Blockage of macrophage migration inhibitory factor (MIF) suppressed uric acid-induced vascular inflammation,smooth muscle cell de-differentiation,and remodeling

Hyperuricemia contributes to vascular injury and dysfunction, yet the potential mechanisms are not well understood. Uric acid (UA) has been found to stimulate macrophage migration inhibitory factor (MIF) up-regulation in renal tubules from rats subjected to UA-induced nephropathy. Given that MIF is able to induce vascular smooth muscle cell (VSMC) de-differentiation (from contractile state to a secretory state), we thus hypothesized that UA-induced vascular injury is via up-regulating of MIF in VSMCs, which enhancing vascular inflammation and VSMC transition. Within a mouse model of UA injection (500?mg/kg, twice/day, 14 days), we measured circulating and vascular MIF levels under UA stimulation at 6?h, day 1, and 14. We tested the efficacy of MIF inhibitor (10?mg/kg, twice/day, 14 days) on UA-induced vascular inflammation and remodeling. High plasma level of UA induced vascular MIF release into the plasma at acute phase. In the chronic phase, the protein level of MIF is up-regulated in the vessels. MIF inhibitor suppressed vascular inflammatory responses, repressed VSMC de-differentiation, and attenuated vascular remodeling and dysfunction following UA stimulation. Knockdown of MIF in cultured VSMCs repressed UA-induced de-differentiation. Our results provided a novel mechanism for MIF-mediated vascular injury in response to UA stimulation, and suggested that anti-MIF interventions may be of therapeutic value in hyperuricemic patients.     

Dimethylthiourea protects against mitochondrial oxidative damage induced by cisplatin in liver of rats

Cisplatin is one of the most effective chemotherapeutic agents. However, at higher doses liver injury may occur. The purpose of this study was to explore whether the hydroxyl radical scavenger dimethylthiourea (DMTU) protects against cisplatin-induced oxidative damage in vivo and to define the mitochondrial pathways involved in cytoprotection. Adult male Wistar rats (200–220 g) were divided into four groups of eight animals each. The control group was treated only with an intraperitoneal (i.p.) injection of saline solution (1 ml/100 g body weight). The DMTU group was given only DMTU (500 mg/kg body weight, i.p), followed by 125 mg/kg body weight, i.p. (twice a day) until sacrifice. The cisplatin group was given a single injection of cisplatin (10 mg/kg body weight, i.p.). The DMTU + cisplatin group was given DMTU (500 mg/kg body weight, i.p.), just before the cisplatin injection (10 mg/kg body weight, i.p.), followed by injections of DMTU (125 mg/kg body weight, i.p.) twice a day until sacrifice (72 h after the treatment). DMTU did not present any direct effect on mitochondria and substantially inhibited cisplatin-induced mitochondrial damage in liver, therefore preventing elevation of AST and ALT serum levels. DMTU protected against (a) decreased hepatic ATP levels; (b) lipid peroxidation; (c) cardiolipin oxidation; (d) sulfhydryl protein oxidation; (e) mitochondrial membrane rigidification; (f) GSH oxidation; (g) NADPH oxidation; (h) apoptosis. Results suggest that antioxidants, particularly hydroxyl radical scavengers, protect liver mitochondria against cisplatin-induced oxidative damage. Several mitochondrial changes were delineated and proposed as interesting targets for cytoprotective strategy.     

Pretreatment with catalase or dimethyl sulfoxide protects alloxan-induced acute lung edema in dogs.

