A Practical Standardized Composite Nutrition Score Based on Lean Tissue Index: Application in Nutrition Screening and Prediction of Outcome in Hemodialysis Population.

OBJECTIVE: Rapid screening and monitoring of nutritional status is mandatory in hemodialysis population because of the increasingly encountered nutritional problems. Considering the limitations of previous composite nutrition scores applied in this population, we tried to develop a standardized composite nutrition score (SCNS) using low lean tissue index as a marker of protein wasting to facilitate clinical screening and monitoring and to predict outcome. DESIGN AND METHODS: This retrospective cohort used 2 databases of dialysis populations from Taiwan between 2011 and 2014. First database consisting of data from 629 maintenance hemodialysis patients was used to develop the SCNS and the second database containing data from 297 maintenance hemodialysis patients was used to validate this developed score. RESULTS: SCNS containing albumin, creatinine, potassium, and body mass index was developed from the first database using low lean tissue index as a marker of protein wasting. When applying this score in the original database, significantly higher risk of developing protein wasting was found for patients with lower SCNS (odds ratio 1.38 [middle tertile vs highest tertile, P < .0001] and 2.40 [lowest tertile vs middle tertile, P < .0001]). The risk of death was also shown to be higher for patients with lower SCNS (hazard ratio 4.45 [below median level vs above median level, P < .0001]). These results were validated in the second database. CONCLUSION: We developed an SCNS consisting of 4 easily available biochemical parameters. This kind of scoring system can be easily applied in different dialysis facilities for screening and monitoring of protein wasting. The wide application of body composition monitor in dialysis population will also facilitate the development of specific nutrition scoring model for individual facility.

Survival and other clinical outcomes of maintenance hemodialysis patients in Taiwan: a 5-year multicenter follow-up study.

The increasing aging and diabetes mellitus (DM) patients in dialysis population make the quality maintenance of dialysis an imperative issue. Recently, an increasing number of dialysis centers were run by private dialysis providers, many of which apply quality assurance programs and performance management systems to dialysis care. We studied patients in dialysis facilities in Taiwan run by a private chain to see clinical outcomes of centers operating under these systemic strategies. Hemodialysis patients from January 1, 2008 to December 31, 2012 in 25 dialysis facilities in Taiwan, which received the management and consultation from a dialysis service provider, NephroCare (NC), were included. Data pivotal to quality of dialysis were analyzed. During a 5-year interval, 5161 hemodialysis patients were included. For volume control, the proportion of patients with weight gain ≥4.5% decreases from 41.7% to 30.2%. Mean Kt/V is 1.74 ± 0.28. Mean albumin level is 3.92 ± 0.38 g/dL. Patients with phosphate <5.5 mg/dL is up to 71.8%. The mean hemoglobin level is 10.70 ± 1.40 g/dL. More than 80% of patients have adequate iron status. Further, 73% of patients use native arteriovenous fistula. Hospitalization-free survival rate was 56% at the fifth year. Patient survival rate at the fifth year was 66.4%. Overall clinical performances were maintained very stable in NC facilities from this temporal data analysis. The hospitalization and survival rate also compare favorably with those reported internationally. These results warrant further studies to justify the application of this kind of quality assurance programs and performance management systems in dialysis care.

Hemojuvelin modulates iron stress during acute kidney injury: improved by furin inhibitor.

AIMS: Free iron plays an important role in the pathogenesis of acute kidney injury (AKI) via the formation of hydroxyl radicals. Systemic iron homeostasis is controlled by the hemojuvelin-hepcidin-ferroportin axis in the liver, but less is known about this role in AKI. RESULTS: By proteomics, we identified a 42 kDa soluble hemojuvelin (sHJV), processed by furin protease from membrane-bound hemojuvelin (mHJV), in the urine during AKI after cardiac surgery. Biopsies from human and mouse specimens with AKI confirm that HJV is extensively increased in renal tubules. Iron overload enhanced the expression of hemojuvelin-hepcidin signaling pathway. The furin inhibitor (FI) decreases furin-mediated proteolytic cleavage of mHJV into sHJV and augments the mHJV/sHJV ratio after iron overload with hypoxia condition. The FI could reduce renal tubule apoptosis, stabilize hypoxic induced factor-1, prevent the accumulation of iron in the kidney, and further ameliorate ischemic-reperfusion injury. mHJV is associated with decreasing total kidney iron, secreting hepcidin, and promoting the degradation of ferroportin at AKI, whereas sHJV does the opposite. INNOVATION: This study suggests the ratio of mHJV/sHJV affects the iron deposition during acute kidney injury and sHJV could be an early biomarker of AKI. CONCLUSION: Our findings link endogenous HJV inextricably with renal iron homeostasis for the first time, add new significance to early predict AKI, and identify novel therapeutic targets to reduce the severity of AKI using the FI.

