Kidney Injury by Unilateral Ureteral Obstruction in Mice Lacks Sex Differences.

INTRODUCTION: Renal fibrosis is a critical event in the development and progression of chronic kidney disease (CKD), and it is considered the final common pathway for all types of CKD. The prevalence of CKD is higher in females; however, males have a greater prevalence of end-stage renal disease. In addition, low birth weight and low nephron number are associated with increased risk for CKD. This study examined the development and severity of unilateral ureter obstruction (UUO)-induced renal fibrosis in male and female wild-type (ROP +/+) and mutant (ROP Os/+) mice, a mouse model of low nephron number. METHODS: Male and female ROP +/+ and ROP Os/+ mice were subjected to UUO, and kidney tissue was collected at the end of the 10-day experimental period. Kidney histological analysis and mRNA expression determined renal fibrosis, tubular injury, collagen deposition, extracellular matrix proteins, and immune cell infiltration. RESULTS: Male and female UUO mice demonstrated marked renal injury, kidney fibrosis, and renal extracellular matrix production. Renal fibrosis and α-smooth muscle actin were increased to a similar degree in ROP +/+ and ROP Os/+ mice with UUO of either sex. There were also no sex differences in renal tubular cast formation or renal infiltration of macrophage in ROP +/+ and ROP Os/+ UUO mice. Interestingly, renal fibrosis and α-smooth muscle actin were 1.5-3-fold greater in UUO-ROP +/+ compared to UUO-ROP Os/+ mice. Renal inflammation phenotypes following UUO were also 30-45% greater in ROP +/+ compared to ROP Os/+ mice. Likewise, expression of extracellular matrix and renal fibrotic genes was greater in UUO-ROP +/+ mice compared to UUO-ROP Os/+ mice. In contrast to these findings, ROP Os/+ mice with UUO demonstrated glomerular hypertrophy with 50% greater glomerular tuft area compared to ROP +/+ with UUO. Glomerular hypertrophy was not sex-dependent in any of the genotypes of ROP mice. These findings provide evidence that low nephron number contributes to UUO-induced glomerular hypertrophy in ROP Os/+ mice but does not enhance renal fibrosis, inflammation, and renal tubular injury. CONCLUSION: Taken together, we demonstrate that low nephron number contributes to enhanced glomerular hypertrophy but not kidney fibrosis and tubular injury. We also demonstrate that none of the changes caused by UUO was affected by sex in any of the ROP mice genotypes.

A SORAFENIB INDUCED MODEL OF GLOMERULAR KIDNEY DISEASE.

Glomerular injury and proteinuria are important pathophysiological features of chronic kidney disease. In the present study, we provide data on a glomerular injury model that was developed using the cancer chemotherapy drug sorafenib. Sorafenib is a tyrosine kinase inhibitor that acts via the vascular endothelial growth factor (VEGF) signaling pathway and is widely used to treat a variety of cancers. On the other hand, sorafenib causes serious renal side effects in patients including the development of chronic kidney disease. The current study aimed to utilize the nephrotoxic property of sorafenib to develop a rat model for chronic kidney disease. We demonstrate that rats administered sorafenib for 8 weeks along with a high salt diet (8% NaCl enriched) develop hypertension (80mmHg higher systolic blood pressure), proteinuria (75% higher), and 4-fold higher glomerular injury compared to vehicle-treated normal control rat. Sorafenib induced glomerular injury was associated with decreased (20-80% lower) renal mRNA expression of key glomerular structural proteins such as nephrin, podocin, synaptopodin, and podoplanin compared to vehicle-treated normal control rat. Renal cortical endothelial-to-mesenchymal transition (EndoMT) was activated in the sorafenib induced glomerular injury model. In the sorafenib treated rats, the renal EndoMT was evident with 20% lower mRNA expression of an endothelial marker WT-1 and 2 to 3-fold higher expression of mesenchymal markers Col III, FSP-1, α-SMA, and vimentin. In conclusion, we developed a rat pre-clinical chronic kidney disease model that manifest glomerular injury. We further demonstrate that the glomerular injury in this model is associated with decreased renal mRNA expression of key glomerular structural proteins and an activated kidney EndoMT.

