In vivo isolated kidney perfusion with tumour necrosis factor alpha (TNF-alpha) in tumour-bearing rats. (33/37740)

Isolated perfusion of the extremities with high-dose tumour necrosis factor alpha (TNF-alpha) plus melphalan leads to dramatic tumour response in patients with irresectable soft tissue sarcoma or multiple melanoma in transit metastases. We developed in vivo isolated organ perfusion models to determine whether similar tumour responses in solid organ tumours can be obtained with this regimen. Here, we describe the technique of isolated kidney perfusion. We studied the feasibility of a perfusion with TNF-alpha and assessed its anti-tumour effects in tumour models differing in tumour vasculature. The maximal tolerated dose (MTD) proved to be only 1 microg TNF-alpha. Higher doses appeared to induce renal failure and a secondary cytokine release with fatal respiratory and septic shock-like symptoms. In vitro, the combination of TNF-alpha and melphalan did not result in a synergistic growth-inhibiting effect on CC 531 colon adenocarcinoma cells, whereas an additive effect was observed on osteosarcoma ROS-1 cells. In vivo isolated kidney perfusion, with TNF-alpha alone or in combination with melphalan, did not result in a significant anti-tumour response in either tumour model in a subrenal capsule assay. We conclude that, because of the susceptibility of the kidney to perfusion with TNF-alpha, the minimal threshold concentration of TNF-alpha to exert its anti-tumour effects was not reached. The applicability of TNF-alpha in isolated kidney perfusion for human tumours seems, therefore, questionable.  (+info)

O-raffinose cross-linking markedly reduces systemic and renal vasoconstrictor effects of unmodified human hemoglobin. (34/37740)

The hemodynamic effects of a 20% exchange-transfusion with different solutions of highly purified human hemoglobin A-zero (A0) were evaluated. We compared unmodified hemoglobin with hemoglobin cross-linked with O-raffinose. Unmodified hemoglobin increased systemic vascular resistance and mean arterial pressure more than the O-raffinose cross-linked hemoglobin solution (by approximately 45% and approximately 14%, respectively). Unmodified hemoglobin markedly reduced cardiac output (CO) by approximately 21%, whereas CO was unaffected by the O-raffinose cross-linked hemoglobin solution. Unmodified and O-raffinose cross-linked hemoglobin solutions increased mean arterial pressure to comparable extents ( approximately 14% and approximately 9%, respectively). Unmodified hemoglobin increased renal vascular resistance 2-fold and reduced the glomerular filtration rate by 58%. In marked contrast, the O-raffinose cross-linked hemoglobin had no deleterious effect on the glomerular filtration rate, renal blood flow, or renal vascular resistance. The extents to which unmodified and O-raffinose cross-linked hemoglobin solutions inactivated nitric oxide also were compared using three separate in vitro assays: platelet nitric oxide release, nitric oxide-stimulated platelet cGMP production, and endothelium-derived relaxing factor-mediated inhibition of platelet aggregation. Unmodified hemoglobin inactivated or oxidized nitric oxide to a greater extent than the O-raffinose cross-linked hemoglobin solutions in all three assays. In summary, O-raffinose cross-linking substantially reduced the systemic vasoconstriction and the decrease in CO induced by unmodified hemoglobin and eliminated the deleterious effects of unmodified hemoglobin on renal hemodynamics and function. We hypothesize that O-raffinose cross-linking reduces the degree of oxidation of nitric oxide and that this contributes to the reduced vasoactivity of this modified hemoglobin.  (+info)

Role of glutamine in human carbohydrate metabolism in kidney and other tissues. (35/37740)

Glutamine is the most abundant amino acid in the human body and is involved in more metabolic processes than any other amino acid. Until recently, the understanding of many aspects of glutamine metabolism was based on animal and in vitro data. However, recent studies using isotopic and balance techniques have greatly advanced the understanding of glutamine metabolism in humans and its role in glucose metabolism in the kidney and other tissues. There is now evidence that in postabsorptive humans, glutamine is an important glucose precursor and makes a significant contribution to the addition of new carbon to the glucose carbon pool. The importance of alanine for gluconeogenesis, viewed in terms of the addition of new carbons, is less than previously assumed. It appears that glutamine is predominantly a renal gluconeogenic substrate, whereas alanine gluconeogenesis is essentially confined to the liver. As shown recently, renal gluconeogenesis contributes 20 to 25% to whole-body glucose production. Moreover, glutamine has been shown not only to stimulate net muscle glycogen storage but also to stimulate gluconeogenesis in normal humans. Finally, in humans with type II diabetes, conversion of glutamine to glucose is increased (more so than that of alanine). The available evidence on the hormonal regulation of glutamine gluconeogenesis in kidney and liver and its alterations under pathological conditions are discussed.  (+info)