Application of Bioimpedance Spectroscopy in Asian Dialysis Patients (ABISAD): serial follow-up and dry weight evaluation.

BACKGROUND: Maintenance of the optimal fluid status in hemodialysis patients is still a challenging task in daily clinical practice. A bioelectric impedance technique has been applied for evaluation of hydration status in the dialysis population in recent years, but serial observations of its role in clinical dry weight determination are lacking. In this study, serial follow-up data of a body composition monitor based on bioimpedance spectroscopy (BCM-BIS) applied in dialysis patients were analyzed to define the technique's feasibility, precision and possible role in dry weight determination. METHODS: BCM-BIS was applied monthly to 194 hemodialysis patients for 6 months. Intra-patient precision was analyzed. Bland-Altman analysis and repeated-measures analysis of variance (ANOVA) were used to define the relationship between the dry weights determined by BCM-BIS and by clinical judgment. RESULTS: The coefficients of variation (CVs) of fluid parameters were <5%. Serial changes in dry weight differences were compared in groups with different post-dialysis hydration status and dry weight differences decreased gradually. Bland-Altman analysis revealed that the range of these differences was significantly narrower towards the latter part of the study. The upper limit of agreement with 95% confidence interval (CI) was 1.47 L and the lower limit was -3.02 L. CONCLUSIONS: BCM-BIS is precise and can be easily applied in the clinical setting. Discrepancy between the dry weights determined by BCM-BIS and by clinical judgment significantly decreased during the study. It is sensitive in dry weight determination, especially for those patients with obvious over-hydration (OH) by BCM-BIS. Patients with post-dialysis OH results beyond some critical values (>1.5 L or <-3 L) should be closely monitored.

Impact of thyroid dysfunction on erythropoietin dosage in hemodialysis patients.

BACKGROUND: Although thyroid diseases exist in patients with renal failure, thyroid function tests are not routine tests in patients on chronic hemodialysis (HD). Therefore, the impact of thyroid diseases on erythropoietin (EPO) dosage in HD patients is not well defined. This study evaluated the relationship between the dose of EPO and the presence or absence of thyroid dysfunction in HD patients. METHODS: This study included 1013 adult patients on HD who did not have a malignancy, liver cirrhosis, thalassemia, iron deficiency, gastrointestinal bleeding, or a major operation within 6 months. Patients were characterized as being euthyroid, or having the sick euthyroid syndrome, primary hypothyroidism, subclinical hypothyroidism, hyperthyroidism, or subclinical hyperthyroidism based on thyroid function tests. Routine biochemistry profiles including an index of the efficiency of HD, along with clinical data over the previous 6-month period, were collected and analyzed. Multiple regression models were employed to assess the relationship between the dose of EPO and the presence or absence of thyroid status. RESULTS: The mean monthly EPO dosages were 77.7±37.0, 70.2±40.6, 90.8±68.4, 78.5±46.7, and 82.3±41.2 µg, respectively, in the sick euthyroid syndrome, euthyroid patients, hypothyroidism, subclinical hypothyroidism, and subclinical hyperthyroidism groups (p<0.05). After adjustment of all other variables in multiple regression, the mean monthly EPO dosage was 19.00±8.59 µg more in hypothyroid patients compared with euthyroid patients (p=0.027). Further, considering an interaction with the presence of diabetes, the mean monthly EPO dosage in patients with either hypothyroidism or subclinical hypothyroidism and diabetes was 54.66±17.12 µg (p=0.001) and 31.51±10.38 µg more than that of euthyroid patients, respectively (p=0.002). CONCLUSIONS: In HD patients, the EPO dosage required to maintain the target hemoglobin level is significantly higher in patients having both hypothyroidism or subclinical hypothyroidism and diabetes than in euthyroid patients.

Improved renal function 12 months after bariatric surgery.

