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Glomerular damage and proteinuria are important pathophysiological signs of chronic kidney disease. This study provides data obtained using a model developed based on the use of the anti-cancer drug sorafenib. Sorafenib is a tyrosine kinase inhibitor that acts through the signaling pathway associated with vascular endothelial growth factor and is widely used to treat various types of cancer. Sorafenib, on the other hand, causes serious side effects in patients, including the development of chronic kidney disease. This study was aimed at using the nephrotoxic properties of sorafenib to model chronic kidney disease in rats. We showed that rats treated with sorafenib for 8 weeks along with a diet high in salt (8% NaCl) develop hypertension with high systolic blood pressure of 80 mmHg, proteinuria with an increase in protein content of 75% higher , and a 4-fold increase in glomerular damage compared to the control group. In case of damage to the renal glomeruli caused by sorafenib, the level of transcripts that are involved in the synthesis of key glomerular proteins such as nephrine, podocin, synaptopodin and subplanin is significantly reduced. Also, when studying this model, activation of the endothelial-mesenchymal transition is observed. In the group of rats treated with sorafenib, the mRNA level for the WT-1 endothelial cell marker was reduced by 20%, while the concentration of the Col III, FSP-1, α-SMA and vimentin mesenchymal cell markers increased by 2–3 times. Thus, we developed a preclinical model of chronic kidney disease, expressed in damage to the renal glomeruli. We also demonstrated that glomerular damage in this model is associated with decreased expression of key structural glomerular proteins and activation of the endothelial-mesenchymal transition of the kidneys.

Epoxyeicosatrienoic acids (EETs) and their synthetic analogs have cardiovascular protective effects. Here, we investigated the action of a novel EET analog EET-B on the progression of post-myocardial infarction (MI) heart failure in spontaneously hypertensive rats (SHR). Adult male SHR were divided into vehicle- and EET-B (10 mg/kg/day; p.o., 9 weeks)-treated groups. After 2 weeks of treatment, rats were subjected to 30-min left coronary artery occlusion or sham operation. Systolic blood pressure (SBP) and echocardiography (ECHO) measurements were performed at the beginning of study, 4 days before, and 7 weeks after MI. At the end of the study, tissue samples were collected for histological and biochemical analyses. We demonstrated that EET-B treatment did not affect blood pressure and cardiac parameters in SHR prior to MI. Fractional shortening (FS) was decreased to 18.4 ± 1.0% in vehicle-treated MI rats compared with corresponding sham (30.6 ± 1.0%) 7 weeks following MI induction. In infarcted SHR hearts, EET-B treatment improved FS (23.7 ± 0.7%), markedly increased heme oxygenase-1 (HO-1) immunopositivity in cardiomyocytes and reduced cardiac inflammation and fibrosis (by 13 and 19%, respectively). In conclusion, these findings suggest that EET analog EET-B has beneficial therapeutic actions to reduce cardiac remodeling in SHR subjected to MI.

Although epoxyeicosatrienoic acid (EET) analogs have performed well in several acute and chronic kidney disease models, targeted delivery of EET analogs to the kidney can be reasonably expected to reduce the level of drug needed to achieve a therapeutic effect and obviate possible side effects. For EET analog kidney-targeted delivery, we conjugated a stable EET analog to folic acid via a PEG-diamine linker. Next, we compared the kidney targeted EET analog, EET-F01, to a well-studied EET analog, EET-A. EET-A or EET-F01 was infused i.v. and plasma and kidney tissue collected. EET-A was detected in the plasma but was undetectable in the kidney. On the other hand, EET-F01 was detected in the plasma and kidney. Experiments were conducted to compare the efficacy of EET-F01 and EET-A for decreasing cisplatin nephrotoxicity. Cisplatin was administered to WKY rats treated with vehicle, EET-A (10 mg/kg i.p.) or EET-F01 (20 mg/kg or 2 mg/kg i.p.). Cisplatin increased kidney injury markers, viz., blood urea nitrogen (BUN), N-acetyl-β-(D)-glucosaminidase (NAG), kidney injury molecule-1 (KIM-1), and thiobarbituric acid reactive substances (TBARS). EET-F01 was as effective as EET-A in decreasing BUN, NAG, KIM-1, TBARS, and renal histological injury caused by cisplatin. Despite its almost 2×-greater molecular weight compared with EET-A, EET-F01 was comparably effective in decreasing renal injury at a 10-fold w/w lower dose. EET-F01 decreased cisplatin nephrotoxicity by reducing oxidative stress and inflammation. These data demonstrate that EET-F01 targets the kidney, allows for a lower effective dose, and combats cisplatin nephrotoxicity. In conclusion, we have developed a kidney targeted EET analog, EET-F01, that demonstrates excellent potential as a therapeutic for kidney diseases.