Human, rat, and mouse kidney cells express functional erythropoietin receptors. (36/37740)

BACKGROUND: Erythropoietin (EPO), secreted by fibroblast-like cells in the renal interstitium, controls erythropoiesis by regulating the survival, proliferation, and differentiation of erythroid progenitor cells. We examined whether renal cells that are exposed to EPO express EPO receptors (EPO-R) through which analogous cytokine responses might be elicited. METHODS: Normal human and rat kidney tissue and defined cell lines of human, rat, and mouse kidney were screened, using reverse transcription-polymerase chain reaction, nucleotide sequencing, ligand binding, and Western blotting, for the expression of EPO-R. EPO's effects on DNA synthesis and cell proliferation were also examined. RESULTS: EPO-R transcripts were readily detected in cortex, medulla, and papilla of human and rat kidney, in mesangial (human, rat), proximal tubular (human, mouse), and medullary collecting duct cells (human). Nucleotide sequences of EPO-R cDNAs from renal cells were identical to those of erythroid precursor cells. Specific 125I-EPO binding revealed a single class of high- to intermediate-affinity EPO-Rs in each tested cell line (kD 96 pm to 1. 4 nm; Bmax 0.3 to 7.0 fmol/mg protein). Western blots of murine proximal tubular cell membranes revealed an EPO-R protein of approximately 68 kDa. EPO stimulated DNA synthesis and cell proliferation dose dependently. CONCLUSION: This is the first direct demonstration, to our knowledge, that renal cells possess EPO-Rs through which EPO stimulates mitogenesis. This suggests currently unrecognized cytokine functions for EPO in the kidney, which may prove beneficial in the repair of an injured kidney while being potentially detrimental in renal malignancies.  (+info)

The prolactin gene is expressed in the mouse kidney. (37/37740)

BACKGROUND: Prolactin (PRL), originally identified as an anterior pituitary hormone exhibiting lactogenic activity, is now recognized as a versatile hormone expressed in a wide variety of tissues. METHODS: In this study, the expression of PRL in the mouse kidney was investigated by solution-phase and in situ reverse transcription-polymerase chain reaction (RT-PCR) methods and immunohistochemistry. RESULTS: Mouse PRL (mPRL) transcript and protein are localized in the parietal epithelial cells of Bowman's capsule. Pit-1 is a positive transcription factor for the expression of the PRL gene. The presence of Pit-1 transcript in the kidney was also assessed by RT-PCR methods. The localization of Pit-1 mRNA coincided well with that of PRL. Immunoreactivity to mouse PRL receptor (mPRL-R) is distributed on the luminal membrane of the proximal tubule cells and the parietal epithelial cells of Bowman's capsule. CONCLUSION: These data indicate that the parietal epithelial cells of Bowman's capsule synthesize PRL de novo and suggest that Pit-1 contributes to the transcriptional regulation of PRL gene expression in the kidney, and PRL expressed in this tissue functions in an autocrine/paracrine fashion.  (+info)

Effect of diabetes and aminoguanidine therapy on renal advanced glycation end-product binding. (38/37740)

BACKGROUND: Advanced glycation end-products (AGEs) have been implicated in the pathogenesis of diabetic nephropathy, and aminoguanidine (AG) has been shown to decrease the accumulation of AGEs in the diabetic kidney. METHODS: This study investigates changes in AGE binding associated with diabetes in the rat kidney using in vitro and in vivo autoradiographic techniques. Male Sprague-Dawley rats were randomized into control and diabetic groups with and without AG treatment and were sacrificed after three weeks. Frozen kidney sections (20 microm) were incubated with [125I]-AGE-RNase or [125I]-AGE-BSA. To localize the AGE binding site, in vivo autoradiography was performed by injection of 15 microCi of [125I]-AGE-BSA into the abdominal aorta of the rat. RESULTS: Low-affinity binding sites specific for AGEs in the renal cortex (IC50 = 0.28 microm) were detected by in vitro autoradiography. There was a significant increase in [125I]-AGE binding in the diabetic kidney, which was prevented by AG treatment. Emulsion autoradiography revealed that binding was localized primarily to proximal tubules in the renal cortex. Renal AGE levels, as assessed by fluorescence or by radioimmunoassay, were increased after three weeks of diabetes. This increase was attenuated by AG therapy. CONCLUSIONS: AGE binding sites are present within the proximal tubules of the kidney and appear to be modulated by endogenous AGE levels. It remains to be determined if these binding sites represent receptors involved in clearance of AGEs or are linked to pathogenic pathways that lead to the development of diabetic nephropathy.  (+info)

