Introducing residents to a career in management: the Physician Management Pathway.

To prepare their students and residents for the practice of medicine in a rapidly changing environment, medical educators must teach the business of medicine as well as the science and art of medicine. Recognizing the value of and demand for physician leaders, the authors sought to introduce residents to diverse aspects of medical administration within the context of an academically rigorous internal medicine residency program. The Physician Management Pathway (PMP) was developed in 1997 to expose interested residents to the management concepts employed by physician managers; to help physician trainees begin to develop the leadership and technical skills they will use as physician managers; to provide career mentoring; to provide experiences that enrich the resident's training in clinical medicine; to enhance resident-directed educational activities; and to produce physicians who can successfully work in any health care environment. The PMP curriculum consists of a monthly seminar series, a preceptorship experience in the second year of residency, and a supervised project in the third year. The program was not designed to provide competency in management, but rather is intended to provide an opportunity for new physicians to explore options in this exciting and changing profession.

Reperfusion injury pathophysiology in sickle transgenic mice.

Reperfusion of tissues after interruption of their vascular supply causes free-radical generation that leads to tissue damage, a scenario referred to as "reperfusion injury." Because sickle disease involves repeated transient ischemic episodes, we sought evidence for excessive free-radical generation in sickle transgenic mice. Compared with normal mice, sickle mice at ambient air had a higher ethane excretion (marker of lipid peroxidation) and greater conversion of salicylic acid to 2,3-dihydroxybenzoic acid (marker of hydroxyl radical generation). During hypoxia (11% O(2)), only sickle mice converted tissue xanthine dehydrogenase to oxidase. Only the sickle mice exhibited a further increase in ethane excretion during restitution of normal oxygen tension after 2 hours of hypoxia. Only the sickle mice showed abnormal activation of nuclear factor-kappaB after exposure to hypoxia-reoxygenation. Allopurinol, a potential therapeutic agent, decreased ethane excretion in the sickle mice. Thus, sickle transgenic mice exhibit biochemical footprints consistent with excessive free-radical generation even at ambient air and following a transient induction of enhanced sickling. We suggest that reperfusion injury physiology may contribute to the evolution of the chronic organ damage characteristic of sickle cell disease. If so, novel therapeutic approaches might be of value.

Acute renal failure: controversies, clinical trials, and future directions.

Acute renal failure (ARF) is an important clinical syndrome. Despite the frequent occurrence of ARF, nephrologists have not made major therapeutic inroads in the treatment or prevention of ARF. This article will speculate as to why it has been so difficult to gain a substantial foothold in the ongoing battle against ARF. First, some of the major controversies regarding the pathogenesis of ARF will be considered. Scientific debates regarding the mechanisms of ARF have greatly enriched the scientific literature, but may have slowed the development of clinically applicable therapies. Controversies regarding the treatment of ARF will then be discussed. Next, the fate of several recent clinical trials in ARF will be examined. Finally, the future directions that research in ARF may pursue will be contemplated.

Nitric oxide-mediated renal epithelial cell injury during hypoxia and reoxygenation.

The potent endothelial-derived vasodilator nitric oxide (NO) has been identified as a protective agent in acute renal failure. However, some recent studies have suggested a detrimental effect of NO on rat proximal tubules exposed to hypoxia and reoxygenation. We determined whether NO metabolites cause intracellular oxidation during hypoxia and reoxygenation and whether this oxidative stress is linked to irreversible cell injury. Primary cultures of rat proximal tubular epithelial cells were studied in a subconfluent stage and subjected to 60 min hypoxia and 30 min reoxygenation. Intracellular oxidation was assessed by monitoring the conversion of nonfluorescent dihydrorhodamine 123 (DHR) to fluorescent rhodamine 123 as a probe for the long-lived oxidant peroxynitrite. Hypoxia and reoxygenation produced a marked increase in cellular generation of oxidant species. Intracellular oxidation of DHR was reduced by approximately 40% when cells were also exposed to the NO synthase inhibitor L-NAME. Oxidation of DHR following hypoxia and reoxygenation was not affected by SOD or DMTU. A combination of SOD and L-NAME was no more effective than L-NAME alone. Hypoxia and reoxygenation produced substantial injury (as LDH release). There was a 40% reduction in LDH release when cells were pretreated with a NO synthase inhibitor. In summary, increased generation of NO capable of inducing intracellular oxidizing reactions and cell death occurred during renal hypoxia and reoxygenation.