Kidney fibrosis is a key event in the development of chronic kidney disease, leading to end-stage renal failure. Unfortunately, there are now few drugs capable of preventing fibrosis in the kidneys, which is accompanied by the progression of chronic kidney disease in the terminal stage of renal failure. The results show the effectiveness of the use of a new dual-acting agent DM509 in the prevention of renal fibrosis using a model of unilateral obstruction of the ureter in mice. DM509 is both a farnesoid X-receptor agonist and a soluble epoxyhydrolase inhibitor. In this study, there were 8-12 week old C57BL/6J males undergoing surgery, which led to the development of unilateral ureteral obstruction and a control group. Mice received DM509 (10 mg/kg/day) or DM509-free solution together with drinking water for 10 days the day before surgery. Samples of kidney and blood tissues were collected at the end of the experiment. In the unilateral ureteral obstruction group, kidney dysfunction was detected, which was accompanied by increased urea nitrogen content in the blood compared to the control group (63 ± 7 vs. 34 ± 6 mg/d). The reduction of urea nitrogen in the blood by 36 % in mice with unilateral ureteral obstruction treated with DM509 is shown compared to mice with this pathology without treatment, which in turn proved the effectiveness of DM509 in preventing renal dysfunction. In mice with unilateral ureteral obstruction, which did not receive DM509, the development of kidney fibrosis with a high content of hydroxyproline in the kidneys and also increased collagen content in histological sections of the kidneys were detected. In the DM509 group, the renal and collagen hydroxyproline content was 34-66 % lower, indicating the effectiveness of this agent in the treatment of renal fibrosis. Thus, we have shown that the new DM509 is effective in preventing renal dysfunction and renal fibrosis using a murine model of unilateral ureteral obstruction.

Genome-wide association studies have found a number of potential genes involved in blood pressure regulation; however, the functional role of many of these candidates has yet to be established. One such candidate gene is CLCN6 , which encodes the transmembrane protein, chloride channel 6 (ClC-6). Although the CLCN6 locus has been widely associated with human blood pressure regulation, the mechanistic role of ClC-6 in blood pressure homeostasis at the molecular, cellular, and physiological levels is completely unknown. In this study, we demonstrate that rats with a functional knockout of ClC-6 on the Dahl Salt-Sensitive rat background (SS- Clcn6 ) have lower diastolic but not systolic blood pressures. The effect of diastolic blood pressure attenuation was independent of dietary salt exposure in knockout animals. Moreover, SS- Clcn6 rats are protected from hypertension-induced cardiac hypertrophy and arterial stiffening; however, they have impaired vasodilation and dysregulated intracellular calcium handling. ClC-6 is highly expressed in vascular smooth muscle cells where it is targeted to the Golgi apparatus. Using bilayer electrophysiology, we provide evidence that recombinant human ClC-6 protein can function as a channel. Last, we demonstrate that loss of ClC-6 function reduces Golgi calcium stores, which may play a previously unidentified role in vascular contraction and relaxation signaling in vascular smooth muscle cells. Collectively, these data indicate that ClC-6 may modulate blood pressure by regulating Golgi calcium reserves, which in turn contribute to vascular smooth muscle function.

Hypertension is a global health problem that increases the risk of stroke, heart failure, and kidney disease. GWAS have linked the CLCN6 gene to blood pressure regulation. CLCN6 encodes the voltage-sensitive chloride channel 6 (ClC-6), an intracellular protein of little-known function. Due to its high expression in vascular smooth muscle cells (VSMCs) and its localization in the Golgi, we hypothesized that ClC-6 is involved in Ca 2+ uptake in the Golgi and that reducing Golgi Ca 2+ levels changes VSMC function and therefore blood pressure, potentially by altering protein trafficking or cell proliferation. Using a knock-out of Clcn6 on the Dahl salt-sensitive rat background, we determined that KO rats have impaired vasodilation in response to 10 μM acetylcholine in mesenteric arteries (82 ± 6% vs 41 ± 13%, N=6 rats, n ≥ 9 vessels, p2+ in response to acetylcholine at a greater frequency than WT. Additionally, arterial stiffness, determined by pulse-wave velocity (PWV) measurements, was greater in KO rats at baseline (4.5 ± 0.3 vs. 3.6 ± 0.2 mm/msec, N≥6 rats, p2+ , confocal microscopy with an organelle-specific Ca 2+ dye and pharmacological agents was used to measure Ca 2+ . SR Ca 2+ stores were depleted with thapsigargin + ATP, followed by release of Golgi Ca 2+ stores by emetine. Total Golgi calcium levels were determined by area under the curve measurements of recorded fluorescence changes. Emetine-sensitive Golgi Ca 2+ stores were significantly reduced in KO rat VSMCs compared to WT. In conclusion, these results suggest that ClC-6 regulates Golgi Ca 2+ levels, and alteration to these stores results in cellular changes that slow or prevent arterial stiffness during the development of hypertension. Future studies will assess vessel fibrosis, cell proliferation, and apoptosis to elucidate the underlying molecular mechanisms impacting vessel stiffness and Ca 2+ signaling.