BACKGROUND: Obesity is a risk factor for developing chronic kidney disease (CKD) that may be improved with bariatric surgical weight reduction. The objective of this study was to investigate changes in the glomerular filtration rate (GFR) in severely obese patients 1 year after bariatric surgery. METHODS: GFR was measured in 233 severely obese patients before and more than 12 months after bariatric surgery. Patients were separated by baseline GFR: hyperfiltration (GFR>125 mL/min), normal (GFR 125-90 mL/min), CKD stage 2 (GFR 89-60 mL/min), and CKD stage 3 (59-30 mL/min). The groups were reanalyzed 12 months after bariatric surgery. RESULTS: Of the 233 patients, 61 (26.2%) had hyperfiltration, 127 (54.5%) were normal, 39 (16.7%) had CKD stage 2, and 6 (2.6%) had CKD stage 3. The mean GFR was 146.4±17.1 mL/min in the hyperfiltration group, 105.7±9.6 mL/min in the normal group, 76.8±16.7 mL/min in the CKD stage 2 group, and 49.5±6.6 mL/min in the CKD stage 3 group. The mean GFR 1 year after weight loss surgery decreased to 133.9±25.7 mL/min in the hyperfiltration group, increased to 114.2±22.2 mL/min in the normal group, increased to 93.3±20.4 mL/min in the CKD stage 2 group, and increased to 66.8±19.3 mL/min in the CKD stage 3 group. CONCLUSIONS: Abnormal renal function was common in severely obese patients. Bariatric surgery-induced weight loss had positive effects on renal function at 1 year after surgery.

The impact of dialysis-requiring acute kidney injury on long-term prognosis of patients requiring prolonged mechanical ventilation: nationwide population-based study.

BACKGROUND: Prolonged mechanical ventilation (PMV) is increasingly common worldwide, consuming enormous healthcare resources. Factors that modify PMV outcome are still obscure. METHODS: We selected patients without preceding mechanical ventilation within the one past year and who developed PMV during index admission in Taiwan's National Health Insurance (NHI) system during 1998-2007 for comparison of mortality and resource use. They were divided into three groups: (1) patients with end-stage renal diseases (ESRD) before the index admission for PMV onset; (2) patients with dialysis-requiring acute kidney injury (AKI-dialysis) during the hospitalization course; and (3) patients without AKI or with non dialysis-requiring AKI during the hospitalization course (non-AKI). We used a random-effects logistic regression model to identify factors associated with mortality. RESULTS: Compared with the other two groups, patients with AKI-dialysis had significantly longer mechanical ventilation, more frequent use of vasopressors, longer intensive care unit/hospital stay and higher inpatient expenditures during the index admission. Relative to non-AKI patients, patients with AKI-dialysis had an elevated mortality hazard; the adjusted relative risk ratios were 1.51 (95% confidence interval [CI]:1.46-1.56), 1.27 (95% CI: 1.23-1.32), and 1.10 (95% CI: 1.08-1.12) for mortality rates at discharge, 3 months, and 4 years after PMV, respectively. Patients with AKI-dialysis also consumed significantly higher total in-patient expenditure than the other two patient groups (p<0.001). CONCLUSIONS: Among patients that need PMV care during an admission, the presence of de novo AKI requiring dialysis significantly increased short and long term mortality, and demand for health care resources.

Advanced age affects the outcome-predictive power of RIFLE classification in geriatric patients with acute kidney injury.

The RIFLE (risk, injury, failure, loss, and end-stage) classification is widely used to gauge the severity of acute kidney injury, but its efficacy has not been formally tested in geriatric patients. To correct this we conducted a prospective observational study in a multicenter cohort of 3931 elderly patients (65 years of age or older) who developed acute kidney injury in accordance with the RIFLE creatinine criteria after major surgery. We studied the predictive power of the RIFLE classification for in-hospital mortality and investigated the potential interaction between age and RIFLE classification. In general, the survivors were significantly younger than the nonsurvivors and more likely to have hypertension. In patients 76 years of age and younger, RIFLE-R, -I, or -F classifications were significantly associated with increased hospital mortality in a stepwise manner. There was no significant difference, however, in hospital mortality in those over 76 years of age between patients with RIFLE-R and RIFLE-I, although RIFLE-F patients had significantly higher mortality than both groups. Thus, the less severe categorizations of acute kidney injury per RIFLE classification may not truly reflect the adverse impact on elderly patients.