THE EFFECT OF COMPOUND DM509 ON KIDNEY FIBROSIS IN THE CONDITIONS OF THE EXPERIMENTAL MODEL.

Renal fibrosis is a critical event in the progression of chronic kidney disease (CKD) to end-stage renal disease (ESRD). Unfortunately, there are few options to target renal fibrosis in order to develop novel anti-fibrotic agents that could prevent CKD progression to ESRD. We evaluated the efficacy of a novel dual-acting molecule, DM509, in preventing renal fibrosis using the unilateral ureteral obstruction (UUO) renal fibrosis mouse model. DM509 acts simultaneously as a farnesoid X receptor agonist (FXRA) and a soluble epoxide hydrolase inhibitor (sEHi). In this study, groups of 8-12 weeks old C57BL/6J male mice went through either UUO or sham surgery (n=6/group). Mice were pre-treated with DM509 (10mg/kg/d) or vehicle administered in drinking water one day prior to the UUO surgery. Sham, vehicle and DM509 treatments continued until day 10 and blood and kidney tissue were collected for biochemical, histological, and gene expression analysis at the end of the treatment protocol. The UUO group exhibited kidney dysfunction with elevated blood urea nitrogen (BUN) compared to the sham group (63±7 vs. 34±6 mg/dL). DM509 treatment prevented renal dysfunction as evident from 36% lower BUN level in the DM509 treated UUO mice compared to UUO mice treated with vehicle. Vehicle treated UUO mice demonstrated renal fibrosis with elevated kidney hydroxyproline content (213±11 vs. 49±9 µg/mg protein) and kidney collagen positive area (13±2% vs. 1.1±0.1%) compared to the sham group. We found that DM509 treatment prevented renal fibrosis and DM509 treated mice had 34-66% lower levels of kidney hydroxyproline and collagen positive renal area compared to vehicle-treated UUO mice. In conclusion, our data provide evidence that the novel dual-acting FXRA and a sEHi, DM509, prevented renal dysfunction and renal fibrosis in UUO mouse model.

Delayed Effects of Acute Radiation Exposure (Deare) in Juvenile and Old Rats: Mitigation by Lisinopril.

Our goal is to develop lisinopril as a mitigator of delayed effects of acute radiation exposure in the National Institute of Allergy and Infectious Diseases program for radiation countermeasures. Published studies demonstrated mitigation of delayed effects of acute radiation exposure by lisinopril in adult rats. However, juvenile or old rats beyond their reproductive lifespans have never been tested. Since no preclinical models of delayed effects of acute radiation exposure were available in these special populations, appropriate rat models were developed to test lisinopril after irradiation. Juvenile (42-d-old, prepubertal) female and male WAG/RijCmcr (Wistar) rats were given 13-Gy partial-body irradiation with only part of one hind limb shielded. Lethality from lung injury between 39-58 d and radiation nephropathy between 106-114 d were recorded. All irradiated-only juvenile rats were morbid from delayed effects of acute radiation exposure by 114 d, while lisinopril (24 mg m d) started 7 d after irradiation and continued improved survival to 88% (p = 0.0015, n ≥ 8/group). Old rats (>483-d-old, reproductively senescent) were irradiated with 13-Gy partial-body irradiation keeping part of one leg shielded and additionally shielding the head in some animals. Irradiated old females developed lethal nephropathy, and all became morbid by 170 d after irradiation, though no rats displayed lethal radiation pneumonitis. Similar results were observed for irradiated geriatric males, though 33% of rats remained alive at 180 d after irradiation. Lisinopril mitigated radiation nephropathy in old rats of both sexes. Finally, comparison of delayed effects of acute radiation exposure between irradiated juvenile, adult, and old rats showed younger rats were more sensitive to delayed effects of acute radiation exposure with earlier manifestation of injuries to some organs.