Epoxyeicosatrienoic acids (EETs) decrease cardiac ischemia-reperfusion injury; however, the mechanism of their protective effect remains elusive. Here, we investigated the cardioprotective action of a novel EET analog, EET-B, in reperfusion and the role of hypoxia-inducible factor (HIF)-1α in such action of EET-B. Adult male rats were subjected to 30 min of left coronary artery occlusion followed by 2 h of reperfusion. Administration of 14,15-EET (2.5 mg/kg) or EET-B (2.5 mg/kg) 5 min before reperfusion reduced infarct size expressed as a percentage of the area at risk from 64.3 ± 1.3% in control to 42.6 ± 1.9% and 46.0 ± 1.6%, respectively, and their coadministration did not provide any stronger effect. The 14,15-EET antagonist 14,15-epoxyeicosa-5( Z)-enoic acid (2.5 mg/kg) inhibited the infarct size-limiting effect of EET-B (62.5 ± 1.1%). Similarly, the HIF-1α inhibitors 2-methoxyestradiol (2.5 mg/kg) and acriflavine (2 mg/kg) completely abolished the cardioprotective effect of EET-B. In a separate set of experiments, the immunoreactivity of HIF-1α and its degrading enzyme prolyl hydroxylase domain protein 3 (PHD3) were analyzed in the ischemic areas and nonischemic septa. At the end of ischemia, the HIF-1α immunogenic signal markedly increased in the ischemic area compared with the septum (10.31 ± 0.78% vs. 0.34 ± 0.08%). After 20 min and 2 h of reperfusion, HIF-1α immunoreactivity decreased to 2.40 ± 0.48% and 1.85 ± 0.43%, respectively, in the controls. EET-B blunted the decrease of HIF-1α immunoreactivity (7.80 ± 0.69% and 6.44 ± 1.37%, respectively) and significantly reduced PHD3 immunogenic signal in ischemic tissue after reperfusion. In conclusion, EET-B provides an infarct size-limiting effect at reperfusion that is mediated by HIF-1α and downregulation of its degrading enzyme PHD3. NEW & NOTEWORTHY The present study shows that EET-B is an effective agonistic 14,15-epoxyeicosatrienoic acid analog, and its administration before reperfusion markedly reduced myocardial infarction in rats. Most importantly, we demonstrate that increased hypoxia-inducible factor-1α levels play a role in cardioprotection mediated by EET-B in reperfusion likely by mechanisms including downregulation of the hypoxia-inducible factor -1α-degrading enzyme prolyl hydroxylase domain protein 3.

Systemic lupus erythematosus (SLE) is a chronic autoimmune inflammatory disorder that causes life threatening renal disease and current therapies are limited with serious side-effects. CYP epoxygenase metabolites of arachidonic acid epoxyeicosatrienoic acids (EETs) demonstrate strong anti-inflammatory and kidney protective actions. We investigated the ability of an orally active EET analog, EET-A to prevent kidney injury in a mouse SLE model. Twenty-weeks old female NZBWF1 (SLE) and age-matched NZW/LacJ (Non SLE) were treated with vehicle or EET-A (10 mg/kg/d, p.o.) for 14 weeks and urine and kidney tissues were collected at the end of the protocol. SLE mice demonstrated marked renal chemotaxis with 30-60% higher renal mRNA expression of CXC chemokine receptors (CXCR) and CXC chemokines (CXCL) compared to Non SLE mice. In SLE mice, the elevated chemotaxis is associated with 5-15-fold increase in cytokine mRNA expression and elevated inflammatory cell infiltration in the kidney. SLE mice also had elevated BUN, serum creatinine, proteinuria, and renal fibrosis. Interestingly, EET-A treatment markedly diminished renal CXCR and CXCL renal mRNA expression in SLE mice. EET-A treatment also reduced renal TNF-α, IL-6, IL-1β, and IFN-γ mRNA expression by 70-80% in SLE mice. Along with reductions in renal chemokine and cytokine mRNA expression, EET-A reduced renal immune cell infiltration, BUN, serum creatinine, proteinuria and renal fibrosis in SLE mice. Overall, we demonstrate that an orally active EET analog, EET-A prevents renal injury in a mouse model of SLE by reducing inflammation.

The introduction of calcineurin inhibitors (CNI) into clinical practice in the late 1970s transformed organ transplantation and led to significant improvement in acute rejection episodes. However, despite their significant clinical utility, the use of these agents is hampered by the development of hypertension and nephrotoxicity, which ultimately lead to end-stage kidney disease and overt cardiovascular outcomes. There are currently no effective agents to treat or prevent these complications. Importantly, CNI-free immunosuppressive regimens lack the overall efficacy of CNI-based treatments and put patients at risk of allograft rejection. Cytochrome P-450 epoxygenase metabolites of arachidonic acid, epoxyeicosatrienoic acids (EETs), have potent vasodilator and antihypertensive properties in addition to many cytoprotective effects, but their effects on CNI-induced nephrotoxicity have not been explored. Here, we show that PVPA, a novel, orally active analog of 14,15-EET, effectively prevents the development of hypertension and ameliorates kidney injury in cyclosporine-treated rats. PVPA treatment reduced proteinuria and renal dysfunction induced by cyclosporine. PVPA inhibited inflammatory cell infiltration into the kidney and decreased renal fibrosis. PVPA also reduced tubular epithelial cell apoptosis, attenuated the generation of reactive oxygen species, and modulated the unfolded protein response that is associated with endoplasmic reticulum stress. Consistent with the in vivo data, PVPA attenuated cyclosporine-induced apoptosis of NRK-52E cells in vitro. These data indicate that the cytochrome P-450/EET system offers a novel therapeutic strategy to treat or prevent CNI-induced nephrotoxicity.