Purinergic receptors mediate cell proliferation and enhanced recovery from renal ischemia by adenosine triphosphate.

Kidney dysfunction after ischemia can be improved by either limiting the initial injury or by enhancing the subsequent proliferative repair process. Adenosine triphosphate (ATP) favorably affects kidney function when it is given shortly after ischemia. We tested whether ATP promotes the proliferative repair response. Rats were subjected to occlusion of the left renal artery for 40 minutes and received an infusion of ATP, 12.5 micromol intravenously over 30 minutes, beginning at reperfusion. Control animals received saline solution or the hydroxyl radical scavenger dimethylthiourea (DMTU). Despite comparable functional protection by DMTU and ATP, only ATP specifically increased DNA synthesis (renal incorporation of tritiated thymidine) to an extent greater than that produced by ischemia alone. In other animals, ribonucleic acid was extracted from kidneys for Northern analysis. Expression of the proto-oncogenes c-fos and c-jun was enhanced in ATP-treated animals as compared with controls. Expression of a histone protein gene (H2b) and thymidine kinase was increased by ischemia but was not additionally affected by ATP. In vitro studies of primary cultures of renal proximal tubule epithelial cells confirmed the ability of ATP to stimulate cellular proliferation as a consequence of stimulation of purinergic P2 receptors, possibly of the P2x subclass. In summary, ATP given after ischemia increased new DNA synthesis and augmented expression of genes critical to cellular proliferation. These beneficial effects were not merely a consequence of limiting initial cellular damage, and they suggest a novel mechanism of action for ATP and other purinergic receptor agonists in renal ischemia.

Acute renal failure in the 21st century: recommendations for management and outcomes assessment.

Acute renal failure (ARF) remains a common and potentially devastating disorder affecting as many of 5% of all hospitalized patients, with a higher prevalence in patients in critical care units. ARF is more frequently observed in the setting of multiorgan dysfunction syndrome (MODS) and in elderly patients with complex disease, where mortality is high. Numerous technical advances have not yet impacted favorably on this high mortality rate. This report summarizes recommendations from participants at the National Institutes of Health Conference: "Acute Renal Failure in the 21st Century," May 6 to 8, 1996, in Bethesda, MD. The focus is on categorizing recent clinically relevant developments in the field and on identification of new research initiatives to transfer a new body of knowledge derived from fundamental studies and laboratory investigation to the management of ARF in the new millennium. The development of a multicenter database through cooperative multicenter studies is advocated. Future studies should define the appropriate outcome measures to assess and emphasize the impact of hemodynamic monitoring, adjunctive agents, and adequacy and modality of renal replacement therapy on outcomes in ARF.

Effects of integrins on proliferation and apoptosis of renal epithelial cells after acute injury.