Epoxyeicosatrienoic acid analog attenuates angiotensin II hypertension and kidney injury.

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.

Telmisartan provides better renal protection than valsartan in a rat model of metabolic syndrome.

BACKGROUND: Angiotensin receptor blockers (ARB), telmisartan, and valsartan were compared for renal protection in spontaneously hypertensive rats (SHR) fed high fat diet. We hypothesized that in cardiometabolic syndrome, telmisartan an ARB with peroxisome proliferators activated receptor-γ (PPAR-γ) activity will offer better renal protection. METHODS: SHR were fed either normal (SHR-NF, 7% fat) or high fat (SHR-HF, 36% fat) diet and treated with an ARB for 10 weeks. RESULTS: Blood pressure was similar between SHR-NF (190 ± 3 mm Hg) and SHR-HF (192 ± 4 mm Hg) at the end of the 10 week period. Telmisartan and valsartan decreased blood pressure to similar extents in SHR-NF and SHR-HF groups. Body weight was significantly higher in SHR-HF (368 ± 5 g) compared to SHR-NF (328 ± 7 g). Telmisartan but not valsartan significantly reduced the body weight gain in SHR-HF. Telmisartan was also more effective than valsartan in improving glycemic and lipid status in SHR-HF. Monocyte chemoattractant protein-1 (MCP-1), an inflammatory marker, was higher in SHR-HF (24 ± 2 ng/d) compared to SHR-NF (14 ± 5 ng/d). Telmisartan reduced MCP-1 excretion in both SHR-HF and SHR-NF to a greater extent than valsartan. An indicator of renal injury, urinary albumin excretion increased to 85 ± 8 mg/d in SHR-HF compared to 54 ± 9 mg/d in SHR-NF. Telmisartan (23 ± 5 mg/d) was more effective than valsartan (45 ± 3 mg/d) in lowering urinary albumin excretion in SHR-HF. Moreover, telmisartan reduced glomerular damage to a greater extent than valsartan in the SHR-HF. CONCLUSIONS: Collectively, our data demonstrate that telmisartan was more effective than valsartan in reducing body weight gain, renal inflammation, and renal injury in a rat model of cardiometabolic syndrome.

Effect of renal sympathetic nerve on adrenergically and angiotensin II-induced renal vasoconstriction in normal Wistar-Kyoto rats.

BACKGROUND: This study examined the effect of renal sympathetic innervation on adrenergically and angiotensin II (Ang II)-induced renal vasoconstriction in Wistar-Kyoto (WKY) rats. METHODS: Forty-eight WKY rats were treated with either losartan (10 mg/kg/day p.o.) or carvedilol (5 mg/kg/day p.o.) or a combination of them (10 mg/kg/day + 5 mg/kg/day p.o.) for 7 days. On day 8, the rats were anaesthetized, and renal vasoconstrictor experiments were carried out. A group of rats was subjected to acute unilateral renal denervation during the acute study. Changes in the renal vasoconstrictor responses were determined in terms of reductions in renal blood flow caused by Ang II, noradrenaline (NA), and methoxamine (ME). RESULTS: In normal animals, losartan decreased (P < 0.05) the renal vasoconstrictor response to Ang II but not to NA or ME. Carvedilol treatment, however, blunted (P < 0.05) the renal vasoconstrictor responses to Ang II and adrenergic agonists. Combination of losartan and carvedilol blunted (P < 0.05) the renal vasoconstrictor response to Ang II but augmented the responses to NA and ME (all P < 0.05). Interestingly, when denervated rats were treated with the same combination, there was a reduction (P < 0.05) in the renal vasoconstrictor responses to Ang II and adrenergic agonists. CONCLUSIONS: Data suggest that the renal sympathetic nerve contributes to adrenergic agonist-mediated renal vasoconstrictions in normal rats. The data further indicate an interactive relationship between renin-angiotensin and sympathetic nervous systems in modulating adrenergically and Ang II-induced renal vasoconstriction in WKY rats.