Arachidonic acid is metabolized to epoxyeicosatrienoic acids (EETs) by CYP epoxygenases, and EETs are kidney protective in multiple pathologies. We determined the ability of an EET analogue, EET-A, to mitigate experimental radiation nephropathy. The kidney expression of the EET producing enzyme CYP2C11 was lower in rats that received total body irradiation (TBI rat) compared with non-irradiated control. At 12 weeks after TBI, the rats had higher systolic blood pressure and impaired renal afferent arteriolar function compared with control, and EET-A or captopril mitigated these abnormalities. The TBI rats had 3-fold higher blood urea nitrogen (BUN) compared with control, and EET-A or captopril decreased BUN by 40–60%. The urine albumin/creatinine ratio was increased 94-fold in TBI rats, and EET-A or captopril attenuated that increase by 60–90%. In TBI rats, nephrinuria was elevated 30-fold and EET-A or captopril decreased it by 50–90%. Renal interstitial fibrosis, tubular and glomerular injury were present in the TBI rats, and each was decreased by EET-A or captopril. We further demonstrated elevated renal parenchymal apoptosis in TBI rats, which was mitigated by EET-A or captopril. Additional studies revealed that captopril or EET-A mitigated renal apoptosis by acting on the p53/Fas/FasL (Fas ligand) apoptotic pathway. The present study demonstrates a novel EET analogue-based strategy for mitigation of experimental radiation nephropathy by improving renal afferent arteriolar function and by decreasing renal apoptosis.

Epoxyeicosatrienoic acids (EETs) contribute to haemodynamics, electrolyte homoeostasis and blood pressure regulation, leading to the concept that EETs can be therapeutically targeted for hypertension. In the present study, multiple structural EET analogues were synthesized based on the EET pharmacophore and vasodilator structure-activity studies. Four EET analogues with 91–119% vasodilatory activity in the isolated bovine coronary artery (EC50: 0.18–1.6 μM) were identified and studied for blood-pressure-lowering in hypertension. Two EET analogues in which the COOH group at carbon 1 of the EET pharmacophore was replaced with either an aspartic acid (EET-A) or a heterocyclic surrogate (EET-X) were administered for 14 days [10 mg/kg per day intraperitoneally (i.p.)]. Both EET-A and EET-X lowered blood pressure in spontaneously hypertensive rats (SHRs) and in angiotensin II (AngII) hypertension. On day 14, the mean arterial pressures in EET analogue-treated AngII-hypertensive and SHRs were 30–50 mmHg (EET-A) and 15–20 mmHg (EET-X) lower than those in vehicle-treated controls. These EET analogues (10 mg/kg per day) were further tested in AngII hypertension by administering orally in drinking water for 14 days and EET-A lowered blood pressure. Additional experiments demonstrated that EET-A inhibits epithelial sodium channel (ENaC) activity in cultured cortical collecting duct cells and reduced renal expression of ENaC subunits in AngII hypertension. In conclusion, we have characterized EET-A as an orally active antihypertensive EET analogue that protects vascular endothelial function and has ENaC inhibitory activity in AngII hypertension.

Cytochrome P450 (CYP) monooxygenases epoxidize the omega-3 polyunsaturated fatty acid (PUFA) docosahexaenoic acid into novel epoxydocosapentaenoic acids (EDPs) that have multiple biological actions. The present study determined the ability of the most abundant EDP regioisomer, 19,20-EDP to reduce kidney injury in an experimental unilateral ureteral obstruction (UUO) renal fibrosis mouse model. Mice with UUO developed kidney tubular injury and interstitial fibrosis. UUO mice had elevated kidney hydroxyproline content and five-times greater collagen positive fibrotic area than sham control mice. 19,20-EDP treatment to UUO mice for 10 days reduced renal fibrosis with a 40%-50% reduction in collagen positive area and hydroxyproline content. There was a six-fold increase in kidney α-smooth muscle actin (α-SMA) positive area in UUO mice compared to sham control mice, and 19,20-EDP treatment to UUO mice decreased α-SMA immunopositive area by 60%. UUO mice demonstrated renal epithelial-to-mesenchymal transition (EMT) with reduced expression of the epithelial marker E-cadherin and elevated expression of multiple mesenchymal markers (FSP-1, α-SMA, and desmin). Interestingly, 19,20-EDP treatment reduced renal EMT in UUO by decreasing mesenchymal and increasing epithelial marker expression. Overall, we demonstrate that a novel omega-3 fatty acid metabolite 19,20-EDP, prevents UUO-induced renal fibrosis in mice by reducing renal EMT.