Effect of diuretic use on 30-day postdialysis mortality in critically ill patients receiving acute dialysis.

BACKGROUND: The impact of diuretic usage and dosage on the mortality of critically ill patients with acute kidney injury is still unclear. METHODS AND RESULTS: In this prospective, multicenter, observational study, 572 patients with postsurgical acute kidney injury receiving hemodialysis were recruited and followed daily. Thirty-day postdialysis mortality was analyzed using Cox's proportional hazards model with time-dependent covariates. The mean age of the 572 patients was 60.8±16.6 years. Patients with lower serum creatinine (p = 0.031) and blood lactate (p = 0.033) at ICU admission, lower predialysis urine output (p = 0.001) and PaO(2)/FiO(2) (p = 0.039), as well as diabetes (p = 0.037) and heart failure (p = 0.049) were more likely to receive diuretics. A total of 280 (49.0%) patients died within 30 days after acute dialysis initiation. The analysis of 30-day postdialysis mortality by fitting propensity score-adjusted Cox's proportional hazards models with time-dependent covariates showed that higher 3-day accumulated diuretic doses after dialysis initiation (HR = 1.449, p = 0.021) could increase the hazard rate of death. Moreover, higher time-varying 3-day accumulative diuretic doses were associated with hypotension (p<0.001) and less intense hemodialysis (p<0.001) during the acute dialysis period. BACKGROUND AND SIGNIFICANCE: Higher time-varying 3-day accumulative diuretic dose predicts mortality in postsurgical critically ill patients requiring acute dialysis. Higher diuretic doses are associated with hypotension and a lower intensity of dialysis. Caution should be employed before loop diuretics are administered to postsurgical patients during the acute dialysis period.

Prevalence of clinical and subclinical thyroid disease in a peritoneal dialysis population.

AIMS: We investigated dialysis duration, dose of erythropoietin (EPO), and clinical manifestations in peritoneal dialysis (PD) patients with subclinical hypothyroidism. METHODS: This cross-sectional study, performed in 3 centers, assessed 122 adult patients on PD for more than 6 months with regard to demographic data, dialysis duration, thyroid function, biochemical data, EPO dose, and clinical manifestations. Thyroid dysfunction was determined by serum thyroid-stimulating hormone, free thyroxine, total thyroxine, total triiodothyronine, antithyroid peroxidase antibodies, and auto-antibodies against thyroglobulin. RESULTS: Of the 122 study patients, 98 (80.3%) were assessed as having euthyroidism; 19 (15.6%), subclinical hypothyroidism; and 5 (4.1%), subclinical hyperthyroidism. The proportion of women (74.2% vs. 57.1%, p = 0.038), the mean duration of PD (58.1 months vs. 37.9 months, p = 0.032), and the weighted mean monthly EPO dose (1.22 µg/kg vs. 1.64 µg/kg, p = 0.009) were significantly higher in the subclinical hypothyroidism group than in the euthyroidism group, but the prevalences of coronary artery disease and cerebrovascular disease were not. From the multivariate model, PD duration was more significant than sex as a risk factor for subclinical hypothyroidism (p = 0.0132). CONCLUSIONS: Subclinical hypothyroidism is frequent in PD patients, especially female patients and patients with a longer PD duration. Compared with euthyroid patients, patients with subclinical hyperthyroidism need a higher dose of EPO to maintain a stable hemoglobin level.

Essential role for Smad3 in angiotensin II-induced tubular epithelial-mesenchymal transition.

Angiotensin II (Ang II) is a key mediator of chronic kidney disease, in which epithelial-mesenchymal transition (EMT) is a critical process mediated by the TGFbeta/Smad signalling pathway. The present study examined the specific role of Smads in Ang II-induced EMT in vitro and in vivo. We found that Ang II signalled through the receptor of AT1, not AT2, to activate Smad2/3 and induce EMT in a normal rat tubular epithelial cell line (NRK52E). Activation of Smads by Ang II was attributed to degradation of an inhibitory Smad7, which was mediated by the AT1-Smurf2-dependent ubiquitin degradation mechanism because blockade of AT1 receptor or knockdown of Smurf2 inhibited Smad7 loss, thereby reducing Smad2/3 activation and EMT in response to Ang II. In contrast, over-expression of Smad7 inhibited Ang II-induced Smad2/3 activation and EMT in NRK52E cells and in a rat model of remnant kidney disease. Moreover, knockdown of Smad3, not Smad2, attenuated Ang II-induced EMT. In conclusion, Ang II activates Smad signalling to induce EMT, which is mediated by a loss of Smad7 through the AT1-Smurf2-dependent ubiquitin degradation pathway. Smad3, but not Smad2, may be a mediator of EMT, while Smad7 may play a protective role in EMT in response to Ang II.