Role of the renal sympathetic nervous system in mediating renal ischaemic injury-induced reductions in renal haemodynamic and excretory functions.

AIM: We investigated the role of renal sympathetic innervation in the deterioration of renal haemodynamic and excretory functions during the early post-ischaemic phase of renal ischaemia/reperfusion injury. METHODS: Anaesthetised male Sprague-Dawley rats were subjected to unilateral renal ischaemia by clamping the left renal artery for 30 min followed by reperfusion. Following acute renal denervation clearance experiments were performed. In a different set of experiments, the renal nerves were electrically stimulated at increasing frequencies and responses in renal blood flow and renal vascular resistance were recorded. RESULTS: Denervated post-ischaemic acute renal failure (ARF) rats showed higher urine flow rate, absolute and fractional sodium excretions, urinary sodium to urinary potassium, glomerular filtration rate and basal renal blood flow but lower basal renal vascular resistance (all p < 0.05 vs innervated ARF rats). Potassium excretion was significantly lower in denervated group as per fractional (p < 0.05 vs innervated ARF rats) but not absolute potassium excretion (p > 0.05 vs innervated ARF rats). The rise in mean arterial pressure and renal vasoconstrictor response to renal nerve stimulation were blunted in denervated ischaemic ARF rats (all p < 0.05 vs innervated ARF rats). Renal histopathology in denervated ARF rats manifested a significantly lower medullary congestion, inflammation and tubular injury compared to innervated counterparts (p < 0.05 vs innervated ARF rats). CONCLUSIONS: The findings strongly suggest the involvement of renal sympathetic tone in the post-ischaemic events of ischaemic ARF, as the removal of its action to a degree ameliorated the post-ischaemic renal dysfunctions.

Effect of TGF-beta1 antisense oligodeoxynucleotide on renal function in chronic renal failure rats.

AIM: The aim of the present study was to investigate the effectiveness of transforming growth factor (TGF)-beta1 antisense oligodeoxynucleotides (ODN) in ameliorating deteriorated kidney function in rats with puromycin-induced chronic renal failure (CRF). METHODS: Saline, puromycin, puromycin+TGF-beta1 antisense ODN or puromycin+scrambled ODN were administered to unilaterally nephrectomized rats. Renal hemodynamic and excretory measurements were taken in the anaesthetized rats that had undergone surgical procedure. RESULTS: It was observed that in the CRF rats, there was a marked reduction in the renal blood flow (RBF), glomerular filtration rate (GFR), severe proteinuria, and almost 6-fold increased fractional excretion of sodium (FE Na+) as compared to that in the control rats (all P<0.05). It was further observed that in the CRF rats, the treatment with TGF-beta1 antisense, but not scrambled ODN, markedly attenuated the reduction of RBF, GFR, and proteinuria and markedly prevented the increase of the FE Na+ (all P<0.05). In addition, the renal hypertrophy in the CRF group (P<0.05 vs non-renal failure control) was markedly attenuated after treatment with TGF-1 antisense ODN (P<0.05). Focal segmental glomerulosclerosis was evident only in the untreated and scrambled ODN-treated CRF groups. An interesting observation of this study was that in the CRF rats, although there was marked attenuating and preventive effects of the TGF-beta1 antisense ODN on the deteriorated renal functions, the antisense treatment did not cause any marked change in the renal expression of TGF-beta1 at the protein level. CONCLUSION: Collectively, the data obtained suggests that TGF-beta1 antisense ODN possesses beneficial effects in puromycininduced chronic renal failure and that the deterioration in morphology and impaired renal function in this pathological state is in part dependent upon the action of TGF-beta1 within the kidney.