We tested the preventive effects of catalase, an enzymatic scavenger of hydrogen peroxide, or dimethyl sulfoxide (DMSO), a hydroxyl radical scavenger, on intravenous alloxan-induced lung edema in four groups of pentobarbital sodium-anesthetized, ventilated dogs for 3 h: saline (20 ml.kg-1.h-1) infusion alone (n = 5), alloxan (75 mg/kg) + saline infusion (n = 5), catalase (150,000 U/kg) + alloxan + saline infusion (n = 5), or DMSO (4 mg/kg) + alloxan + saline infusion (n = 5). Catalase or DMSO significantly prevented the increase in plasma thromboxane B2 and 6-keto-prostaglandin F1 alpha over 3 h after alloxan and the accumulation of extravascular lung water after 3 h [3.95 +/- 0.52 (SE) g/g with catalase, 3.06 +/- 0.42 g/g with DMSO] but not early pulmonary arterial pressor response. An electron microscopic study indicated that catalase or DMSO significantly reduced the endothelial cellular damages after alloxan. These findings strongly suggest that hydrogen peroxide and hydroxyl radical are major mediators responsible for intravenous alloxan-induced edematous lung injury in anesthetized ventilated dogs.     

Enhanced intrarenal receptor-mediated prorenin activation in chronic progressive anti-thymocyte serum nephritis rats on high salt intake

Despite suppression of the circulating renin-angiotensin system (RAS), high salt intake (HSI) aggravates kidney injury in chronic kidney disease. To elucidate the effect of HSI on intrarenal RAS, we investigated the levels of intrarenal prorenin, renin, (pro)renin receptor (PRR), receptor-mediated prorenin activation, and ANG II in chronic anti-thymocyte serum (ATS) nephritic rats on HSI. Kidney fibrosis grew more severe in the nephritic rats on HSI than normal salt intake. Despite suppression of plasma renin and ANG II, marked increases in tubular prorenin and renin proteins without concomitant rises in renin mRNA, non-proteolytically activated prorenin, and ANG II were noted in the nephritic rats on HSI. Redistribution of PRR from the cytoplasm to the apical membrane, along with elevated non-proteolytically activated prorenin and ANG II, was observed in the collecting ducts and connecting tubules in the nephritic rats on HSI. Olmesartan decreased cortical prorenin, non-proteolytically activated prorenin and ANG II, and apical membranous PRR in the collecting ducts and connecting tubules, and attenuated the renal lesions. Cell surface trafficking of PRR was enhanced by ANG II and was suppressed by olmesartan in Madin-Darby canine kidney cells. These data suggest the involvement of the ANG II-dependent increase in apical membrane PRR in the augmentation of intrarenal binding of prorenin and renin, followed by nonproteolytic activation of prorenin, enhancement of renin catalytic activity, ANG II generation, and progression of kidney fibrosis in the nephritic rat kidneys on HSI. The origin of the increased tubular prorenin and renin remains to be clarified. Further studies measuring the urinary prorenin and renin are needed.     

Adaptational effect of dimethylsulfoxide during chronic emotional-painful stress in rats

Intraperitoneal injection of hydroxyl radical scavenger dimethyl sulfoxide (DMSO) to rats (1 g/kg body weight, daily for 3 weeks) did not change their behaviour in the open field and had no effect on blood pressure and heart and respiratory rates. DMSO injection before stress prevented changes usually induced by chronic (three-week) emotional-painful stress. DMSO increased superoxide dismutase activity in brain homogenates and serum. It is suggested that hydroxyl radicals (OH) play the key role in the realization of stress-induced effects and that molecular mechanisms of anti-stress effects of DMSO include both scavenging of hydroxyl radicals and activation of superoxide dismutase.     

Dimethylthiourea decreases acute pulmonary edema induced by phorbol myristate acetate in isolated blood-perfused lung of the rat

Acute pulmonary edema can be induced by phorbol myristate acetate (PMA). Oxygen radicals released from the neutrophils have been considered to play an important role in the pathogenesis of PMA-induced pulmonary edema. In the present experiment, we studied the effect of dimethylthiourea (DMTU) on PMA-induced pulmonary injuries in isolated perfused lungs of rats. DMTU is a potent scavenger of the hydroxyl radical and hydrogen peroxide. PMA infusion into the isolated lung increased pulmonary arterial pressure (delta PAP) by 37.8 +/- 3.9 mmHg. The lung weight gain (LWG) and lavage albumin concentration (LAC) amounted to 6.2 +/- 1.2 g and 102.0 +/- 22.9 mg/dl, respectively. DMTU (100 mM) pretreatment significantly reduced the PAP increase (delta PAP = 4.6 +/- 0.8 mmHg, p less than 0.001), LWG (0.3 +/- 0.1 g, p less than 0.01) and LAC (25.3 +/- 1.7 mg/dl, p less than 0.01). Additional in vitro experiments demonstrated that DMTU depressed the chemiluminescence released from neutrophils activated by PMA (17.9 +/- 2.6 mV.min to 2.6 +/- 0.5 mV.min, p less than 0.01). The results suggest that DMTU, a scavenger of toxic radicals, decreases the lung edema through both attenuation of pulmonary hypertension and protection of vascular permeability from PMA injury.     

