Urinary Albumin Excretion Levels Research Articles

#2439 The association between vitamin D deficiency and sarcopenia in patients with pre-dialysis CKD

Abstract Background and Aims Sarcopenia is characterized by a progressive and widespread loss of skeletal muscle, leading to muscle weakness and frailty. While primary sarcopenia is linked to aging, secondary sarcopenia can independently occur, especially in chronic conditions like chronic kidney disease (CKD). Investigating sarcopenia in CKD patients has become a focal point in research due to its significant impact on the quality of life, morbidity, and mortality of these individuals. Despite reports connecting vitamin D deficiency, prevalent in CKD, to sarcopenia in the general population, there is limited information regarding the specific relationship between vitamin D and sarcopenia in CKD. This study aims to explore the association between vitamin D deficiency and sarcopenia in patients with pre-dialysis CKD. Method This cross-sectional study enrolled 569 patients with pre-dialysis chronic kidney disease (CKD), defined as having an estimated glomerular filtration rate (eGFR) less than 60 ml/min/m2. Sarcopenia was assessed using computed tomography (CT)-derived skeletal muscle index (SMI), calculated based on the cross-sectional area of skeletal muscles at the 3rd lumbar vertebra. Using Korean-specific SMI cutoffs, patients were categorized into sarcopenic and non-sarcopenic groups. Logistic regression analyses were employed to explore associations between sarcopenia and baseline variables, including active vitamin D, specifically 1,25-dihydroxyvitamin D [1,25(OH)2D] level. Results Among the 569 patients, 278 had CKD stage 3, 198 had CKD stage 4, and 93 had CKD stage 5. The prevalence of sarcopenia increased with advancing CKD stage (CKD stage 3 vs. stage 4 vs. stage 5: 20.1% vs. 31.3% vs. 41.9%, P < 0.001). Univariate logistic regression analysis revealed an association between 1,25(OH)2D levels and sarcopenia. Additionally, age, male sex, diabetes, body mass index, eGFR, urinary albumin excretion levels, and serum levels of albumin, high-sensitive C-reactive protein (hsCRP), and intact parathyroid hormone were associated with sarcopenia. In multivariable logistic regression analysis, 1,25(OH)2D levels remained independently associated with sarcopenia (per 1 pg/dl increase, odds ratio [OR]: 0.84; 95% confidence interval [CI]: 0.80–0.87, P < 0.001). Age (per 1 year increase, OR: 1.04, 95% CI: 1.02–1.07, P < 0.001), diabetes (vs. non-diabetes, OR: 2.33, 95% CI: 1.39–3.89, P = 0.001), serum albumin (per 1 mg/dl increase, OR: 0.39, 95% CI: 0.22–0.71, P = 0.0023), and hsCRP (per 1 mg/dl increase, OR: 1.91, 95% CI: 1.25–2.92, P = 0.003) were also independently associated with sarcopenia. Conclusion This study revealed an independent association between 1,25(OH)2D levels and sarcopenia derived from CT scans in pre-dialysis CKD patients. This association retained significance even after adjusting for traditional risk factors for sarcopenia, such as age, diabetes, serum albumin, and hsCRP. Further prospective studies are necessary to determine whether therapy with exogenous 1,25(OH)2D drugs can improve sarcopenia in these patients.