Vascular endothelial growth factor inhibitors (VEGFi) are known to cause hypertension and renal injury that severely limits their use as an anticancer therapy. We hypothesized that the angiotensin-converting enzyme inhibitor captopril not only prevents hypertension, but also decreases renal injury caused by the VEGFi sorafenib. Rats were administered sorafenib (20 mg/kg per day) alone or in combination with captopril (40 mg/kg per day) for 4 weeks. Sorafenib administration increased blood pressure, which plateaued by day 10. Concurrent treatment with captopril for 4 weeks resulted in a 30 mmHg decrease in blood pressure compared with sorafenib alone (155 ± 5 vs 182 ± 6 mmHg, respectively; P

Cu(II) and Zn(II) complexes of Schiff bases obtained by condensation of sulfamethoxazole with salicylaldehyde/pyridoxal were prepared and characterized by microanalytical, thermogravimetric, magnetic, and spectroscopic data. The Cu (II) and Zn (II) complexes of salicylidenesulfamethoxazole were found to be five coordinate while all others were six coordinate. The electron paramagnetic resonance spectral lines exhibited rhombic distortion from axial symmetry, with g ‖ > g ⊥ > g e, in the case of the Cu (II) complexes. The geometry of the complexes appears to be square–pyramidal or octahedral. All the compounds under investigation possess antibacterial activity against the strains tested. The antibacterial activity showed the following trend: Cu (II) complexes > Zn (II) complexes > Schiff base ligands > parent drugs. All the copper complexes were found to be active against kaolin paw edema whereas the parent drugs were inactive.

Cerebral arterial myogenic and autoregulatory responses are impaired in Fawn Hooded hypertensive (FHH) rats. Cerebral autoregulatory responses are restored in the congenic rat strain in which a segment of chromosome 1 from the Brown Norway (BN) rat was transferred into the FHH genetic background (FHH.1BN). The impact of this region on cerebral arterial dilator responses remains unknown. Aminopeptidase is a gene that was transferred into the FHH genetic background to generate the FHH.1BN rats and is responsible for degradation of the vasodilator bradykinin. Thus, we hypothesized that FHH rats will have increased aminopeptidase P levels with impaired cerebral arterial responses to bradykinin compared to BN and FHH.1BN rats. We demonstrated higher cerebral arterial expression of aminopeptidase P in FHH compared to BN rats. Accordingly, we demonstrated markedly impaired cerebral arterial dilation to bradykinin in FHH compared to BN rats. Interestingly, aminopeptidase P expression was lower in FHH.1BN compared to FHH rats. Decreased aminopeptidase P levels in FHH.1BN rats were associated with increased cerebral arterial bradykinin-induced dilator responses. Aminopeptidase P inhibition by apstatin improved cerebral arterial bradykinin dilator responses in FHH rats to a level similar to FHH.1BN rats. Unlike bradykinin, cerebral arterial responses to acetylcholine were similar between FHH and FHH.1BN groups. These findings indicate decreased bradykinin bioavailability contributes to impaired cerebral arterial dilation in FHH rats. Overall, these data indicate an important role of aminopeptidase P in the impaired cerebral arterial function in FHH rat.

Epoxyeicosatrienoic acids (EETs) contribute to blood pressure regulation leading to the concept that EETs can be therapeutically targeted for hypertension and the associated end organ damage. In the present study, we investigated anti-hypertensive and kidney protective actions of an EET analog, EET-B in angiotensin II (ANG II)-induced hypertension. EET-B was administered in drinking water for 14 days (10 mg/kg/d) and resulted in a decreased blood pressure elevation in ANG II hypertension. At the end of the two-week period, blood pressure was 30 mmHg lower in EET analog-treated ANG II hypertensive rats. The vasodilation of mesenteric resistance arteries to acetylcholine was impaired in ANG II hypertension; however, it was improved with EET-B treatment. Further, EET-B protected the kidney in ANG II hypertension as evidenced by a marked 90% decrease in albuminuria and 54% decrease in nephrinuria. Kidney histology demonstrated a decrease in renal tubular cast formation in EET analog-treated hypertensive rats. In ANG II hypertension, EET-B treatment markedly lowered renal inflammation. Urinary monocyte chemoattractant protein-1 excretion was decreased by 55% and kidney macrophage infiltration was reduced by 52% with EET-B treatment. Overall, our results demonstrate that EET-B has anti-hypertensive properties, improves vascular function, and decreases renal inflammation and injury in ANG II hypertension.

