Effects of Tissue Pressure on Transgene Expression Characteristics via Renal Local Administration Routes from Ureter or Renal Artery in the Rat Kidney.
ABSTRACT: We previously developed a renal pressure-mediated transfection method (renal pressure method) as a kidney-specific in vivo gene delivery system. However, additional information on selecting other injection routes and applicable animals remains unclear. In this study, we selected renal arterial and ureteral injections as local administration routes and evaluated the characteristics of gene delivery such as efficacy, safety, and distribution in pressured kidney of rat. Immediately after the naked pDNA injection, via renal artery or ureter, the left kidney of the rat was pressured using a pressure controlling device. Transfection efficiency of the pressured kidney was about 100-fold higher than that of the injection only group in both administration routes. The optimal pressure intensity in the rat kidney was 1.2 N/cm2 for renal arterial injection and 0.9 N/cm2 for ureteral injection. We found that transgene expression site differs according to administration route: cortical fibroblasts and renal tubule in renal arterial injection and cortical and medullary tubule and medullary collecting duct in ureteral injection. This is the first report to demonstrate that the renal pressure method can also be effective, after renal arterial and ureteral injections, in rat kidney.
Project description:BACKGROUND AND OBJECTIVES:Mesenchymal stem cells (MSCs) have been shown to ameliorate cisplatin-induced acute kidney injury (AKI). The present study compares the efficacy of different routes of MSCs administration on kidney damage and regeneration after cisplatin-induced AKI. METHODS:A single intraperitoneal injection of cisplatin (5 mg/kg) was used to induce AKI in 160 rats. MSCs (5×10?) were given by either intravenous, intra-arterial or kidney sub capsular injection one day after cisplatin injection. Suitable control groups were included. Rats were sacrificed at 4, 7, 11 and 30 days after cisplatin injection. Kidney function parameters, kidney tissue oxidative stress markers, and scoring for renal tissue injury, regeneration and chronicity were all determined. RESULTS:MSCs by any routes were able to ameliorate kidney function deterioration and renal tissue damage induced by cisplatin. The overall results of the three routes were equal. Differences between the different routes in one parameter were transient and inconsistent with other parameters. CONCLUSIONS:Changing the route of MSCs injection does not have a major influence on the outcome. Future evaluation should focus on differences between the routes of administration considering the long term safety.
Project description:Experimentally renal tissue hypoxia appears to play an important role in the pathogenesis of chronic kidney disease (CKD) and arterial hypertension (AHT). In this study we measured renal tissue oxygenation and its determinants in humans using blood oxygenation level-dependent magnetic resonance imaging (BOLD-MRI) under standardized hydration conditions. Four coronal slices were selected, and a multi gradient echo sequence was used to acquire T2* weighted images. The mean cortical and medullary R2* values (?=?1/T2*) were calculated before and after administration of IV furosemide, a low R2* indicating a high tissue oxygenation. We studied 195 subjects (95 CKD, 58 treated AHT, and 42 healthy controls). Mean cortical R2 and medullary R2* were not significantly different between the groups at baseline. In stimulated conditions (furosemide injection), the decrease in R2* was significantly blunted in patients with CKD and AHT. In multivariate linear regression analyses, neither cortical nor medullary R2* were associated with eGFR or blood pressure, but cortical R2* correlated positively with male gender, blood glucose and uric acid levels. In conclusion, our data show that kidney oxygenation is tightly regulated in CKD and hypertensive patients at rest. However, the metabolic response to acute changes in sodium transport is altered in CKD and in AHT, despite preserved renal function in the latter group. This suggests the presence of early renal metabolic alterations in hypertension. The correlations between cortical R2* values, male gender, glycemia and uric acid levels suggest that these factors interfere with the regulation of renal tissue oxygenation.
Project description:Activation of the renin-angiotensin system may initiate chronic kidney disease. We hypothesised that renal hypoxia is a consequence of hemodynamic changes induced by angiotensin II and occurs prior to development of severe renal damage. Male Sprague-Dawley rats were infused continuously with angiotensin II (350?ng/kg/min) for 8 days. Mean arterial pressure (n?=?5), cortical (n?=?6) and medullary (n?=?7) oxygenation (pO2) were continuously recorded by telemetry and renal tissue injury was scored. Angiotensin II increased arterial pressure gradually to 150?±?18?mmHg. This was associated with transient reduction of oxygen levels in renal cortex (by 18?±?2%) and medulla (by 17?±?6%) at 10?±?2 and 6?±?1?hours, respectively after starting infusion. Thereafter oxygen levels normalised to pre-infusion levels and were maintained during the remainder of the infusion period. In rats receiving angiotensin II, adding losartan to drinking water (300?mg/L) only induced transient increase in renal oxygenation, despite normalisation of arterial pressure. In rats, renal hypoxia is only a transient phenomenon during initiation of angiotensin II-induced hypertension.