We sought to determine the importance of integrins for recovery after acute tubular injury and to investigate the signal transduction pathways for integrin effects on cell cycle regulation involving proliferation and apoptosis. Primary cultures of rat renal proximal tubule epithelial cells were exposed to a superoxide-generating system to induce injury in the absence of overt necrosis. Integrin function was antagonized by the integrin recognition sequence tetrapeptide Gly-Arg-Gly-Asp (GRGD) or monoclonal antibody to beta 1-integrin. Injured cells had reduced thymidine uptake compared with normal cells. The presence of GRGD during recovery from injury caused a further 44% reduction in DNA synthesis but did not affect DNA synthesis in normal cells. Injured cells had an increased proportion of apoptosis that was further accentuated by exposed to GRGD during recovery. Integrin antagonism also stimulated apoptosis in uninjured cells. To investigate signal transduction mechanisms for this effect of integrins, inhibitors and activators of protein tyrosine kinase (PTK) and protein kinase C (PKC) were evaluated. Activation of PKC stimulated cellular proliferation, whereas inhibitors of PKC and PTK had no significant effect. Genistein, a PTK inhibitor, induced apoptosis in normal cells, mimicking the effect of integrin inhibition. On the other hand, PMA, an activator of PKC, prevented cells from becoming apoptotic when exposed to injury plus GRGD. The phosphorylation status of intracellular proteins was evaluated by immunoblotting with antiphosphotyrosine antibody. A similar pattern of decreased phosphorylation was observed after either integrin inhibition, injury, both, or PTK inhibition. These findings suggest that kinase cascades are involved in the effects of integrins on renal epithelial cell proliferation and apoptosis. After injury, an interaction between cells and the extracellular matrix is required for cells to proliferate and contribute to repair rather than to enter an apoptotic pathway.

Glomerular disease and lung transplantation.

Three patients with lung or heart/lung transplants developed nephrotic-range proteinuria 2 to 5 years posttransplantation. Kidney biopsy showed focal segmental glomerulosclerosis in two patients and probable focal sclerosis in the third. A retrospective review of postmortem kidney specimens from 18 lung transplant recipients who died did not indicate additional cases of glomerular disease. The three patients with glomerular disease after lung transplantation had very few clinical similarities other than nephrotic-range proteinuria and lung transplantation. Their underlying lung diseases were different, and their posttransplantation courses were very different in terms of pulmonary function, cyclosporine nephrotoxicity, and other complications. We did not find in the literature previous reports of de novo focal segmental glomerulosclerosis or other glomerular lesions after lung transplantation. We suspect that additional cases will be identified in the future.

Hazards of antioxidant combinations containing superoxide dismutase.

Oxygen free radical scavengers protect against ischemia/reperfusion injury of the kidney in vivo and against hypoxia/reoxygenation (H/R) injury of renal cells in several in vitro systems. In an attempt to maximize renal protection we tested several antioxidants in combination; the individual components had previously reduced reoxygenation injury of hypoxic renal epithelial cells. Both glutathione (GSH; 1 mM) and Cu,Zn-SOD provided significant protection against posthypoxic injury. Surprisingly, the combination of Cu,Zn-SOD plus GSH eliminated protection entirely and was highly toxic to normoxic cells. The toxicity of Cu,Zn-SOD+GSH was not prevented by the iron chelator deferoxamine and was only slightly reduced by the hydroxyl scavenger DMTU. Catalase reversed the toxicity of Cu,Zn-SOD+GSH and provided net protection. Direct measurement of intracellular peroxides using 2,7-dichlorofluorescein quantitated by laser cytometry also revealed enhanced generation of peroxides by cells during H/R when Cu,Zn-SOD+GSH was present. GSSG was less toxic than GSH when combined with Cu,Zn-SOD. Importantly, the combination of Mn-SOD+GSH provided superior protection to either agent alone. In the presence of added GSH, heated or autoclaved Cu,Zn-SOD was still toxic, whereas SOD free of chelatable Cu++ was benign. In the presence of GSH, Cu++ derived from SOD may promote the formation of toxic thionyl radicals, metal-centered radicals, and/or H2O2, thereby causing cell injury. Great care should be used in designing and interpreting studies employing combinations of antioxidants.

Extracellular iron chelators protect kidney cells from hypoxia/reoxygenation.