Tyrosine kinase inhibitors and immunosuppressants perturb the myo-inositol but not the betaine cotransporter in isotonic and hypertonic MDCK cells. (39/37740)

BACKGROUND: The sodium/myo-inositol cotransporter (SMIT) and the betaine cotransporter (BGT1) are essential for the accumulation of myo-inositol and betaine, and hence cell survival in a hypertonic environment. The underlying molecular mechanism involves an increase in transcription of the SMIT and BGT1 genes through binding of a trans-acting factor to enhancer elements in the 5' flanking region of both genes, resulting in increased mRNA abundance and increased activity of the cotransporters. Current evidence regarding transcriptional and post-transcriptional regulation indicates that both cotransporters are regulated in parallel. METHODS: To investigate the signal transduction of hypertonic stress, we examined the effect of tyrosine kinase inhibitors and immunosuppressants on the hypertonicity-induced activity of the two cotransporters in Madin-Darby canine kidney (MDCK) cells. RESULTS: None of the agents studied affected BGT1 activity in isotonic or hypertonic conditions. Treatment of MDCK cells with genistein, a tyrosine kinase inhibitor, increased SMIT activity in hypertonic but not isotonic conditions. The stimulation of SMIT by genistein was accompanied by a parallel increase in mRNA abundance. In contrast, treating cells with tyrphostin A23, another tyrosine kinase inhibitor, or cyclosporine A, an immunosuppressant, inhibited SMIT activity in hypertonic cells. FK506, another immunosuppressant, increased SMIT activity, but only in isotonic conditions. CONCLUSIONS: These results provide the first evidence of divergent regulatory pathways modulating SMIT and BGT activity.  (+info)

Sodium reabsorption and distribution of Na+/K+-ATPase during postischemic injury to the renal allograft. (40/37740)

BACKGROUND: A loss of proximal tubule cell polarity is thought to activate tubuloglomerular feedback, thereby contributing to glomerular filtration rate depression in postischemic acute renal failure (ARF). METHODS: We used immunomicroscopy to evaluate the segmental distribution of Na+/K+-ATPase in tubules of recipients of cadaveric renal allografts. Fractional excretion (FE) of sodium and lithium was determined simultaneously. Observations were made on two occasions: one to three hours after graft reperfusion (day 0) and again on post-transplant day 7. An inulin clearance below or above 25 ml/min on day 7 was used to divide subjects into groups with sustained (N = 15) or recovering (N = 16) ARF, respectively. RESULTS: In sustained ARF, the fractional excretion of sodium (FENa) was 40 +/- 6% and 11 +/- 5%, and the fractional excretion of lithium (FELi) was 76 +/- 5% and 70 +/- 2% on days 0 and 7, respectively. Corresponding findings in recovering ARF were 28 +/- 2% and 6 +/- 2% for the FENa and 77 +/- 4% and 55 +/- 3% (P < 0.05 vs. sustained) for FELi. Na+/K+-ATPase distribution in both groups was mainly basolateral in distal straight and convoluted tubule segments and collecting ducts. However, Na+/K+-ATPase was poorly retained in the basolateral membrane of proximal convoluted and straight tubule segments in sustained and recovering ARF on both days 0 and 7. CONCLUSIONS: We conclude that loss of proximal tubule cell polarity for Na+/K+-ATPase distribution is associated with enhanced delivery of filtered Na+ to the macula densa for seven days after allograft reperfusion. Whether an ensuing activation of tubuloglomerular feedback is an important cause of glomerular filtration rate depression in this form of ARF remains to be determined.  (+info)