Peroxisome proliferator-activated receptor alpha plays a crucial role in L-carnitine anti-apoptosis effect in renal tubular cells.

BACKGROUND: L-carnitine is synthesized mainly in the liver and kidneys from lysine and methionine from dietary sources. Many reports have shown that L-carnitine can protect certain cells against the toxicity of several anticancer and toxic agents, although the detailed mechanism is poorly understood. In this study, we investigated the protective effect of L-carnitine and its molecular mechanism in renal tubular cells undergoing gentamicin-induced apoptosis. METHODS: Rat tubular cell line (NRK-52E) and mice were used as the model system. Gentamicin-induced apoptosis in renal tubular cells was examined using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labelling. We introduced short interfering RNA transfection and gene-deficient mice to investigate the protective mechanism of L-carnitine. RESULTS: We found that L-carnitine inhibited gentamicin-induced reactive oxygen species generation and correlative apoptotic pathways, resulting in the protection of NRK-52E cells from gentamicin-induced apoptosis. The treatment of L-carnitine also lessened gentamicin-induced renal tubular cell apoptosis in mice. L-carnitine was found to increase the prostacyclin (PGI(2)) generation in NRK-52E cells. The siRNA transfection for PGI(2) synthase significantly reduced L-carnitine-induced PGI(2) and L-carnitine's protective effect. We found that the activity of the potential PGI(2) nuclear receptor, peroxisome proliferator-activated receptor alpha (PPARalpha), was elevated by L-carnitine treatment. The siRNA-mediated blockage of PPARalpha considerably reduced the anti-apoptotic effect of L-carnitine. In PPARalpha-deficient mice, L-carnitine treatment also lost the inhibitory effect on gentamicin-induced apoptosis in kidneys. CONCLUSIONS: Based on these findings, we suggest that L-carnitine protects renal tubular cells from gentamicin-induced apoptosis through PGI(2)-mediated PPARalpha activation.

Urotensin II induces transactivation of the epidermal growth factor receptor via transient oxidation of SHP-2 in the rat renal tubular cell line NRK-52E.

Urotensin-II (UII) is a potent vasoactive peptide that has been implicated in cardiac fibrosis and renal diseases. However, the role played by UII in renal tissues is largely unknown. In this study, we investigated the effects of human UII (hUII) on rat renal proximal tubular cells of the NRK-52E line and the role of Src homology 2-containing phosphotyrosine phosphatase (SHP-2) in the hUII-induced transactivation of the epidermal growth factor receptor (EGFR). Exposure to hUII at low concentrations significantly induced proliferation in NRK-52E cells; this effect was inhibited by treatment with an ERK1/2 inhibitor (PD98059). UII treatment increased the phosphorylation of EGFR and induced the generation of reactive oxygen species (ROS). Treatment of the ROS scavenger N-acetyl-cysteine (NAC) inhibited EGFR transactivation and ERK phosphorylation induced by hUII. SHP-2 was found to interact with EGFR and be transiently oxidized following the hUII treatment. In SHP-2 knockdown cells, UII-induced phosphorylation of EGFR was less influenced by NAC, and significantly suppressed by heparin binding (HB)-EGF neutralizing antibody. Our data suggest that the ROS-mediated oxidation of SHP-2 is essential for the hUII-induced mitogenic pathway in NRK-52E cells.

Peroxisomal proliferator-activated receptor-alpha protects renal tubular cells from doxorubicin-induced apoptosis.

