Biomedical subjects

W Lieberthal

Publications and source records attributed to W Lieberthal.

At least 19 recordsLinked to original sources

PubMed2001

Anemia-induced increase in the bleeding time: implications for treatment of nonsurgical blood loss.

BACKGROUND: Preoperative bleeding time (BT) does not correlate with postoperative bleeding in patients subjected to surgical procedures. A significant positive correlation has been reported between the BT 2 hours after cardiopulmonary bypass surgery and the nonsurgical blood loss during the first 4 hours after bypass surgery. This study was done to investigate the effect of Hct and platelet count on the BT measurement in normal, healthy men and women. STUDY DESIGN AND METHODS: To assess the relative effect of RBCs and platelets on the BT, 22 healthy male and 7 healthy female volunteers were subjected to the removal of 2 units of RBCs (360 mL), followed by the return of the platelet-rich plasma (PRP) from both units and the infusion of 1000 mL of 0.9-percent NaCl. Four of the men and all seven women received their RBCs 1 hour after their removal. Shed blood levels of thromboxane B(2) (TXB(2)), 6-keto prostaglandin F(1 alpha), and peripheral venous Hct were measured. BTs were measured in 15 men and 13 women before and after a plateletpheresis procedure to collect 3.6 x 10(11) platelets per unit. RESULTS: The 2-unit RBC apheresis procedure produced a 60-percent increase in the BT associated with a 15-percent reduction in the peripheral venous Hct and a 9-percent reduction in the platelet count. The plateletpheresis procedure produced a 32-percent decrease in the platelet count, no change in peripheral venous Hct, and no change in the BT. After the removal of 2 units of RBCs, the shed blood TXB(2) level decreased significantly. Reinfusion of 2 units of RBCs restored the BT and restored the TXB(2) level to the baseline levels. CONCLUSION: The acute reduction in Hct produced a reversible platelet dysfunction manifested by an increase in BT and a decrease in the shed blood TXB(2) level at the template BT site. Return of the RBCs restored both the BT and the shed blood TXB(2) level to normal. The platelet dysfunction observed with the reduction in Hct was due in part to a reduction in shed blood TXB(2) and other, unknown mechanisms.

Adult↗

PubMed2001

Rapamycin impairs recovery from acute renal failure: role of cell-cycle arrest and apoptosis of tubular cells.

The immunosuppressive effect of rapamycin is mediated by inhibition of interleukin-2-stimulated T cell proliferation. We report for the first time that rapamycin also inhibits growth factor-induced proliferation of cultured mouse proximal tubular (MPT; IC(50) ~1 ng/ml) cells and promotes apoptosis of these cells by impairing the survival effects of the same growth factors. On the basis of these in vitro data, we tested the hypothesis that rapamycin would impair recovery of renal function after ischemic acute renal failure induced in vivo by renal artery occlusion (RAO). Rats given daily injections of rapamycin or vehicle were subjected to RAO or sham surgery. Rapamycin had no effect on the glomerular filtration rate (GFR) of sham-operated animals. In rats subjected to RAO, GFR fell to comparable levels 1 day later in vehicle- and rapamycin-treated rats (0.25 +/- 0.08 and 0.12 +/- 0.05 ml. min(-1). 300 g(-1), respectively) (P = not significant). In vehicle-treated rats subjected to RAO, GFR increased to 0.61 +/- 0.08 ml. min(-1). 300 g(-1) on day 3 (P < 0.02 vs. day 1) and then rose further to 0.99 +/- 0.09 ml. min(-1). 300 g(-1) on day 4 (P < 0.02 vs. day 3). By contrast, GFR did not improve in rapamycin-treated rats subjected to RAO over the same time period. Rapamycin also increased apoptosis of tubular cells while markedly reducing their proliferative response after RAO. Furthermore, rapamycin inhibited activation of 70-kDa S6 protein kinase (p70(S6k)) in cultured MPT cells as well as in the renal tissue of rats subjected to RAO. We conclude that rapamycin severely impairs the recovery of renal function after ischemia-reperfusion injury. This effect appears to be due to the combined effects of increased tubular cell loss (via apoptosis) and profound inhibition of the regenerative response of tubular cells. These effects are likely mediated by inhibition of p70(S6k).