Inability of dimethylthiourea to limit tissue necrosis during acute myocardial infarction in rabbits.

This study examined the effect of treatment with dimethylthiourea (DMTU), a highly cell-permeable scavenger of hydroxyl radicals, on tissue necrosis in rabbit hearts during myocardial ischemia and reperfusion. Sixty-two rabbits underwent 45 minutes of coronary occlusion with, or without, coronary reperfusion for 3 hours. A saline vehicle, or DMTU (500 mg/kg intravenously [iv]) was administered over 45 minutes starting either 10 minutes before or 10 minutes after coronary occlusion, or 10 minutes before coronary reperfusion. Anatomic risk zone size was assessed using microsphere autoradiography, and the area of necrosis was determined using tetrazolium staining. Cardiac hemodynamics and risk zone size were similar for all treatment groups. No differences were observed in the extent of tissue necrosis (normalized to risk zone size) for saline- and DMTU-treated rabbits subjected to 45 minutes (61.2 +/- 23.1% vs. 70.6 +/- 16.5%) or 225 minutes (82.8 +/- 5.4% vs. 78.3 +/- 5.9%) of permanent coronary occlusion without reperfusion. Similarly, tissue necrosis in rabbits with 45 minutes coronary occlusion followed by 3 hours reperfusion was not significantly reduced when DMTU was administered either 10 minutes before coronary occlusion, 10 minutes after coronary occlusion, or 10 minutes before coronary reperfusion (67.0 +/- 9.9%; 57.6 +/- 10.6%; 68.3 +/- 13.3%) compared to saline-treated controls (76.6 +/- 10.5%). These results demonstrate that the hydroxyl radical scavenger DMTU does not appear to influence the progression of myocyte injury in this experimental model of acute myocardial infarction.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of poly(ADP-ribose)polymerase in cisplatin-induced injury in LLC-PK1 cells

Acute renal failure is a dose-limiting factor during cisplatin chemotherapy. We have previously shown in rats that the hydroxyl radical scavenger edaravone reverses cisplatin-induced in vivo renal damage. In the present study, the role of poly(ADP-ribose) polymerase (PARP) in cisplatin nephrotoxicity was investigated in porcine tubular cells LLC-PK1. Cell injury was elicited by transient exposure to 500 microM cisplatin for 1 h or continuous exposure to 30 microM cisplatin for 24 h. Various hydroxyl radical scavengers reversed cell damage in a transient but not permanent model. The cell injury seemed to be necrosis and apoptosis in transient and permanent models, respectively, as assessed by TUNEL method and Annexin V stain. PARP inhibitors such as 3-aminobenzamide and benzamide inhibited cell damage in transient but not permanent model. PARP-dependent cell injury was also observed after transient exposure to hydroxyl radical-generating solution. We demonstrated for the first time the activation of PARP in renal tubular cells by transient cisplatin exposure, as determined by immunofluorescent stain with anti-poly(ADP-ribose) antibody. Moreover, ATP was depleted by transient exposure to cisplatin or hydroxyl radical, both of which were reversed by PARP inhibitors. These findings suggest that hydroxyl radical generation followed by PARP activation contributes to the necrotic cell injury caused by a transient exposure to cisplatin.     