Nephrotoxicity severely limits the use of the anticancer drug cisplatin. Oxidative stress, inflammation, and endoplasmic reticulum (ER) stress contribute to cisplatin-induced nephrotoxicity. We developed novel orally active epoxyeicosatrienoic acid (EET) analogs and investigated their prophylactic effect in cisplatin-induced nephrotoxicity in rats. Cisplatin-induced nephrotoxicity was manifested by increases in blood urea nitrogen, plasma creatinine, urinary N-acetyl-β-(d)-glucosaminidase activity, kidney injury molecule 1, and histopathology. EET analogs (10 mg/kg/d) attenuated cisplatin-induced nephrotoxicity by reducing these renal injury markers by 40–80% along with a 50–70% reduction in renal tubular cast formation. This attenuated renal injury is associated with reduced oxidative stress, inflammation, and ER stress evident from reduction in related biomarkers and in the renal expression of genes involved in these pathways. Moreover, we demonstrated that the attenuated nephrotoxicity correlated with decreased apoptosis that is associated with 50–90% reduction in Bcl-2 protein family mediated proapoptotic signaling, reduced renal caspase-12 expression, and a 50% reduction in renal caspase-3 activity. We further demonstrated in vitro that the protective activity of EET analogs does not compromise the anticancer effects of cisplatin. Collectively, our data provide evidence that EET analogs attenuate cisplatin-induced nephrotoxicity by reducing oxidative stress, inflammation, ER stress, and apoptosis without affecting the chemotherapeutic effects of cisplatin.—Khan, Md. A. H., Liu, J., Kumar, G., Skapek, S. X., Falck, J. R., Imig, J. D. Novel orally active epoxyeicosatrienoic acid (EET) analogs attenuate cisplatin nephrotoxicity.

Cardiometabolic syndrome occurs with obesity and consists of pathophysiological factors that increase the risk for cardiovascular events. Soluble epoxide hydrolase inhibition (sEHi) is a novel therapeutic approach that exerts renal and cardiovascular protection. Although sEHi as a therapeutic approach is promising, it could be more effective for the treatment of cardiometabolic syndrome when combined with peroxisome proliferator activated receptor γ (PPARγ) agonists. We hypothesized that the PPARγ agonist, rosiglitazone in combination with a sEHi (tAUCB) will provide synergistic actions to decrease blood pressure, improve vascular function, decrease inflammation, and prevent renal damage in spontaneously hypertensive obese rats (SHROB). SHROB were treated with rosiglitazone, tAUCB or the combination of tAUCB and rosiglitazone for four-weeks and compared with spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats. Blood pressure increased in SHROB (164±7 mmHg) and decreased 10 mmHg when treated with rosiglitazone, tAUCB, or tAUCB and rosiglitazone. Mesenteric artery dilation to the KATP channel opener pinacidil was attenuated in SHROB (EMax =77±7%), compared with WKY (EMax =115±19) and SHR (EMax = 93±12%). Vasodilation to pinacidil was improved by rosiglitazone (EMax = 92±14%) but not tAUCB. Renal macrophage infiltration increased in SHROB and significantly decreased with rosiglitazone or tAUCB and rosiglitazone treatment. Albuminuria was increased in SHROB (90±20 mg/d) and was significantly decreased by the combination of tAUCB and rosiglitazone (37±9 mg/d). Glomerular injury in SHROB was also significantly decreased by tAUCB and rosiglitazone. These results indicate that even though sEHi or PPARγ agonist have benefits when used individually, the combination is more beneficial for the multidisease features in cardiometabolic syndrome.

The present study explored the hypothesis that a prolonged 8 weeks exposure to a high fructose intake suppresses adrenergic and angiotensin II (Ang II)-mediated vasoconstriction and is associated with a higher contribution of α1D-adrenoceptors. A total of thirty-two Sprague–Dawley rats received either 20 % fructose solution (FFR) or tap water (control, C) to drink ad libitum for 8 weeks. Metabolic and haemodynamic parameters were assessed weekly. The renal cortical vasoconstrictor responses to noradrenaline (NA), phenylephrine (PE), methoxamine (ME) and Ang II were determined in the presence and absence of BMY7378 (α1D-adrenoceptor antagonist). FFR had increased blood pressure, plasma levels of glucose, TAG and insulin. FFR expressed reduced renal vascular responses to adrenergic agonists and Ang II (NA: 50 %, PE: 50 %, ME, 65 %, Ang II: 54 %). Furthermore in the C group, the magnitude of the renal cortical vasoconstriction to all agonists was blunted in the presence of the low or high dose of BMY7378 (NA: 30 and 31 %, PE: 23 and 33 %, ME: 19 and 44 %, Ang II: 53 and 77 %), respectively, while in the FFR, vasoconstriction was enhanced to adrenergic agonists and reduced to Ang II (NA: 8 and 83 %, PE: 55 %, ME, 2 and 177 %, Ang II: 61 and 31 %). Chronic high fructose intake blunts vascular sensitivity to adrenergic agonists and Ang II. Moreover, blocking of the α1D-adrenoceptor subtype results in enhancement of renal vasoconstriction to adrenergic agonists, suggesting an inhibitory action of α1D-adrenoceptors in the FFR. α1D-Adrenoceptors buffer the AT1-receptor response in the renal vasculature of normal rats and fructose feeding suppressed this interaction.

