gene disruption is discussed. For a candidate gene to be unequivocally shown to be involved in organ development, three conditions are necessary. First, the gene must be spatially expressed correctly relative to the developing organ. Second, the gene has to be temporally expressed in a correct manner. Finally, when that gene is disrupted, normal organ development must not occur. There are now 11 genes that satisfy these conditions and thus have been shown to be crucial for metanephric kidney development: WT-1, Pax-2, c-ret, GDNF, alpha8beta1, Wnt-4, BF-2, BMP-7, PDGF B, PDGFRbeta, and alpha3beta1. These genes and their probable roles in kidney development are discussed, and some molecular pathways are suggested. Finally, the applications, limitations, and future trends in single gene disruption studies are discussed. Single gene disruption already has generated a wealth of information about kidney development and mammalian development in general. It is likely that this information is only the beginning, and many startling and profound discoveries can be expected in the years to come both from the utilization of knockout mice that already exist and those that will be created.

Differentiation of rat neonatal ventral prostates grown in a serum-free organ culture system.

BACKGROUND: Organ culture methods have long been used in the study of the prostate because effects of drugs and hormones can be examined in the absence of systemic effects. METHODS: Neonatal rat ventral prostates (VP) were grown on Millipore filters floating on fluid medium composed of Dulbecco's modified Eagle's medium/Ham's F-12 supplemented with insulin, transferrin, and hydrocortisone, and in the presence or absence of testosterone (T, 10(-8)M). RESULTS: In the presence of T, ductal lumen formation occurred, ductal branching was extensive, and basal and luminal epithelial cells were identified by immunocytochemistry based on their distinctive cytokeratin profile. In the absence of T, ductal lumen formation did not occur, basal and luminal epithelial cells failed to differentiate, and there was a marked decrease in prostatic organ size relative to glands grown with T. Interestingly, DNA synthesis, as measured by counts per min (CPM) for 3H-thymidine incorporation, showed that DNA synthesis per microgram DNA at 7 days of organ culture was not inhibited by lack of T. Androgen receptor expression is another marker of prostatic epithelial differentiation, and it occurred in both the presence and absence of T. CONCLUSIONS: Growth and differentiation of the neonatal rat prostate in vitro occur in a manner similar to that of the developing prostate in vivo, demonstrating that organ cultures of neonatal rat ventral prostates provide a faithful model for studying rat prostatic development and differentiation under serum-free conditions.

Keratinocyte growth factor (KGF) can replace testosterone in the ductal branching morphogenesis of the rat ventral prostate.

Prostatic growth occurs through ductal elongation and branching into the mesenchyme. Ductal branching morphogenesis in the prostate is elicited by androgens via mesenchymal-epithelial interactions mediated by paracrine influences from mesenchyme. The role of keratinocyte growth factor (KGF) was investigated in the developing prostate as KGF has been suggested to be a paracrine acting factor. KGF transcripts were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) in neonatal rat ventral prostates (VPs) in vivo, in VPs cultured in vitro, and in isolated VP mesenchyme. KGF receptor was detected in VP's by RT-PCR and was localized specifically to the epithelium by in situ hybridization. KGF was investigated as a potential paracrine mediator during androgen-induced prostatic development by examining neonatal rat VPs cultured for 6 days under serum-free conditions using a basal medium supplemented only with insulin and transferrin. When testosterone (10(-9) to 10(-8) M) was added to the basal medium, VPs grew and underwent ductal branching morphogenesis similar to that in situ. Neutralization of endogenous KGF with a monoclonal antibody to KGF (anti-KGF) or a soluble KGF receptor peptide inhibited androgen-stimulated VP growth (DNA content) and reduced the number of ductal end buds after 6 days of culture. When KGF (50 or 100 ng/ml) was added to the basal medium in the absence of testosterone, VP growth and ductal branching morphogenesis were stimulated. The number of ductal end buds was about 70% of that obtained with an optimal dose of testosterone (10(-8)M), and DNA content of VP's cultured with 100 ng/ml KGF was equivalent to that of glands cultured with testosterone. The stimulatory effect of KGF was partially blocked by cyproterone acetate, a steroidal anti-androgen. These data imply that KGF plays an important role as a mesenchymal paracrine mediator of androgen-induced epithelial growth and ductal branching morphogenesis in the rat VP.

Phenylacetate is an inhibitor of prostatic growth and development in organ culture.