Acute Kidney Injury↗

PubMed2000

Effects of hemoglobin-based oxygen-carrying solutions in anesthetized rats with acute ischemic renal failure.

The effects of three hemoglobin solutions were compared with those of iso-oncotic human serum albumin in rats with ischemic renal failure and sham-operated controls. Unmodified and alpha-alpha cross-linked hemoglobins both increase mean arterial pressure and systemic vascular resistance and reduce cardiac output substantially and to a comparable extent. In contrast, omicron-raffinose cross-linked hemoglobin has no deleterious effect on any of these parameters. In sham-operated rats unmodified hemoglobin reduces the glomerular filtration rate (GFR) by approximately 30%, whereas neither of the two cross-linked hemoglobins has any adverse effect on GFR in this group. None of the three hemoglobin solutions exacerbated the degree to which GFR was reduced by ischemia-reperfusion injury. Also, the degree of tubular necrosis induced by ischemia-reperfusion injury was also comparable in all groups. We conclude the following: (1) omicron-raffinose cross-linking, but not alpha-alpha cross-linking, ameliorates the effects of unmodified hemoglobin on vascular resistance and cardiac output; (2) both forms of cross-linking reduce the nephrotoxicity exhibited by unmodified hemoglobin in sham-operated rats; and (3) none of the hemoglobin solutions exacerbate renal injury induced by ischemia-reperfusion.

Acute Kidney Injury↗

PubMed2000

Acute renal failure. II. Experimental models of acute renal failure: imperfect but indispensable.

Acute renal failure (ARF) due to ischemic or toxic renal injury, a clinical syndrome traditionally referred to as acute tubular necrosis (ATN), is a common disease with a high overall mortality of approximately 50%. Little progress has been made since the advent of dialysis more than 30 years ago in improving this outcome. During this same period, a considerable amount of basic research has been devoted to elucidating the pathophysiology of ATN. The ultimate goal of this research is to facilitate the development of therapeutic interventions that either prevent ARF, ameliorate the severity of tubular injury following an acute ischemic or toxic renal insult, or accelerate the recovery of established ATN. This research endeavor has been highly successful in elucidating many vascular and tubular abnormalities that are likely to be involved in ischemic and toxic ARF. This information has led to impressive advances in the development of a number of different pharmacological interventions that are highly effective in ameliorating the renal dysfunction in animal models of ARF. Although these developments are exciting and promising, enthusiasm of investigators involved in this endeavor has been tempered somewhat by the results of a few recent clinical studies of patients with ATN. These trials, designed to examine the efficacy in humans of some of the interventions effective in animal models of ARF, have resulted in little or no benefit. This is therefore an important time to reevaluate the approaches we have taken over the past three to four decades to develop new and effective treatments for ATN in humans. The major goals of this review are 1) to evaluate the relevance and utility of the experimental models currently available to study ischemic and toxic renal injury, 2) to suggest novel experimental approaches and models that have the potential to provide advantages over methods currently available, 3) to discuss ways of integrating results obtained from different experimental models of acute renal injury and of evaluating the relevance of these findings to ATN in humans, and 4) to discuss the difficulties inherent in clinical studies of ATN and to suggest how studies should be best designed to overcome these problems.

Acute Kidney Injury↗

PubMed2000

Mouse proximal tubular cell-cell adhesion inhibits apoptosis by a cadherin-dependent mechanism.

Adhesion of epithelial cells to matrix is known to inhibit apoptosis. However, the role of cell-cell adhesion in mediating cell survival remains uncertain. Primary cultures of mouse proximal tubular (MPT) cells were used to examine the role of cell-cell adhesion in promoting survival. When MPT cells were deprived of both cell-matrix and cell-cell adhesion, they died by apoptosis. However, when incubated in agarose-coated culture dishes (to prevent cell-matrix adhesion) and at high cell density (to allow cell-cell interactions), MPT cells adhered to one another and remained viable. Expression of E-cadherin among suspended, aggregating cells increased with time. A His-Ala-Val (HAV)-containing peptide that inhibits homophilic E-cadherin binding prevented cell-cell aggregation and promoted apoptosis of MPT cells in suspension. By contrast, inhibition of potential beta(1)-integrin-mediated interactions between cells in suspension did not prevent either aggregation or survival of suspended cells. Aggregation of cells in suspension activated phosphatidylinositol 3-kinase (PI3K), an event that was markedly reduced by the presence of the HAV peptide. LY-294002, an inhibitor of PI3K, also inhibited survival of suspended cells. In summary, we provide novel evidence that MPT cells, when deprived of normal cell-matrix interactions, can adhere to one another in a cadherin-dependent fashion and remain viable. Survival of aggregated cells depends on activation of PI3K.