Peroxisome proliferator-activated receptor-alpha (PPAR-alpha) is a transcription factor and has been reported to inhibit cisplatin-mediated proximal tubule cell death. In addition, doxorubicin (Adriamycin)-induced nephrosis in rats is a commonly used experimental model for pharmacological studies of human chronic renal diseases. In this study, we investigated the protective effect of PPAR-alpha on doxorubicin-induced apoptosis and its detailed mechanism in NRK-52E cells and animal models. The mRNA level of PPAR-alpha was found to be reduced by doxorubicin treatment in NRK-52E cells. PPAR-alpha overexpression in NRK-52E cells significantly inhibited doxorubicin-induced apoptosis and the quantity of cleaved caspase-3. Endogenous prostacyclin (PGI(2)) augmentation, which has been reported to protect NRK-52E cells from doxorubicin-induced apoptosis, induced the translocation and activation of PPAR-alpha. The transformation of PPAR-alpha short interfering RNA was applied to silence the PPAR-alpha gene, which abolished the protective effect of PGI(2) augmentation in doxorubicin-treated cells. To confirm the protective role of PPAR-alpha in vivo, PPAR-alpha activator docosahexaenoic acid (DHA) was administered to doxorubicin-treated mice, and it has been shown to significantly reduce the doxorubicin-induced apoptotic cells in renal cortex. However, this protective effect of DHA did not exist in PPAR-alpha-deficient mice. In NRK-52E cells, the overexpression of PPAR-alpha elevated the activity of catalase and superoxide dismutase and inhibited doxorubicin-induced reactive oxygen species (ROS). PPAR-alpha overexpression also inhibited the doxorubicin-induced activity of nuclear factor-kappaB (NF-kappaB), which was associated with the interaction between PPAR-alpha and NF-kappaB p65 subunit as revealed in immunoprecipitation assays. Therefore, PPAR-alpha is capable of inhibiting doxorubicin-induced ROS and NF-kappaB activity and protecting NRK-52E cells from doxorubicin-induced apoptosis.

Tetramethylpyrazine protects rat renal tubular cell apoptosis induced by gentamicin.

BACKGROUND: Gentamicin, a widely used antibiotic for the treatment of bacterial infection, can cause nephrotoxicity. Tetramethylpyrazine (TMP) is a compound purified from the rhizome of Ligusticum wallichi (Chuanxiong) and has been found to protect against ischaemia-reperfusion injury, nephritis and alcohol-induced toxicity in rat kidneys. METHODS: We used rat renal tubular cells (RTCs), NRK-52E, in this study. The cytotoxicity of gentamicin was checked with transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL) staining, and the generation of reactive oxygen species was measured using the fluorescent probe 2,7-dichlorofluorescein. We evaluated several apoptotic parameters: cleaved caspase levels, tumour necrosis factor (TNF-alpha) excretion and nuclear factor Kappa B (NF-kappaB) activity. We also examined the TMP protective effect on gentamicin-induced apoptosis in rat kidneys. RESULTS: The results of this study showed that gentamicin was found to markedly induce apoptosis in NRK-52E cells in a dose-dependent manner; that TMP expressed a dose-dependent protective effect against gentamicin-induced apoptosis; that pre-treatment of the cells with 50 or 100 microM of TMP effectively decreased the reactive oxygen species formation induced by gentamicin; that TMP was found to inactivate the gentamicin-stimulated activities of caspase-3, caspase-8 and caspase-9, to inhibit gentamicin-induced release of cytochrome c, as well as to raise the expression of Bcl-x(L); that TMP inhibited the gentamicin-induced TNF-alpha excretion, and inactivated the transcription factor NF-kappaB; and that the TMP treatment significantly reduced apoptotic injury in rat RTCs. CONCLUSIONS: Based on the results of this study, we suggest that TMP can attenuate gentamicin-induced oxidative stress and apoptotic injury in rat RTCs, and that its character may have therapeutic potential for patients with renal diseases.

Tetramethylpyrazine attenuates adriamycin-induced apoptotic injury in rat renal tubular cells NRK-52E.