PURPOSE: Benign prostatic hyperplasia (BPH) is the most common proliferative disease affecting men, and the obstructive uropathy it causes results in serious morbidity and financial cost. Phenylacetate (PA) is a small molecule that is a product of phenylalanine metabolism and is normally present in the mammalian circulation at very low levels. It has long been safely used in humans to treat the hyperammonemia resulting from urea synthesis disorders and liver failure. It has recently been investigated as an anticancer agent because it decreased growth and increased differentiation of a variety of human neoplasms, including prostate cancer in which a phase I trial has recently been completed. MATERIALS AND METHODS: Because of PA's growth-inhibitory effects on a variety of cell lines and the idea that BPH is due to a reawakening of embryonic-like inductive activity in prostatic stromal cells, which then induce development of epithelial nodules, we examined the effect of PA on serum-free organ cultures of developing rat prostates. RESULTS: We found that PA markedly decreased rat prostatic growth and ductal morphogenesis at concentrations that have previously been well tolerated in patients. In ventral prostates grown for 7 days in organ culture, histodifferentiation was inhibited as measured by a marked decrease in ductal lumen formation and ductal branching morphogenesis. This inhibition of differentiation was confirmed by using cytokeratin antibodies specific for basal and luminal cells. Synthesis of DNA was also significantly decreased per organ with PA. The growth inhibitory effects of PA were reversible, and the mechanism did not appear to be due to glutamine or glycine deprivation, or androgen receptor inhibition. CONCLUSIONS: In common with earlier studies, we found that PA inhibits prostatic growth; however, in our organ culture system, differentiation was also largely inhibited. These studies indicate that there may be a role for PA in treating BPH or in elucidating the mechanism by which it occurs since BPH apparently involves the neoformation of ductal-acinar tissue in aged men via mechanisms fundamentally similar if not identical to those in fetal prostatic development.

Urothelial transformation into functional glandular tissue in situ by instructive mesenchymal induction.

It is generally believed that adult tissue is terminally differentiated. The ureter is derived from the metanephric diverticulum which, along with the derivatives of the metanephric mesoderm, forms the kidney. In our experiments, the left ureters of adult male athymic mouse hosts were severed below the kidney, and mesenchyme from neonatal rat seminal vesicles (SVM) was grafted to the cut end of the ureter, thus bringing adult mouse ureter epithelium (URE) in contact with neonatal rat SVM. After four to eight weeks, the in situ tissue recombinants were harvested, and the epithelial secretory proteins recovered. In 5 of 11 cases, an induction occurred, resulting in an in situ transformation of the non-glandular transitional epithelium of the adult mouse ureter into the simple columnar epithelium of the seminal vesicle (SV). Functional cytodifferentiation was examined in these neonatal rat SVM + adult mouse URE tissue recombinants using antibodies against SV-specific secretory (SVS) proteins of the mouse and rat. From the cut end of the ureter, the adult URE was induced to undergo SV morphogenesis, to express SV cytodifferentiation, and to produce the complete spectrum of major SVS proteins characteristic of the mouse. The induced seminal vesicle epithelium (SVE) also expressed androgen receptors (AR) which are not seen in urothelial tissue. Staining with Hoechst dye 33258, which can distinguish cells of mouse and rat origin, further demonstrated that the induced SVE was indeed of mouse origin and not a contaminant of the inducing rat SVM. in addition, neonatal mouse vaginal mesenchyme was grafted in situ beneath the bladder mucosa of adult male mice, and the host animals were killed after three months. The vaginal mesenchyme implanted into the bladders induced prostate-like acini, indicating that the above reprogramming of adult organs in situ is not an isolate occurrence. These results set a precedent for the "recreation" of new vital organs, such as the kidney, in situ by demonstrating that adult epithelial cells retain a developmental plasticity equivalent to their undifferentiated fetal counterparts and are capable of being reprogrammed in situ to express a completely new morphological, biochemical, and functional phenotype.

Unreliability of the abdominal fat pad biopsy in the evaluation of nephrosis: report of 3 consecutive cases.

In the workup of unexplained nephrotic syndrome in the elderly patient, renal biopsy has shown amyloidosis to be the cause in 15-30% of the cases. Most of the cases of amyloidosis are primary and are, therefore, treatable with alkylating agents, albeit at a high level of toxicity. Abdominal fad pad biopsy has been suggested as a minimally invasive, low-cost method for diagnosing amyloidosis that is 100% specific. We report our experience with 3 consecutive cases of fat pad biopsy in the workup of unexplained nephrosis in the elderly patient: including the first false positive reported with respect to nephrotic renal disease, a false negative, and a true positive. We feel that in an elderly patient with unexplained nephrosis though the abdominal fat pad biopsy may be helpful, the patient should not be committed to a regimen with potentially very high toxicity on the basis of a positive fat pad biopsy alone. We recommend that the more invasive renal biopsy be performed should therapy with alkylating agents be contemplated.

Laparoscopic cholecystectomy and ERCP with sphincterotomy in an elderly patient with situs inversus.

Situs inversus viscerum (transposition of the viscera) is a rare condition with a genetic predisposition that is autosomal recessive. We present a patient with situs inversus, cholelithiasis, and choledocholithiasis who was successfully treated via laparoscopic cholecystectomy and endoscopic retrograde cholangiopancreatography with sphincterotomy. This paper further expands the application of these techniques and shows that they can be safely and effectively applied in the setting of situs inversus, although attention must be paid to the details of left-right reversal.

Bilateral anterior cerebral artery infarction resulting from explosion-type injury to the head and neck.

A 43-year-old woman suffered a blast-type injury to the head and neck. She subsequently developed bilateral internal carotid artery occlusion and bilateral anterior cerebral artery infarction not demonstrated by magnetic resonance imaging scan 24 hours after the explosion, but confirmed by a second scan 8 days after the explosion. In patients with blast-type injury to the head and neck who develop coma with a nonfocal neurological exam, the possibility of bilateral carotid artery occlusion and bilateral ischemic infarction should be considered.