Amino Acid Sequence↗

PubMed2000

Acute renal failure. III. The role of growth factors in the process of renal regeneration and repair.

This review, which is the final installment in a series devoted to controversial issues in acute renal failure (ARF) (3, 47), will examine available information regarding the role of growth factors in ARF. In general, studies in this area have fallen into two broad categories: 1) those that have examined the renal expression of genes encoding growth factors or transcriptional factors associated with the growth response that is induced after ARF, and 2) those that have examined the efficacy of exogenously administered growth factors in accelerating recovery of renal function in experimental models of ARF. Despite the vast amount of information that has accumulated in these two areas of investigation, our understanding of the mechanisms involved in the process of regeneration and repair after ARF, and the role of growth factors in this response, remains rudimentary. This overview, contributed to by a number of experts in the field, is designed to summarize present knowledge and to highlight potentially fertile areas for future research in this area.

Acute Kidney Injury↗

PubMed1999

Plasmalogens as endogenous antioxidants: somatic cell mutants reveal the importance of the vinyl ether.

Exposure of plasmalogen-deficient variants of the murine cell line RAW 264.7 to short-term (0-100 min) treatment with electron transport inhibitors antimycin A or cyanide (chemical hypoxia) resulted in a more rapid loss of viability than in the parent strain. Results suggested that plasmalogen-deficient cells were more sensitive to reactive oxygen species (ROS) generated during chemical hypoxia; the mutants could be rescued from chemical hypoxia by using the antioxidant Trolox, an alpha-tocopherol analogue, and they were more sensitive to ROS generation by plumbagin or by rose bengal treatment coupled with irradiation. In addition, the use of buffers containing 2H2O greatly enhanced the cytotoxic effect of chemical hypoxia, suggesting the involvement of singlet oxygen. We used the unique enzymic deficiencies displayed by the mutants to differentially restore either plasmenylethanolamine (the major plasmalogen species normally found in this cell line) or its biosynthetic precursor, plasmanylethanolamine. Restoration of plasmenylethanolamine, which contains the vinyl ether, resulted in wild-type-like resistance to chemical hypoxia and ROS generators, whereas increasing levels of its precursor, which bears the saturated ether, had no effect on cell survival. These findings identify the vinyl ether double bond as a crucial element in cellular protection under these conditions and support the hypothesis that plasmalogens, through the vinyl ether, act as antioxidants to protect cells against ROS. These phospholipids might protect cells from ROS-mediated damage during events such as chemical hypoxia.

Animals↗

PubMed1999

Albumin is a major serum survival factor for renal tubular cells and macrophages through scavenging of ROS.

We have previously shown that lysophosphatidic acid (LPA), an abundant serum lipid that binds with high affinity to albumin, is a potent survival factor for mouse proximal tubular cells and peritoneal macrophages. We show here that BSA also has potent survival activity independent of bound lipids. Delipidated BSA (dBSA) protected cells from apoptosis induced by FCS withdrawal at concentrations as low as 1% of that in FCS. dBSA did not activate phosphatidylinositol 3-kinase, implying that its survival activity occurs via a mechanism distinct from that for most cytokines. On the basis of the following evidence, we propose that dBSA inhibits apoptosis by scavenging reactive oxygen species (ROS): 1) FCS withdrawal leads to ROS accumulation that is inhibitable by dBSA; 2) during protection from apoptosis, sulfhydryl and hydroxyl groups of dBSA are oxidized; and 3) chemical blockage of free sulfhydryl groups or preoxidation of dBSA with H(2)O(2) removes its survival activity. Moreover, dBSA confers almost complete protection from cell death in a well-established model of oxidative injury (xanthine/xanthine oxidase). These results implicate albumin as a major serum survival factor. Inhibition of apoptosis by albumin occurs through at least two distinct mechanisms: carriage of LPA and scavenging of ROS.