Sex-Based Differences in Proximal Tubule Endocytic Capacity in Mice

Women are more protected from acute kidney injury, and chronic kidney disease (CKD) in women progresses more slowly to end-stage renal disease compared to men. CKD involves changes in the integrity of the glomerular barrier function with consequent proteinuria. The kidney proximal tubule (PT) normally reclaims albumin and other filtered proteins via receptor-mediated endocytosis facilitated by the multiligand receptors megalin and cubilin. PT health, and ultimately kidney function, is impacted by the uptake of excess albumin and by nephrotoxic drugs that enter cells in a megalin/cubilin dependent manner. Recent transcriptomic studies in mice have shown differences in the expression of megalin and cubilin between males and females and across the S1, S2, and S3 sub-segments that comprise the PT. Additionally, reduced reabsorption of water and salt by the PT has been demonstrated in female rodents. These differences in PT receptor expression and functionality likely contribute to sex-based differences in PT endocytic capacity. To determine if there are sex-based differences in the recovery of filtered proteins by the PT, urine and kidney tissue were collected from five male and five female 13-week-old C57BL/6 mice. Urinary albumin was quantified by ELISA. Kidneys were processed for Western blotting and immunofluorescence staining. We used imaging-based approaches to quantify the expression and distribution of albumin, megalin, and cubilin in SGLT2-positive S1 and SGLT2-negative, OAT1-positive S2 segments. Under normal conditions, S1 had greater intensity of subapical albumin staining in both males and females. However, females had greater intensity of albumin staining in S2 normalized to S1 compared to males. Female mice had lower levels of urinary albumin excretion when normalized to creatinine than males. Female mice had greater expression of total megalin (1.5x) and cubilin (1.4x) and decreased expression of NHE3 (0.7x) in the kidney cortex. Male and female mice had similar levels of megalin expression and surface localization in S1, however female mice had greater expression and surface localization of megalin in S2. Males had greater colocalization of megalin with lysosomal marker, LAMP1. In both males and females, the colocalization of megalin with LAMP1 decreased from S1 to S2. Together, these data suggest that, at baseline in female mice, S2 has greater endocytic capacity and contributes more to the uptake of normally filtered albumin than in males. This increased capacity could explain the reduced urinary excretion of albumin observed in rodent injury models and in women with CKD. Current studies are focused on incorporating these sex-based differences into a multiscale mathematical model of protein uptake along the length of the PT. National Institutes of Health: T32 DK007052, F31 DK121394, R01 DK125049, R01 DK118726, R01 DK083785, S10 OD021627, S10 OD028596, U54 DK137329, ASN Foundation for Kidney Research Pre-Doctoral Fellowship Award. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Baicalein improves renal interstitial fibrosis by inhibiting the ferroptosis in vivo and in vitro

Evidence indicates that Baicalein can ameliorate renal interstitial fibrosis by inducing myofibroblast apoptosis and inhibit the RLS3-induced ferroptosis in melanocytes. However, the relationship between renal interstitial fibrosis and anti-ferroptosis affected by Baicalein remains unclear. In our study, the anti-fibrosis and anti-ferroptosis effects of Baicalein were assessed in a rat model induced by the UUO method in vivo, and the effects of Baicalein on Erastin-induced ferroptosis of renal MPC-5 cells were examined by Western blot of fibrosis-related and ferroptosis-related proteins in vitro. In the UUO-induced rat model, Baicalein decreased kidney weight loss, improved renal function assessed the biomarks of urinary albumin excretion, serum creatine, and BUN levels, and reduced renal tubular injury. Furthermore, Baicalein inhibited renal ferroptosis by reducing ROS and MDA levels and increasing SOD and GSH levels in the UUO rat model. In addition, Baicalein potently reduced the expression of fibrosis-related proteins such as TGF-β1, a-SMA, and Smad-2 to prevent renal interstitial fibrosis, and increased the expression of ferroptosis-related proteins such as SLC7A11, GPX4, and FTH to inhibit ferroptosis both in vitro and in vivo. Taken together, Baicalein exerts anti-fibrosis activity by reducing the ferroptosis response on the UUO-induced rat model and renal MPC5 cells. Therefore, Baicalein, as a novel therapeutic method on kidney diseases with strong activity in suppressing ferroptosis, could be a potential alternative treatment for renal interstitial fibrosis.

Plasma plasminogen activator inhibitor-1 as a predictive marker of type 2 diabetic nephropathy in Egyptian patients

Background The study investigates the relationship between Plasminogen activator inhibitor-1 (PAI-1) levels and the progression of diabetic nephropathy (DN). The fibrinolytic system is impaired in type 2 diabetes mellitus (DM), leading to elevated PAI-1 levels, increasing the potential for renal fibrosis and worsening nephropathy. Patients and methods The research involved 196 type 2 DM patients and 50 healthy control individuals divided into four groups based on Urinary albumin excretion (UAE). Results The diabetic group had significantly higher PAI-1 levels than the control group. The study also found that diabetic patients with macroalbuminuria had higher PAI-1 levels than the control group. Patients with higher UAE and PAI-1 levels had significantly higher serum creatinine concentrations. The study found a significant negative link with creatinine clearance and a significant positive correlation between PAI-1 and serum creatinine and 24-h urine albumin in terms of PAI-1. Conclusions There was a notable positive association with the 24-h UAE, suggesting that increases in PAI-1 were linked to a decline in renal function.