Project description:Sirtuin 1 (Sirt1) is a NAD+-dependent deacetylase that exerts many of the pleiotropic effects of oxidative metabolism. Due to local hypoxia and hypertonicity, the renal medulla is subject to extreme oxidative stress. Here, we set out to investigate the role of Sirt1 in the kidney. Our initial analysis indicated that it was abundantly expressed in mouse renal medullary interstitial cells in vivo. Knocking down Sirt1 expression in primary mouse renal medullary interstitial cells substantially reduced cellular resistance to oxidative stress, while pharmacologic Sirt1 activation using either resveratrol or SRT2183 improved cell survival in response to oxidative stress. The unilateral ureteral obstruction (UUO) model of kidney injury induced markedly more renal apoptosis and fibrosis in Sirt1+/- mice than in wild-type controls, while pharmacologic Sirt1 activation substantially attenuated apoptosis and fibrosis in wild-type mice. Moreover, Sirt1 deficiency attenuated oxidative stress-induced COX2 expression in cultured mouse renal medullary interstitial cells, and Sirt1+/- mice displayed reduced UUO-induced COX2 expression in vivo. Conversely, Sirt1 activation increased renal medullary interstitial cell COX2 expression both in vitro and in vivo. Furthermore, exogenous PGE2 markedly reduced apoptosis in Sirt1-deficient renal medullary interstitial cells following oxidative stress. Taken together, these results identify Sirt1 as an important protective factor for mouse renal medullary interstitial cells following oxidative stress and suggest that the protective function of Sirt1 is partly attributable to its regulation of COX2 induction. We therefore suggest that Sirt1 provides a potential therapeutic target to minimize renal medullary cell damage following oxidative stress.
Project description:BACKGROUND: The objective of this study was to test the effect of removal of a ureteral obstruction (renal calculus) from anesthetized patients on the perfusion index (PI), as measured by a pulse oximeter, and on the estimated glomerular filtration rate (eGFR). PATIENTS AND METHODS: This prospective study enrolled 113 patients with unilateral ureteral obstructions (kidney stones) who were scheduled for ureteroscopy (URS) laser lithotripsy. One urologist graded patient hydronephrosis before surgery. A pulse oximeter was affixed to each patient's index finger ipsilateral to the intravenous catheter, and a non-invasive blood pressure cuff was placed on the contralateral side. Ipsilateral double J stents and Foley catheters were inserted and left indwelling for 24 h. PI and mean arterial pressure (MAP) were determined at baseline, 5 min after anesthesia, and 10 min after surgery; eGFR was determined at admission, 1 day after surgery, and 14 days after surgery. RESULTS: Patients with different grades of hydronephrosis had similar age, eGFR, PI, mean arterial pressure (MAP), and heart rate (HR). PI increased significantly in each hydronephrosis group after ureteral stone disintegration. None of the groups had significant post-URS changes in eGFR, although eGFR increased in the grade I hydronephrosis group after 14 days. The percent change of PI correlates significantly with the percent change of MAP, but not with that of eGFR. CONCLUSION: Our results demonstrate that release of a ureteral obstruction leads to a concurrent increase of PI during anesthesia. Measurement of PI may be a valuable tool to monitor the successful release of ureteral obstructions and changes of microcirculation during surgery. There were also increases in eGFR after 14 days, but not immediately after surgery.
Project description:Sympathetic activation and the kidney play critical roles in hypertension and chronic heart failure. The role of the kidney in sympathetic activation is still not well known. In this study, we revealed an excitatory renal reflex (ERR) in rats induced by chemical stimulation of the kidney that regulated sympathetic activity and blood pressure. The ERR was induced by renal infusion of capsaicin, and evaluated by the changes in renal sympathetic outflow, blood pressure, and heart rate. Renal infusion of capsaicin dose-dependently increased the contralateral renal sympathetic nerve activity, mean arterial pressure, and heart rate. Capsaicin in the cortico-medullary border had greater effects than in the cortex or medulla. Intravenous infusion of capsaicin had no significant effects. The effects of renal infusion of capsaicin were abolished by ipsilateral renal denervation, but were not affected by bilateral sinoaortic denervation. Renal infusion of capsaicin increased the ipsilateral renal afferent activity. The ERR was also induced by renal infusion of bradykinin, adenosine, and angiotensin II, but not by ATP. Renal infusion of capsaicin increased c-Fos expression in the paraventricular nucleus (PVN) of hypothalamus. Lesion of neurons in the PVN with kainic acid abolished the capsaicin-induced ERR. These findings indicate that chemical stimulation of kidney causes an excitatory reflex, leading to sympathetic activation, pressor response, and accelerated heart rate. The PVN is an important central nucleus in the pathway of the ERR.