Animals↗

PubMed1999

ATP depletion increases tyrosine phosphorylation of beta-catenin and plakoglobin in renal tubular cells.

This study examines the hypothesis that the loss of integrity of the junctional complex induced by ATP depletion is related to alterations in tyrosine phosphorylation of the adherens junction proteins beta-catenin and plakoglobin. ATP depletion of cultured mouse proximal tubular (MPT) cells induces a marked increase in tyrosine phosphorylation of both beta-catenin and plakoglobin. The tyrosine phosphatase inhibitor vanadate has the same effect in ATP-replete (control) monolayers, whereas genistein, a tyrosine kinase inhibitor, reduces phosphorylation of both proteins in ATP-replete monolayers and prevents the hyperphosphorylation of these proteins with ATP depletion. This study also demonstrates that the fall in the transepithelial resistance of MPT monolayers induced by ATP depletion can be reproduced by treatment of ATP-replete monolayers with vanadate, whereas genistein substantially ameliorates the fall in transepithelial resistance induced by ATP depletion. Also, using immunofluorescence microscopy it was demonstrated that ATP depletion results in a marked diminution of E-cadherin staining in the basolateral membrane of MPT cells. Vanadate mimics this effect of ATP depletion, whereas genistein ameliorates the reduction in the intensity of E-cadherin staining induced by ATP depletion. Because it is has been well established that hyperphosphorylation of the catenins leads to dissociation of the adherens junction and to dysfunction of the junctional complex, it is proposed that the increase in tyrosine phosphorylation of catenins observed in MPT cells during ATP depletion contributes to the loss of function of the junctional complex associated with sublethal injury.

Adenosine Triphosphate↗

PubMed1999

O-raffinose cross-linking markedly reduces systemic and renal vasoconstrictor effects of unmodified human hemoglobin.

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.

Animals↗

PubMed1998

Lysophosphatidic acid is a major serum noncytokine survival factor for murine macrophages which acts via the phosphatidylinositol 3-kinase signaling pathway.

Lysophosphatidic acid (LPA) is the smallest and structurally simplest of all the glycerophospholipids. It occurs normally in serum and binds with high affinity to albumin, while retaining its biological activity. The effects of LPA are pleiotropic and range from mitogenesis to stress fiber formation. We show a novel role for LPA: as a macrophage survival factor with potency equivalent to serum. Administration of LPA protects macrophages from apoptosis induced by serum deprivation, and protection is equivalent to that with conventional survival factors such as macrophage colony stimulating factor. The ability of LPA to act as a survival factor is mediated by the lipid kinase phosphatidylinositol 3-kinase (PI3K), since LPA activated both the p85-p110 and p110gamma isoforms of PI3K and macrophage survival was blocked completely by wortmannin or LY294002, two mechanistically dissimilar inhibitors of PI3K. pp70(s6k), a downstream kinase activated by PI3K, also contributes to survival, because inhibitors of pp70(s6k), such as rapamycin, blocked macrophage survival in the presence of LPA. Modified forms of LPA and phospholipids, such as phosphatidylcholine and phosphatidylethanolamine, had no survival effect, thereby showing the specificity of LPA. These results show that LPA acts as a potent macrophage survival factor. Based on striking similarities between our LPA and serum data, we suggest that LPA is a major noncytokine survival factor in serum.

Androstadienes↗

PubMed1998

Biology of ischemic and toxic renal tubular cell injury: role of nitric oxide and the inflammatory response.

Acute ischemic or toxic injury to the kidney induces alterations in the expression of many genes. Some of these molecular responses have been termed 'maladaptive' because they exacerbate the tubular damage induced by the initiating insult. Some maladaptive responses include alterations in the activity of nitric oxide synthases and expression of cytokines and adhesion molecules that mediate an inflammatory response. This review focuses on the role of nitric oxide and inflammation in influencing the course of acute renal failure due to ischemic and toxic tubular injury.

Acute Kidney Injury↗

PubMed1998

Graded ATP depletion can cause necrosis or apoptosis of cultured mouse proximal tubular cells.