Type-2 diabetes mellitus (DM-2) is a conformational disease involving intrinsically disordered islet amyloid polypeptide (IAPP), in which a structural transition from physiological polypeptide to pathological deposits takes place. Different factors acquired or inherited, contribute to endoplasmic reticulum stress and/or impair mitochondrial function which leads to conformational changes in IAPP intermediates and ultimately produces oligomers of an anti-parallel crossed β-pleated sheets that eventually accumulate as space-occupying lesions within the islets. Clusters of IAPP monomers form a pore which is linked to channel-like behavior in planar bilayers, indicating that these oligomeric IAPP pores could become incorporated into membranes and alter its barrier properties. Identification of nucleating residues and the residues responsible for this oligomeric tendency could improve understanding of structure–function relationships as well as the molecular mechanism of folding and aggregation of IAPP contributing to the onset of DM-2. A combination of biological, chemical or physical approaches is required to be extensively pursued for the development of a successful anti-amyloidogenic agent to prevent this malady. Exploring the hypothesis of π-stacking may be a better option to control IAPP aggregation if researchers go through the mechanism of π–π interaction, which provides entropy driven energy and direction for self-assembly to control amyloidogenic aggregation.

This paper reports the changes in various physical properties associated with the derivatization of sulfamethoxazole. The properties studied include moisture content, crystallanity, particle size distribution, porosity, flow, compressibility and compactability. It was found that the derivatives, salicylidene-sulfamethoxazole-Zn(II) • H₂O and salicylidene-sulfamethoxazole-Cu(II) • H₂O are crystalline substances. The moisture content was found to be highest in salicylidene-sulfamethoxazole-Zn(II) • H₂O followed by salicylidene-sulfamethoxazole-Cu(II) • H₂O and sulfamethoxazole. The copper complex contained only chemically bonded water, whereas the zinc complex contained both bonded and absorbed water, which affected the strength of the tablets prepared from the three materials accordingly. The particle size decreased on derivatization and complexation with metal ions and the trend was: sulfamethoxazole salicylidene-sulfamethoxazole-Cu(II) • H₂O salicylidene-sulfamethoxazole-Zn(II) • H₂O. This trend was represented by better compactability shown by salicylidene-sulfamethoxazole-Cu(II) • H₂O and salicylidene-sulfamethoxazole-Zn(II) • H₂O as compared with sulfamethoxazole. Salicylidene-sulfamethoxazole-Zn(II) • H₂O had the highest porosity followed by salicylidene-sulfamethoxazole-Cu(II) • H₂O, and sulfamethoxazole; this resulted in better compressibility behavior of the complexes. Thus it was observed that salicylidene-sulfamethoxazole-Cu(II) • H₂O and salicylidene-sulfamethoxazole-Zn(II) • H₂O formed stronger compacts. The values of angle of repose and flow rate show better flow properties for salicylidene-sulfamethoxazole-Cu(II) • H₂O as compared with sulfamethoxazole and salicylidene-sulfamethoxazole-Zn(II) • H₂O. It was concluded that derivatization substantially changed the pharmaceutical properties, which have important role to play in formulation of solid dosage form.

The mechanisms involved in the initiation of human labor are largely unknown. Understanding the molecular pathways is fundamental in both the development of effective therapeutic strategies and intervention to prevent preterm labor. Prostaglandins are bioactive lipids and members of the eicosanoids family, derived from arachidonic acid, which act in a paracrine or autocrine manner and function via binding to specific G-protein-coupled receptors, activating intracellular signaling and gene transcription. Prostaglandins have a central role in the maintenance of pregnancy and initiation of labor, with the change from uterine quiescence to a contractile state facilitated by differential expression of prostaglandin receptors within the myometrium and fetal membranes. Clinical evidence for the key role of prostaglandins in human parturition is evident from their successful exploitation as exogenous agents for the induction of labor and the role of prostaglandin synthase inhibitors as a preventative therapy for preterm labor. This review aims to focus on prostaglandin synthesis and metabolism and how differential regulation of prostaglandins and their receptors in gestational tissues interact in the initiation of labor.