Mechanism of action of an old antibiotic revisited: Role of calcium ions in protonophoric activity of usnic acid

Usnic acid (UA), an old antibiotic and one of the first described mitochondrial uncouplers, has demonstrated many beneficial activities, such as antimicrobial, antiviral, antitumour and anti-inflammatory properties. Here, we performed a thorough investigation of effects of usnic acid and its analogues on artificial planar bilayer lipid membrane (BLM), rat liver mitochondria and bacteria. Surprisingly enough, all of the three hydroxyl groups of UA appeared to be involved in its proton-shuttling activity on BLM. We ascribed this fact to an ability of UA to form complexes with calcium ions, aiding it in cycling protons across the membrane. Actually, the addition of calcium ions markedly stimulated the UA-induced electrical current across BLM. By using the calcium ionophore A23187, we proved the involvement of calcium ions in the UA uncoupling action on isolated rat liver mitochondria. The calcium-chelating property of UA was demonstrated here by the method of extracting metal ions into a hydrophobic phase. Modification of any of the hydroxyl groups in UA dramatically reduced not only the UA-induced current across BLM and the UA-mediated calcium extraction, but also the uncoupling activity of UA in mitochondria and the inhibiting effect of UA on the growth of Bacillus subtilis. The ability of UA to cause dissipation of membrane potential in isolated liver mitochondria and bacterial cells was shown here for the first time. In view of the data obtained, the protonophoric activity of UA is considered to make a significant contribution to its antibacterial action.     

Renal functional responses to selective intrarenal renin inhibition in Cyp1a1-Ren2 transgenic rats with ANG II-dependent malignant hypertension

Angiotensin (ANG) II-dependent hypertension is characterized by increases in intrarenal ANG II levels, derangement in renal hemodynamics, and augmented tubular sodium reabsorptive capability. Increased nephron expression of renin-angiotensin system components, such as angiotensinogen by proximal tubule cells and renin by collecting duct principal cells, has been associated with an augmented ability of the kidney to form ANG II in hypertensive states. However, the contribution of de novo intrarenal ANG II production to the development and maintenance of ANG II-dependent hypertension remains unclear. The present study was performed to determine the effects of selective intrarenal renin inhibition on whole kidney hemodynamics and renal excretory function in Cyp1a1-Ren2 rats with ANG II-dependent malignant hypertension in the absence of the confounding influence of associated reductions in mean arterial pressure (MAP). Male Cyp1a1-Ren2 transgenic rats were induced to develop malignant hypertension, anesthetized, and surgically prepared for intrarenal administration of the direct renin inhibitor aliskiren (0.01 mg/kg). Following acute aliskiren treatment, urine flow and sodium excretion increased (10.5 ± 1.1 to 15.9 ± 1.9 μl/min, P < 0.001; 550 ± 160 to 1,370 ± 320 neq/min, P < 0.001, respectively) and ANG II excretion decreased (120 ± 30 to 63 ± 17 fmol/h, P < 0.05). There were no significant changes in MAP, glomerular filtration rate, estimated renal plasma flow, plasma ANG II levels, or protein excretion. The present findings demonstrate that selective renal renin inhibition elicits diuretic and natriuretic responses in Cyp1a1-Ren2 rats with ANG II-dependent malignant hypertension. Elevated intraluminal ANG II levels likely act to augment tubular reabsorptive function and, thereby, contribute to the elevated blood pressure in Cyp1a1-Ren2 rats with ANG II-dependent malignant hypertension.     

Dimethylthiourea does not ameliorate reperfusion lung injury in dogs or rabbits

We previously demonstrated that in vivo reperfusion of a dog lung after 48 h of pulmonary arterial (PA) ischemia results in pulmonary edema with a significant infiltrate of polymorphonuclear leukocytes. We hypothesized that the injury resulted from production of hydroxyl radical by activated neutrophils. In the current study, we attempted to prevent the injury in both dogs and rabbits with dimethylthiourea (DMTU), a scavenger of hydroxyl radical. After 48 h of left PA occlusion in 18 dogs, DMTU was administered to 9 animals and 9 were not treated. The occlusion was then released, and the dogs were killed 4 h later. Reperfusion resulted in a drop in leukocyte count and left lung edema, but there was no difference between treated and untreated animals. The wet-to-dry ratios of the lungs in the treated group were 5.76 +/- 0.44 (SE) on the reperfused left side and 4.50 +/- 0.06 (P less than 0.05) on the right side. In the untreated groups the comparable ratios were 5.73 +/- 0.31 and 4.92 +/- 0.10 (P less than 0.05 for right vs. left). Histological examination revealed significant differences between the right and left lungs in the extent of intra-alveolar granulocytes and macrophages but did not reveal differences between the treated and untreated animals. To ensure that neither the model nor the lack of response to DMTU was species specific, we then developed a rabbit model of reperfusion edema.(ABSTRACT TRUNCATED AT 250 WORDS)     