Tetramethylpyrazine (TMP), a compound purified from Rhizoma Ligustici, is a widely used active ingredient in Chinese herbal medicine to treat cardiovascular diseases on account of its vasodilatory actions and antiplatelet activity. Studies have shown that TMP can remove oxygen free radicals and protect rat kidney from ischemia-reperfusion injury. In addition, adriamycin-induced nephrosis in rats is commonly used in pharmacological studies of human chronic renal diseases. Apoptosis of renal tubular cells has been reported in adriamycin-treated rats. To examine the therapeutic potential of TMP on chronic progressive renal diseases, adriamycin-induced injury in rat renal tubular cells NRK-52E has been used to monitor its protective effect. In TUNEL staining, TMP showed a dose-dependent protective effect against adriamycin-induced apoptosis in NRK-52E cells. Pretreatment of the cells with 10 or 100 microM of TMP effectively decreased the reactive oxygen species (ROS) formation induced by adriamycin, as measured in fluorescent assays. TMP was found to reduce the adriamycin-stimulated activities of caspase-3, caspase-8 and caspase-9, inhibit adriamycin-induced release of cytochrome C, and elevate the expression of Bcl-x (L). TMP was also able to inhibit the death receptor signaling pathway and suppress the activation of transcription factor NF-kappaB in adriamycin-treated NRK-52E cells. Based on the results of this study, we suggest that TMP can attenuate adriamycin-induced oxidative stress and apoptotic injury in NRK-52E cells, and that it may have therapeutic potential for patients with renal diseases. TMP: tetramethylpyrazine LDH: lactate dehydrogenase ROS: reactive oxygen species DCF: 2',7'-dichlorofluorescein TNF-alpha: tumor necrosis factor-alpha TUNEL: terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling.

Celecoxib induces heme-oxygenase expression in glomerular mesangial cells.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are frequently used as analgesics. They inhibit cyclooxygenases (COX), preventing the formation of prostaglandins, including prostacyclin and thromboxane. A serious side effect of COX-1 and COX-2 inhibitors is renal damage. To investigate the molecular basis of the renal injury, we evaluated the expression of the stress marker, heme oxygenase-1 (HO-1), in celecoxib-stimulated mesangial cells. We report here that a COX-2 selective NSAID, celecoxib, induced a concentration- and time-dependent increase of HO-1 expression in glomerular mesangial cells. Celecoxib-induced HO-1 protein expression was inhibited by actinomycin D and cycloheximide, suggesting that de novo transcription and translation are required in this process. N-acetylcysteine, a free radical scavenger, strongly decreased HO-1 expression, suggesting the involvement of reactive oxygen species (ROS). Celecoxib-induced HO-1 expression was attenuated by pretreatment of the cells with SP 600125 (a specific JNK inhibitor), but not SB 203580 (a specific p38 MAPK inhibitor), or PD 98059 (a specific MEK inhibitor). Consistently, celecoxib activated c-Jun N-terminal kinase (JNK) as demonstrated by kinase assays and by increasing phosphorylation of this kinase. N-acetylcysteine reduced the stimulatory effect of celecoxib on stress kinase activities, suggesting an involvement of JNK in HO-1 expression. On the other hand, LY 294002, a phosphatidylinositol 3-kinase (PI-3K)-specific inhibitor, prevented the enhancement of HO-1 expression. This effect was correlated with inhibition of the phosphorylation of the PDK-1 downstream substrate Akt/protein kinase B (PKB). In conclusion, our data suggest that celecoxib-induced HO-1 expression in glomerular mesangial cells may be mediated by ROS via the JNK-PI-3K cascade.

Blockade of NFkappaB activation and renal inflammation by ultrasound-mediated gene transfer of Smad7 in rat remnant kidney.

BACKGROUND: Transforming growth factor-beta (TGF-beta) in renal fibrosis has been well studied, but little attention has been paid to the potential role of TGF-beta in the resolution of renal inflammation. We hypothesize that TGF-beta exerts its anti-inflammation properties by stimulating its negative signaling pathway involving Smad7. METHODS: A rat remnant kidney model was treated with a doxycycline-regulated Smad7 gene or control empty vector using an ultrasound-microbubble (Optison)-mediated system. Smad7 transgene expression within the kidney was tightly controlled by the addition of doxycycline in the daily drinking water. All animals were euthanized at week 4 for examination of inflammatory responses. RESULTS: Real-time polymerase chain reaction (PCR) and immunohistochemistry revealed that gene transfer of Smad7 resulted in a substantial inhibition of interleukin-1beta (IL-1beta) and tumor necrosis factor alpha (TNFalpha) expression (all P < 0.01 vs. control). This was associated with the attenuation of histology damage, proteinuria, serum creatinine, and an increase in creatinine clearance (all P < 0.05). In addition, overexpression of Smad7 significantly inhibited renal inflammation, including ICAM-1, iNOS, and accumulation of macrophages and T cells in both glomeruli and tubulointerstitium. Furthermore, gene transfer of Smad7 also substantially blocked nuclear factor kappa B (NFkappaB) activation in the rat remnant kidney (P < 0.01). CONCLUSION: TGF-beta/Smad7 signaling plays a critical role in the resolution of renal inflammation in rat remnant kidney model. Inhibition of NFkappaB activation is a key mechanism by which Smad7 suppresses renal inflammation, which suggests a crosstalk pathway between NFkappaB and Smad7. The ability of Smad7 to inhibit renal inflammation indicates that ultrasound-microbubble-mediated Smad7 gene therapy may represents a new therapeutic strategy for glomerulonephritis.