The mechanisms of cell death induced by ATP depletion were studied in primary cultures of mouse proximal tubular (MPT) cells. Graded ATP depletion, ranging in severity from approximately 2 to 70% of control levels, was induced by incubating cells with either antimycin or 2-deoxyglucose, with varying concentrations of dextrose. We found that cells subjected to ATP depletion below approximately 15% of control died uniformly of necrosis. In contrast, cells subjected to ATP depletion between approximately 25 and 70% of control all died by apoptosis. The rapidity of cell death was proportional to the severity of reduction of cell ATP content and was independent of the mechanism of cell death. Renal growth factors, epidermal growth factor (EGF) and high-dose insulin, did not ameliorate apoptotic cell death induced by ATP depletion. We conclude that ATP depletion can cause either necrosis or apoptosis in MPT cells. Furthermore, we have identified a narrow range of ATP depletion (approximately 15 to 25% of control) representing a threshold that determines whether cells die by necrosis or apoptosis.

Adenosine Triphosphate↗

PubMed1998

Acute renal failure. I. Relative importance of proximal vs. distal tubular injury.

For more than 15 years, there has been an ongoing debate regarding the nephron segment(s) most severely injured in acute renal failure (ARF) induced by an ischemic or toxic insult. Although some investigators have argued that the proximal tubule (and particularly the S3 segment) is the major target of injury in ARF, others have held the view that damage to the distal nephron [particularly the medullary thick ascending limb (MTAL) segment] plays a more important role in this disease. In this discussion, the first of three on different aspects of ARF that have been hotly debated, we have invited several experts to discuss their opinions on this issue. The goals of this first discussion (and the subsequent two articles in this forum) are to establish areas of consensus in each area of controversy and also to identify unanswered questions that represent important areas for future research.

Animals↗

PubMed1998

Role of superoxide in apoptosis induced by growth factor withdrawal.

We have examined the role of reactive oxygen species (ROS) in apoptosis induced by growth factor deprivation in primary cultures of mouse proximal tubular (MPT) cells. When confluent monolayers of MPT cells are deprived of all growth factors, the cells die by apoptosis over a 10- and 14-day period. Both epidermal growth factor (EGF) and high-dose insulin directly inhibit apoptosis of MPT cells deprived of growth factors. Growth factor deprivation results in an increase in the cellular levels of superoxide anion while apoptosis of MPT cells induced by growth factor withdrawal is inhibited by a number of antioxidants and scavengers of ROS. Growth factor deprivation also results in activation of caspase activity, which is inhibited by EGF and high-dose insulin as well as by the ROS scavengers and antioxidants that inhibit apoptosis. The cell-permeant caspase inhibitor, z-Val-Ala-Asp-CH2F (zVAD-fmk), prevents the increase in caspase activity and markedly inhibits apoptosis induced by growth factor deprivation. However, zVAD-fmk had no effect on the increased levels of superoxide associated with growth factor deprivation. Thus we provide novel evidence that ROS play an important role in mediating apoptosis associated with growth factor deprivation. ROS appear to act upstream of caspases in the apoptotic pathway. We hypothesize that oxidant stress, induced by growth factor withdrawal, represents a signaling mechanism for the default pathway of apoptosis.

Amino Acid Chloromethyl Ketones↗

PubMed1998

Necrosis and apoptosis in acute renal failure.

Renal tubular cells that are lethally injured after an acute ischemic or nephrotoxic insult to the kidney can die by necrosis or apoptosis. Necrosis is usually the result of overwhelming and severe cellular ATP depletion. In contrast, there are many potential causes of apoptosis in acute renal failure (ARF). These include cytotoxic events not severe enough to induce necrosis, a relative deficiency of renal growth factors, and loss of cell-matrix or cell-cell adhesive interactions. In some situations, receptor-mediated events induced by tumor necrosis factor-alpha (TNF-alpha) or Fas (CD95) may play a role in apoptosis in ARF. Necrosis and apoptosis are distinct morphologically and biochemically. Necrosis results in an early loss of plasma membrane integrity, the release of injurious substances from the cytosol, and an inflammatory reaction in the surrounding tissue that is readily detected morphologically. In contrast, apoptosis is characterized by progressive cell shrinkage with condensation and fragmentation of nuclear chromatin. Apoptotic cells ultimately break up into plasma membrane-bound vesicles called "apoptotic bodies" that are rapidly phagocytosed by macrophages and neighboring epithelial cells. In experimental models of ARF in vivo, apoptosis of renal tubular cells has been shown to occur in two distinct phases. The first phase of apoptosis occurs early on, between 12 and 48 hours after the acute ischemic or nephrotoxic insult. The second phase of apoptosis occurs many days later, during the recovery phase of ARF. Tubular cell apoptosis occurring shortly after the acute insult probably contributes to tubular cell loss and the tubular dysfunction associated with ARF. In contrast, the apoptosis associated with the recovery phase has been postulated to contribute to the remodeling of injured tubules and to facilitate their return to a normal structural and functional state. Therapeutic interventions that inhibit or promote apoptosis of renal tubular cells have the potential for minimizing renal dysfunction and accelerating recovery after ARF.