Uric acid decreases NO production and increases arginase activity in cultured pulmonary artery endothelial cells

Elevated levels of serum uric acid (UA) are commonly associated with primary pulmonary hypertension but have generally not been thought to have any causal role. Recent experimental studies, however, have suggested that UA may affect various vasoactive mediators. We therefore tested the hypothesis that UA might alter nitric oxide (NO) levels in pulmonary arterial endothelial cells (PAEC). In isolated porcine pulmonary artery segments (PAS), UA (7.5 mg/dl) inhibits acetylcholine-induced vasodilation. The incubation of PAEC with UA caused a dose-dependent decrease in NO and cGMP production stimulated by bradykinin or Ca(2+)-ionophore A23187. We explored cellular mechanisms by which UA might cause reduced NO production focusing on the effects of UA on the l-arginine-endothelial NO synthase (eNOS) and l-arginine-arginase pathways. Incubation of PAEC with different concentrations of UA (2.5-15 mg/dl) for 24 h did not affect l-[(3)H]arginine uptake or activity/expression of eNOS. However, PAEC incubated with UA (7.5 mg/dl; 24 h) released more urea in culture media than control PAEC, suggesting that arginase activation might be involved in the UA effect. Kinetic analysis of arginase activity in PAEC lysates and rat liver and kidney homogenates demonstrated that UA activated arginase by increasing its affinity for l-arginine. An inhibitor of arginase (S)-(2-boronoethyl)-l-cysteine prevented UA-induced reduction of A23187-stimulated cGMP production by PAEC and abolished UA-induced inhibition of acetylcholine-stimulated vasodilation in PAS. We conclude that UA-induced arginase activation is a potential mechanism for reduction of NO production in PAEC.     

Digital vascular imaging and selective renin sampling in evaluation of vascular anatomy in renal transplant recipients.

Sixty-five renal transplant recipients underwent digital vascular imaging of the graft and simultaneous selective venous sampling for plasma renin activity. Renal artery stenosis was found in seven patients but did not appear to be functionally important. Diffuse intrarenal arterial attenuation was found in seven patients and was associated with impaired graft function and perfusion; it may indicate chronic rejection. Lower pole hypoperfusion was found in nine patients without impaired graft function or perfusion; its clinical relevance is uncertain. Aneurysmal dilatation of the main renal artery was found in two patients. Severe hypertension was common in patients with these three major abnormalities, but a causal association between the abnormality and hypertension could rarely be inferred. It may be the abnormalities on digital vascular imaging, especially diffuse intrarenal arterial attenuation and lower pole hypoperfusion, are secondary to severe hypertension. Digital vascular imaging with simultaneous selective venous sampling for plasma renin activity is useful in evaluating the vascular anatomy of the grafted kidney and in assessing any abnormality found. The combined procedure was well tolerated by all patients with no complications and no incidence of acute tubular dysfunction or proteinuria after the investigation.     