Prostaglandins are bioactive lipids and important mediators of uterine relaxation as well as contraction during pregnancy and labour. E series prostaglandins may directly contract or relax myometrium in a dose-dependent manner, with the relaxatory effects mediated through the prostanoid receptors EP(2) and EP(4). The aim of this study was to evaluate the pharmacological effects of prostaglandin analogues on isolated pregnant rat uterine contractility, at 10(-15) to 10(-9) M concentrations. Uterine strips from rats at 19 days of gestation were set up in organ baths at 37 degrees C, bathed in Krebs buffer and gassed with 95% O(2)/5% CO(2). Spontaneous contractions were recorded via a force transducer. Concentration ranges of 10(-15)-10(-9) M of PGE(2), PGF(2alpha) and a range of prostaglandin analogues were applied non-cumulatively to the tissues. Spontaneous contractions were recorded for 12 min post dose. Amplitude, frequency, baseline tone and percent contractility over 10 min periods were analysed. PGE(2), butaprost, 9-keto fluprostenol, 11-keto fluprostenol, 9-keto fluprostenol isopropyl ester, AL8810 and 15(S)-15-methyl PGE(2) all caused a decrease in percent contractility (P

The characteristics of carnosine (beta-alanyl-L-histidine) transport have been studied using purified brush-border membrane vesicles from mouse small intestine. Uptake curves did not exhibit any overshoot phenomena, and were similar under Na+, K+ or choline+ gradient conditions (extravesicular greater than intravesicular). However, uptake of histidine showed an overshoot phenomenon in the presence of a Na+-gradient. There was no detectable hydrolysis of carnosine during 15 min of incubation with membrane vesicles under conditions used for transport experiments. Analysis of intravesicular contents further showed the complete absence of the constituent free amino acids of carnosine, and indicates that intact carnosine is transported. Studies on the effect of concentration on peptide uptake revealed that transport occurred by a saturable process conforming to Michaelis-Menten kinetics with a Km of 9.6 +/- 1.4 mM and a Vmax of 2.9 +/- 0.2 nmol/mg protein per 0.4 min. Uptake of carnosine was inhibited by both di- and tripeptides with a maximum inhibition of 68% by glycyl-L-leucyltyrosine. These results clearly demonstrate that carnosine is transported intact by a carrier-mediated, Na+-independent process.

This study characterizes the transport of [1-14C]glycyl-L-proline into purified brush border membrane vesicles prepared from human small intestine. Time-course uptake curves of glycyl-L-proline were similar under sodium thiocyanate or potassium thiocyanate gradient conditions (extravesicular greater than intravesicular) and did not show any overshoot phenomena. The transport of glycine and proline, however, was stimulated by the presence of sodium gradient. Measurement of peptide uptake with increasing medium osmolarity showed that glycyl-L-proline was transported into an osmotically reactive intravesicular space with insignificant binding to the surface of the vesicles. Only 2% of the glycyl-L-proline in the incubation media was hydrolyzed after 10 min of incubation. Also, there was no hydrolysis of peptide transported into the intravesicular space. The effects of increasing concentrations of glycyl-L-proline on uptake showed that uptake of the peptide was saturable and conformed to Michaelis-Menten kinetics with a Km of 4.1 +/- 0.5 mM and a Vmax of 1.53 +/- 0.07 nmol/mg protein X 0.5 min. Free amino acids did not inhibit the transport of glycyl-L-proline while dipeptides and tripeptides exerted appreciable inhibition (up to 60%). Our results show that human small intestinal brush border membrane vesicles transport glycyl-L-proline as an intact peptide by a carrier-mediated, Na+-independent process.

760 Background: Somatic and germline DDR gene alterations in PC have been postulated to positively predict response to DNA damaging cytotoxic agents. Due to the relatively high prevalence of germline DDR gene alterations, germline testing is recommended in all pts with PC. We examined whether somatic CGP can be used to reliably identify PC pts that merit germline testing. Methods: We retrospectively reviewed the electronic medical records of PC pts who underwent both somatic CGP (utilizing the Foundation One assay) and germline testing. DDR gene mutations were categorized as somatic-pathogenic, somatic-variant of uncertain significance (VUS), germline-pathogenic and germline-VUS. For somatic testing, DNA was extracted from formalin fixed paraffin embedded (FFPE) clinical specimens and CGP was done on hybrid-capture, adaptor ligation based libraries to a mean coverage depth of > 600 for up to 315 genes plus 47 introns from 19 genes frequently rearranged in cancer. Germline genetic testing was performed on submitted blood or saliva samples, utilizing commercial assays; next generation or Sanger sequencing of all coding regions and adjacent intronic nucleotides were performed. Results: Ninety-three pts had somatic CGP data, 51 (55%) pts had both somatic CGP and germline data available. Among the 51 pts with both germline and somatic data available, DDR gene alterations that were somatic-pathogenic, germline-pathogenic, somatic-VUS and germline-VUS were present in 7 (13.7%), 7 (13.7%), 23 (45.1%) and 16 (31.4%) pts, respectively. Of the 7 pts with somatic-pathogenic alterations, 5 (71%) had a concordant germline alteration and of the 7 pts with germline-pathogenic alterations, 5 (71%) had a concordant somatic alteration. Of the 23 pts with somatic-VUSs, 12 (52%) had a concordant germline VUS and of the 16 pts with germline-VUSs, 12 (75%) had a concordant somatic VUS. Conclusions: Both somatic and germline DDR gene alterations are common in PC pts. Despite the relatively high concordance rate between somatic and germline pathogenic DDR gene alterations, somatic CGP will miss approximately one fourth of the germline DDR gene alterations.