PS-B08-10: COMBINATION OF SYSTEMIC ATRAP DEFICIENCY WITH ANGIOTENSIN II STIMULATION ACCELERATES THE DEVELOPMENT OF DIABETIC NEPHROPATHY IN STREPTOZOTOCIN TREATED MICE

Objective: One of the obstacles toward understanding the pathophysiology of diabetic nephropathy (DN) has been the lack of reliable animal models that faithfully replicate features of human DN. Angiotensin II (Ang II) type 1 receptor (AT1R)-associated protein (ATRAP) promotes internalization of AT1R from the cell surface into the cytoplasm, resulting in the suppression of AT1R signaling pathway. We have recently reported that systemic ATRAP deficiency exaggerates streptozotocin (STZ)-induced DN via an activation of renal renin-angiotensin system (RAS) (Haruhara K, et al. Kidney Int 2022). However, STZ-induced diabetic ATRAP knockout mice still exhibited a modest increase in albuminuria with limited pathological changes just in glomerulus. To establish more robust DN models, we examined the effect of Ang II stimulation on the development of DN in STZ-treated ATRAP knockout mice. Design and method: Male eight weeks old C57BL/6 mice (Ctrl) and systemic ATRAP-knockout mice (KO) were divided into three groups: 1) Ctrl-STZ, 2) Ctrl-STZ-Ang II, and 3) KO-STZ-Ang II. Hyperglycemia was induced by intraperitoneal injection of 55 mg/kg STZ for consecutive 5 days. From 4 weeks after STZ administration, Ang II (1000 ng/kg/min) was continuously administered by osmotic mini-pumps for 6 weeks. During the experimental period, body weights and levels of blood glucose were measured every 2 weeks. At 6 and 10 weeks after STZ administration, 24-hour urine samples were collected in metabolic cages, and urinary albumin excretion (UAE) was evaluated by ELISA. At the end of experimental period, mice were euthanized to evaluate renal pathological changes. Results: During the experimental period, body weight gain and levels of blood glucose were not significantly different between the three groups. Nonetheless, although levels of UAE at 6 weeks after STZ administration were still modest and comparable between the three groups, levels of UAE only in the KO-STZ-Ang II group were remarkably increased compared to those in the Ctrl-STZ and Ctrl-STZ-Ang II groups at 10 weeks after STZ administration (679.1 ± 583.7 vs 69.9 ± 10.0 vs 61.3 ± 8.2 μg/day, respectively). Furthermore, in the KO-STZ-Ang II group, exacerbation of mesangial expansion and interstitial fibrosis was observed. Conclusions: In STZ-induced diabetic mice, combination of systemic ATRAP deficiency with Ang II stimulation accelerated the development of DN, suggesting a potential to become a promising mouse model replicating key features of human DN.