The antioxidants dimethylsulfoxide and dimethylthiourea affect the immediate adhesion responses of larval haemocytes from 3 lepidopteran insect species

Antioxidants, dimethylsulfoxide (DMSO) and dimethylthiourea (DMTU), at concentrations not affecting the viability of blood cells (haemocytes) from the larval stage of 3 lepidopteran insects - Galleria mellonella, Lymantria dispar, and Malacosoma disstria - differed in their influence on the innate binding of haemocytes to glass, bacteria to haemocytes, and on humoral responses to alien materials. In vitro DMSO had little effect, whereas DMTU substantially impaired the adhesion of the haemocyte types, the plasmatocytes and granular cells, to slides as well as the attachment of Bacillus subtilis to these haemocytes. Although both antioxidants increased lysozyme and phenoloxidase activities, there was no correlation of enzyme activity and haemocyte adhesion responses, possibly reflecting sequestered radicals. Nitric oxide and hydroxyl radicals offset the DMTU effect. In the absence of antioxidants, inactivate protein kinases A (PKA) and C (PKC) enhanced haemocyte aggregation. In general, DMSO, as opposed to DMTU, did not alter the effects of PKA and PKC activators and inhibitors on haemocyte aggregation or of PKC and PKA activities. High concentrations of DMSO and all levels of DMTU, although inhibiting PKA and PKC, inhibited haemocyte adhesion to slides. Comparable results occurred for DMTU-treated haemocytes incubated with B. subtilis. In vivo DMSO, unlike DMTU, did not impair plasmatocyte or granular cell responses to foreign materials, including bacterial removal from the haemolymph and nodulation.     

Dimethylthiourea, but not dimethylsulfoxide, reduces canine myocardial infarct size

We studied the effect of treatment with two diffusible, low molecular weight scavengers of toxic oxygen metabolites, dimethylthiourea (DMTU) and dimethylsulfoxide (DMSO), on canine infarcts caused by 90 min of ischemia and 3 h of reperfusion. Infarct size was determined by incubating ventricular slices with triphenyl tetrazolium chloride. Areas at risk were determined by autoradiography of 99Tc microspheres injected in vivo during ischemia and were similar (p greater than 0.05) in DMTU, DMSO, and saline treated dogs. However, the ratio of infarct size to area at risk was reduced (p less than 0.05) in dogs treated 30 min before reperfusion with 500 mg/kg DMTU (31.1 +/- 4.6%, n = 9) compared with saline treated dogs (53.4 +/- 4.6% n = 9). In contrast, the ratio of infarct size to area at risk was not significantly different (p greater than 0.05) in dogs treated with 2000 mg/kg DMSO 30 min before reperfusion (43.7 +/- 4.3%) compared to saline treated dogs. The serum concentration of DMTU (4.5 mM) was one-tenth that of DMSO (48 mM) in early reperfusion. Therefore, DMTU but not DMSO protected against post-ischemic cardiac reperfusion injury.     

Endotoxemia causes increased lung tissue lipid peroxidation in unanesthetized sheep

Our purpose was to determine whether lipid peroxidation of lung tissue, a reflection of O2 radical injury, occurs with endotoxin, and whether the degree of tissue change corresponds with the degree of increased protein permeability. Unanesthetized adult sheep with lung lymph fistulas were given Escherichia coli endotoxin at a dose of 2 micrograms/kg (n = 34). Tissue lipid peroxidation was measured using the thiobarbituric acid assay for malondialdehyde (MDA). The MDA content of lung tissue in nanomoles per gram increased from a control value of 48 +/- 8 to 98 +/- 18 at 5 h postendotoxin (2 micrograms/kg), whereas lung lymph protein transport (Cp), was increased 3- to 4-fold. The MDA content returned to base line with Cp by 24 h postendotoxin. Six sheep given endotoxin were pretreated with 12.5 mg/kg of ibuprofen, and six were infused with dimethylthiourea (DMTU) 0.75 g/kg. With ibuprofen, Cp was only increased 2.5- to 3-fold and MDA was increased to 69 +/- 15 nmol/g. With DMTU, the increase in Cp was comparable to that with endotoxin alone, as was the MDA of lung tissue with a value of 92 +/- 12 nmol/g. The correlation of tissue MDA with Cp in all animals was 0.83. We conclude that lipid peroxidation occurs in lung tissue after a moderately severe endotoxin injury with the degree of change corresponding to the degree of increased Cp.     