Ultrasound-microbubble-mediated gene transfer of inducible Smad7 blocks transforming growth factor-beta signaling and fibrosis in rat remnant kidney.

Transforming growth factor (TGF)-beta1 has been shown to play a critical role in hypertensive nephropathy. We hypothesized that blocking TGF-beta1 signaling could attenuate renal fibrosis in a rat model of remnant kidney disease. Groups of six rats were subjected to 5/6 nephrectomy and received renal arterial injection of a doxycycline-regulated Smad7 gene or control empty vector using an ultrasound-microbubble-mediated system. Smad7 transgene expression within the kidney was tightly controlled by the addition of doxycycline in the daily drinking water. All animals were euthanized at week 4 for renal functional and histological examination. Hypertension of equivalent magnitude (190 to 200 mmHg) developed in both Smad7- and empty vector-treated rats. However, treatment with Smad7 substantially inhibited Smad2/3 activation and prevented progressive renal injury by inhibiting the rise of 24-hour proteinuria (P < 0.001) and serum creatinine (P < 0.001), preserving creatinine clearance (P < 0.05), and attenuating renal fibrosis and vascular sclerosis such as collagen I and III expression (P < 0.01) and myofibroblast accumulation (P < 0.001). In conclusion, TGF-beta/Smad signaling plays a critical role in renal fibrosis in a rat remnant kidney model. The ability of Smad7 to block Smad2/3 activation and attenuate renal and vascular sclerosis demonstrates that ultrasound-mediated Smad7 gene therapy may be a useful therapeutic strategy for the prevention of renal fibrosis in association with hypertension.

Advanced glycosylation end products induce inducible nitric oxide synthase (iNOS) expression via a p38 MAPK-dependent pathway.

BACKGROUND: Advanced glycosylation end products (AGEs) accumulation in tissue has been implicated in diabetic related complications, including diabetic nephropathy. Activation of peroxisome proliferator activated receptor-gamma (PPAR-gamma) ameliorates diabetic nephropathy. METHODS: In the present study, we investigated the effects of AGEs on inducible nitric oxide synthase (iNOS) expression and nitric oxide production, and the effects of rosiglitazone, an activator of PPAR-gamma, on AGE-induced iNOS expression and nitrite release in glomerular mesangial cells. RESULTS: AGEs caused a dose- and time-dependent increase of iNOS induction and nitrite accumulation in mesangial cells. A protein tyrosine kinase inhibitor (genistein), or a p38 mitogen-activated protein kinase (MAPK) inhibitor (SB203580) suppressed AGE-induced iNOS expression and nitrite release from mesangial cells. Addition of bovine serum albumin (BSA)-AGEs to mesangial cells increased p38 MAPK activities. Activation of PPAR-gamma by rosiglitazone inhibited AGE-induced iNOS expression, nitrite release, and p38 MAPK activation in mesangial cells. AGE-stimulated nitrite release was attenuated by pretreatment with anti-tumor necrosis factor-alpha (TNF-alpha) and anti-transforming growth factor-beta (TGF-beta) antibodies. AGE-induced iNOS expression was inhibited by treatment with a nuclear factor-kappaB (NF-kappaB) inhibitor, pyrrolidone dithiocarbamate. Addition of BSA-AGEs to mesangial cells stimulated p65 NF-kappaB translocation from the cytosol to the nucleus. CONCLUSION: These data suggest that cytokine release, NF-kappaB and p38 MAPK-dependent pathways may play a role in AGE-induced iNOS expression and subsequent nitric oxide production in mesangial cells. Rosiglitazone may prevent AGE-induced iNOS expression by interfering with p38 MAPK activity.