Project description:Hypoxia inducible factor (HIF)-1alpha, a transcription factor, is abundantly expressed in the renal medulla and regulates many oxygen-sensitive genes such as nitric oxide synthase, cyclooxygenase-2, and heme oxygenase-1. Given the important roles of these genes in the control of arterial pressure, the present study was to test the hypothesis that HIF-1alpha-mediated gene activation serves as an antihypertensive pathway by regulating renal medullary function and sodium excretion. HIF-1alpha decoy oligodeoxynucleotides (ODNs) or scrambled ODNs were transfected into the renal medulla in uninephrectomized Sprague-Dawley rats. Two weeks after ODN transfection, the HIF-1alpha binding activities were significantly inhibited by 45%, and high salt-induced increases of nitric oxide synthase-2 and heme oxygenase-1 transcriptions were also inhibited by 70% and 61% in the renal medulla from decoy rats. The natriuretic responses and increases of renal medullary blood flow responding to the elevations of renal perfusion pressure were significantly blunted by 50% and 37% in decoy rats. Intravenously acute sodium loading increased medullary blood flow and urinary sodium excretion, which was remarkably attenuated in decoy rats. In decoy rats, high salt intake caused a greater positive sodium balance. Consequently, arterial pressure was remarkably increased (from 118+/-1.9 to 154+/-6.3 mm Hg) in decoy rats but not in control rats when the rats were challenged with a high salt diet. There was no blood pressure change in decoy rats that were maintained in normal salt diet. In conclusion, HIF-1alpha-mediated gene activation importantly participates in the regulation of renal medullary function and long-term arterial blood pressure.
Project description:We created a rat renal congestion model and investigated the effect of renal congestion on hemodynamics and molecular mechanisms. The inferior vena cava (IVC) between the renal veins was ligated by suture in male Sprague-Dawley rats to increase upstream IVC pressure and induce congestion in the left kidney only. Left kidney congestion reduced renal blood flow, glomerular filtration rate, and increased renal interstitial hydrostatic pressure. Tubulointerstitial and glomerular injury and medullary thick ascending limb hypoxia were observed only in the congestive kidneys. Molecules related to extracellular matrix expansion, tubular injury, and focal adhesion were upregulated in microarray analysis. Renal decapsulation ameliorated the tubulointerstitial injury. Electron microscopy captured pericyte detachment in the congestive kidneys. Transgelin and platelet-derived growth factor receptors, as indicators of pericyte-myofibroblast transition, were upregulated in the pericytes and the adjacent interstitium. With the compression of the peritubular capillaries and tubules, hypoxia and physical stress induce pericyte detachment, which could result in extracellular matrix expansion and tubular injury in renal congestion. Overall design: Three rats were operated. Three days after surgery, congestive left kidney were compared to control right kidney. Cortex and outer medulla are separately analyzed.
Project description:Increased central venous pressure in congestive heart failure causes renal dysfunction; however, the underlying mechanisms are unclear. We created a rat renal congestion model and investigated the effect of renal congestion on hemodynamics and molecular mechanisms. The inferior vena cava (IVC) between the renal veins was ligated by suture in male Sprague-Dawley rats to increase upstream IVC pressure and induce congestion in the left kidney only. Left kidney congestion reduced renal blood flow, glomerular filtration rate, and increased renal interstitial hydrostatic pressure. Tubulointerstitial and glomerular injury and medullary thick ascending limb hypoxia were observed only in the congestive kidneys. Molecules related to extracellular matrix expansion, tubular injury, and focal adhesion were upregulated in microarray analysis. Renal decapsulation ameliorated the tubulointerstitial injury. Electron microscopy captured pericyte detachment in the congestive kidneys. Transgelin and platelet-derived growth factor receptors, as indicators of pericyte-myofibroblast transition, were upregulated in the pericytes and the adjacent interstitium. With the compression of the peritubular capillaries and tubules, hypoxia and physical stress induce pericyte detachment, which could result in extracellular matrix expansion and tubular injury in renal congestion.
Project description:Conventional methods used for measuring regional renal blood flow, such as laser-Doppler flowmetry, are highly invasive, and each measurement is restricted to a discrete location. The aim of this study was to determine whether ultrasound imaging in conjunction with enhanced contrast agent (microbubbles; Vevo MicroMarker, VisualSonics) could provide a viable noninvasive alternative. This was achieved by determining changes in renal cortical and medullary rate of perfusion in response to a bolus injection of endothelin-1 (ET-1; 0.6, 1.0, or 2.0 nmol/kg) and comparing these responses to those observed in separate groups of mice with conventional laser-Doppler methods. Intravenous infusion of ET-1 in anesthetized male C57bl/6 mice resulted in a dose-dependent increase in mean arterial pressure and a dose-dependent decrease in total renal blood flow as measured by pulse-wave Doppler. ET-1 infusion resulted in a dose-dependent decrease in regional kidney perfusion as measured by both ultrasound with enhanced contrast agent and laser-Doppler measurements, verifying the use of ultrasound to measure regional kidney perfusion. Noted limitations of ultrasound imaging compared with laser-Doppler flowmetry included a lower degree of sensitivity to changes in tissue perfusion and the inability to assess rapid or transient changes in tissue perfusion. In conclusion, ultrasound represents an effective and noninvasive method for the measurement of relatively short-term, steady-state changes in regional blood flow in the mouse kidney.