Iron is an important contributor to reoxygenation injury because of its ability to promote hydroxyl radical formation. In previous in vivo studies, we demonstrated that iron chelators that underwent glomerular filtration provided significant protection against postischemic renal injury. An in vitro system was employed to further characterize the protection provided by extracellular iron chelators. Primary cultures of rat proximal tubular epithelial cells were subjected to 60 min hypoxia and 30 min reoxygenation (H/R). During H/R, there was a 67% increase in ferrozine-detectable iron in cell homogenates and increased release of iron into the extracellular space. Cells pretreated with either deferoxamine (DFO) or hydroxyethyl starch-conjugated deferoxamine (HES-DFO), an iron chelator predicted to be confined to the extracellular space, were greatly protected against lethal cell injury. To further localize the site of action of DFO and HES-DFO, tracer quantities of 59Fe were added to DFO or HES-DFO, and their distribution after 2 h was quantitated. Less than 0.1% of DFO entered the cells, whereas essentially none of the HES-DFO was cell-associated. These findings suggest that iron was released during hypoxia/reoxygenation and caused lethal cell injury. Iron chelators confined to the extracellular space provided substantial protection against injury.

Lateral mobility of Na,K-ATPase and membrane lipids in renal cells. Importance of cytoskeletal integrity.

Because membrane fluidity is an important determinant of membrane function, the lateral diffusion rate (DL) of the membrane protein Na,K-ATPase was determined in intact renal proximal tubule epithelial cells by the technique of fluorescence redistribution after photobleaching (FRAP). In normal cells the DL of Na,K-ATPase in the basal membrane was 3.31 x 10(-10) cm2/sec. Treatment with cytochalasin D to promote actin filament depolymerization caused a sevenfold increase in DL. Exposure of cells to a Ca(2+)-free medium or to hypoxia and reoxygenation, which have similar disruptive effects on the cytoskeleton, also caused increases in DL. Disruption of actin microfilament structure also increased the mobile fraction of Na,K-ATPase. Using a confocal laser microscopic technique only 14.9% of total Na,K-ATPase was observed to reside in the apical membrane domain of normal cells. Microfilament depolymerization caused this fraction to increase to 47.7%. Thus, the translocation of Na,K-ATPase from the basolateral to the apical domain induced by cytoskeletal protein dysfunction was enabled by an increased rate of lateral diffusion of Na,K-ATPase. The behavior of a variety of membrane lipids following actin depolymerization was more heterogeneous. Some lipids showed a similar increase in DL, whereas others showed very little dependence upon the cytoskeleton for lateral restraint.

Cytochrome P-450 mediates tissue-damaging hydroxyl radical formation during reoxygenation of the kidney.

Renal reperfusion injury results from oxygen radical generation. During reoxygenation of hypoxic kidney cells, xanthine oxidase produces superoxide radical, which eventuates in hydroxyl radical formation by the Fenton reaction. This reaction, catalyzed by transition metals such as iron, is particularly important because hydroxyl radical is highly reactive with a wide variety of biomolecules. We tested the hypothesis that this catalytic function is fostered by iron released from the heme moiety of cytochrome P-450. Primary cultures of rat proximal tubule epithelial cells studied in a subconfluent stage were subjected to 60 min of hypoxia and 30 min of reoxygenation. When cells were pretreated with one of three cytochrome P-450 inhibitors (piperonyl butoxide, cimetidine, or ketoconazole), lethal cell injury was attenuated. There was the expected increase in O2-. production during hypoxia/reoxygenation that cytochrome P-450 inhibitors did not prevent; on the other hand, inhibitors did prevent reoxygenation-induced hydroxyl radical formation. Analogously, the increase in catalytic iron (bleomycin-detectable iron) that accompanies hypoxia/reoxygenation did not occur in the presence of cytochrome P-450 inhibitors. In vivo studies confirmed a protective effect of cytochrome P-450 inhibition because glomerular filtration rate was better preserved in rats pretreated with cimetidine and then subjected to renal artery occlusion. In summary, several chemically distinct cytochrome P-450 inhibitors reduced iron release, and thereby, hydroxyl radical formation and reoxygenation-induced lethal cell injury, without inhibiting superoxide radical formation. We conclude that highly labile P-450 may act as an Fe-donating catalyst for Fenton reaction production of HO.-mediated reperfusion injury.