Acute Kidney Injury↗

PubMed1997

Fatty acid-induced cytotoxicity: differences in susceptibility between MDCK cells and primary cultures of proximal tubular cells.

We have compared the cytotoxicity of exogenously added fatty acid (oleic acid) and that of endogenous free fatty acids released from cell membranes by phospholipase A2 in primary cultures of mouse proximal tubular (MPT) cells and in Madine-Darby canine kidney (MDCK) cells. Exposure of MPT cell monolayers to oleic acid (125 mmol/L) for 2 hours resulted in severe irreversible injury to 70% +/- 4% of MPT cells. In striking contrast, only 8% +/- 3% of MDCK cells were killed by the same insult. This striking difference in the response to exogenous oleate by MPT and MDCK cells was associated with modest and comparable reductions in cell adenosine triphosphate (ATP) content in both cell types. Chemical anoxia induced by cyanide plus deoxyglucose (CN-DOG) in the absence of glucose was associated with greater injury in MPT cells (45% +/- 6% killed) than in MDCK cells (16% +/- 5% cells killed) despite severe and comparable depletion of cell ATP content in both MPT cells (96.0% +/- 0.6% reduction) and MDCK cells (96.0% +/- 0.5% reduction). The release of endogenous fatty acids by the exposure of cells to exogenous phospholipase A2 caused mild injury in both cell types that was more severe in MPT cells than in MDCK cells. The combined insult of phospholipase A2 and chemical anoxia for 2 hours caused substantially greater cell injury in both MPT and MDCK cells than either intervention alone, but the combined insult was still more damaging to MPT cells (73% +/- 4% killed) than to MDCK cells (30% +/- 4% killed). We conclude that the cell membrane in MDCK cells is intrinsically more resistant to fatty acid-induced injury than the lipid membrane in MPT cells.

Adenosine Triphosphate↗

PubMed1997

Heat stress ameliorates ATP depletion-induced sublethal injury in mouse proximal tubule cells.

The role of prior heat stress (HS) in ameliorating changes in the actin cytoskeleton and the loss of tight junction integrity that accompany ATP depletion was examined. Mouse proximal tubule cells in primary culture were exposed to sodium cyanide (CN) in the absence of dextrose for 1 h, a maneuver that produced equivalent degrees of ATP depletion in control and in HS cells. After ATP depletion, actin stress fibers were completely disrupted in control cells. In contrast, HS cells with elevated HSP-72 content showed preservation of stress fibers after CN exposure. ATP depletion in control and HS cells produced similar and reversible depletion of the G-actin pool without altering total actin content. Integrity of the tight junction was assessed by transepithelial electrical resistance (TER) and unidirectional flux of lucifer yellow (LY, mol wt 482). After CN alone, the nadir in TER was lower than that of HS + CN cells (51.6 +/- 2.5 vs. 96.2 +/- 3.2 omega x cm2, respectively; P < 0.05). After 30-min recovery, TER of HS + CN recovered to control values (277 +/- 7.2 vs. 227 +/- 6.6 omega x cm2; P > 0.05), whereas CN did not (165 +/- 7.3 vs. 227 +/- 6.6 omega x cm2; P < 0.05). Changes in LY flux paralleled those in TER. HS is associated with preservation of the actin cytoskeleton and improved integrity of the tight junction after sublethal ATP depletion injury. These protective effects may contribute to the preservation of epithelial cell polarity and function following an ischemic insult.

Actins↗