Synthesis of creatol, a hydroxyl radical adduct of creatinine and its increase by puromycin aminonucleoside in isolated rat hepatocytes

Creatol is a hydroxyl radical adduct of creatinine and the precursor of methylguanidine (MG), a uremic toxin. We investigate the synthesis of creatol and MG from creatinine and the effect of substances that affect the hydroxyl radical in isolated rat hepatocytes. In the presence of increasing concentrations of creatinine, rising level of creatol were found after 2 h incubation in Krebs-Henseleit bicarbonate buffer. However, further increase of creatol was not observed after 4 and 6 h incubations. On the other hand, MG after 2 h incubation achieved a level of about 50% that of creatol and increased depending on both the creatinine concentration and the incubation period. DMSO, a hydroxyl radical scavenger decreased the generation of creatol and MG by about 50% at 2.5 mM and the inhibition depended on DMSO concentration. Puromycin aminonucleoside (PAN) increased both by about 170%. These findings demonstrated that hepatocytes synthesize creatol prior to MG and are inhibited by a hydroxyl radical scavenger. They also show that PAN increased hydroxyl radical generation in tissue cells.     

HIV-induced kidney cell injury: role of ROS-induced downregulated vitamin D receptor

Reactive oxygen species (ROS) have been demonstrated to contribute to HIV-induced tubular cell injury. We hypothesized that HIV-induced ROS generation may be causing tubular cell injury through downregulation of vitamin D receptor (VDR) and associated downstream effects. In the present study, HIV not only downregulated tubular cell VDR expression but also inflicted DNA injury. On the other hand, EB-1089, a VDR agonist (VD), inhibited both downregulation of VDR and tubular cell DNA injury in the HIV milieu. H(2)O(2) (an O(-) donor) directly downregulated tubular cell VDR, whereas catalase, a free radical scavenger, inhibited HIV-induced downregulation of tubular cell VDR expression. HIV also stimulated the tubular cell renin-angiotensin system (RAS) through downregulation of VDR. Because losartan (an ANG II blolcker) partially inhibited HIV-induced tubular cell ROS generation while ANG II directly stimulated tubular cell ROS generation, it appears that HIV-induced ROS production was partly contributed by the RAS activation. VD not only inhibited HIV-induced RAS activation but also attenuated tubular cell ROS generation. Tubular cells displayed double jeopardy in the HIV milieu induction of double-strand breaks and attenuated DNA repair; additionally, in the HIV milieu, tubular cells exhibited enhanced expression of phospho-p53 and associated downstream signaling. A VDR agonist and an ANG II blocker not only preserved expression of tubular cell DNA repair proteins but also inhibited induction of double-strand breaks. In in vivo studies, renal cortical sections of Tg26 mice displayed attenuated expression of VDR both in podocytes and tubular cells. In addition, renal cortical sections of Tg26 mice displayed enhanced oxidative stress-induced kidney cell DNA damage. These findings indicated that HIV-induced tubular cell downregulation of VDR contributed to the RAS activation and associated tubular cell DNA damage. However, both VD and RAS blockade provided protection against these effects of HIV.     

Autophagy-dependent EIF2AK3 activation compromises ursolic acid-induced apoptosis through upregulation of MCL1 in MCF-7 human breast cancer cells

Ursolic acid (UA) is a pentacyclic triterpenoid with promising cancer chemopreventive properties. A better understanding of the mechanisms underlying anticancer activity of UA is needed for further development as a clinically useful chemopreventive agent. Here, we found that both endoplasmic reticulum (ER) stress and autophagy were induced by UA in MCF-7 human breast cancer cells. Surprisingly, ER stress was identified as an effect rather than a cause of UA-induced autophagy. Autophagy-dependent ER stress protected the cells from UA-induced apoptosis through EIF2AK3-mediated upregulation of MCL1. Activation of MAPK1/3 but not inhibition of MTOR pathway contributed to UA-induced cytoprotective autophagy in MCF-7 cells. Our findings uncovered a novel cellular mechanism involved in the anticancer activity of UA, and also provided a useful model to study biological significance and mechanisms of autophagy-mediated ER stress.