PS-C27-2: RISK OF PSEUDO-MICROALBUMINURIA DURING MENSTRUAL SITUATION IN APPARENTLY HEALTHY WORKERS

Background: Measurement of urinary albumin excretion (UAE) is useful for the prediction of future cardiovascular event and is now used not only in clinical but also in health-check up or industrial settings. Especially, microalbuminuria (MA) is an important category of early vascular marker to alert lifestyle modification and/or pharmacological treatment. In the working female population, however, examination of urine in above settings could encounter menstrual situation. Contamination of menstrual blood into urine could induce pseudo-microalbuminuria. However, it has never been studied in apparently healthy women how much of contamination of menstrual blood could increase the risk of pseudo-microalbuminuria. To address this issue, we examined the relationship between urine occult blood level and UAE or frequency of MA cases during menstrual and non-menstrual situation in apparently healthy workers. Subjects and method: We studied 735 female school workers who participated in the Miyagi Karoshi Prevention study. Urinary albumin excretion adjusted for creatinine (UAE) from morning spot urine sample was examined together with annual health check-up in 2019 and 2020. Menstrual situation was confirmed by a questionnaire at the time of examination. One hundred and forty-four non-diabetic, non-hypertensive women without chronic kidney disease (36.9 ± 9.2yrs) were studied either during menstrual or non-menstrual situation in the 2 consecutive years. We compared clinical data during menstrual and non-menstrual situation and then examined the dose-response relationship between urine occult blood category (negative to three plus) and UAE or frequency of MA cases at menstrual and non-menstrual situation. Results: Menstrual situation was associated with significantly higher UAE (4.6 vs. 3.4 mg/gCr, p < 0.001), systolic BP (113.6 ± 13.1 vs. 111.3 ± 12mmHg, p = 0.015) and uric acid level (4.3 ± 0.8 vs. 4.10.9 ± mg/dl, p = 0.004) and was associated with significantly lower high density lipoprotein cholesterol (70.0 ± 14.4 vs. 72.7 ± 16.3 mg/dl, p < 0.001) and eGFR(85.0 ± 14.6 vs, 87.7 ± 16.9 ml/min/1.73m2, p = 0.003) as compared with non-menstrual situation. In menstrual situation, UAE was dose-dependently increased with an increase in urine occult blood category (p < 0.001). Frequency of MA in each urine blood category was 0, 0, 4.5, 0 and 17% (p = 0.001), respectively. UAE was not correlated with systolic BP or other parameters during menstrual situation. In non-menstrual situation, there was no correlation with UAE and urine occult blood level. Conclusion: Risk of pseudo-microalbuminuria could abruptly rise if urine occult blood level reach three plus in menstruated women.

The rs3931283/PVT1 and rs7158663/MEG3 polymorphisms are associated with diabetic kidney disease and markers of renal function in patients with type 2 diabetes mellitus.

Long non-coding RNAs (lncRNAs) are key regulators of gene expression. Some studies have reported the association of polymorphisms in lncRNA genes with diabetes mellitus (DM) and its chronic complications, including diabetic kidney disease (DKD); however, the results are still inconclusive. Thus, we investigated the association of the rs3200401/MALAT1, rs1894720/MIAT, rs3931283/PVT1, rs11993333/PVT1, rs5749201/TUG1, and rs7158663/MEG3 polymorphisms with DKD in patients with type 2 DM (T2DM). This study comprised 902 patients with T2DM and DKD (cases) and 394 patients with T2DM without DKD (controls). The six polymorphisms of interest were genotyped by real-time PCR using TaqMan probes. Frequency of the rs3931283/PVT1 G/G genotype was 36.2% in cases and 31.9% in controls (P = 0.331). After adjustment for gender, glycated hemoglobin, HDL cholesterol, ethnicity, hypertension, and diabetic retinopathy, the G/G genotype was associated with risk for DKD (OR = 1.625, 95% CI 1.020-2.588; P = 0.041). The rs3931283/PVT1 G/G genotype was also associated with higher urinary albumin excretion levels compared to A allele carriers (P = 0.017). No difference was found in rs7158663/MEG3 genotype frequencies between T2DM controls and DKD patients (OR = 1.087, 95% CI 0.686-1.724; P = 0.722). However, the rs7158663/MEG3 G/G genotype was associated with protection against severe DKD (OR = 0.694, 95% CI 0.488-0.989; P = 0.043, for patients with severe DKD vs. T2DM controls). The rs7158663/MEG3 G/G genotype was also associated with lower creatinine levels (P = 0.007) and higher estimated glomerular filtration rate (P = 0.010) compared to A allele carriers. No association was found between the rs11993333/PVT1, rs3200401/MALAT1, rs1894720/MIAT, and rs5749201/TUG1 polymorphisms and DKD or its laboratory markers. The rs3931283/PVT1 G/G and rs7158663/MEG3 G/G are associated with DKD and markers of renal function in T2DM patients from a Brazilian population.

IMBALANCE IN BONE MORPHOGENIC PROTEINS 2 AND 7 IS ASSOCIATED WITH HYPERTENSION AND ARTERIAL STIFFNESS IN CHRONIC KIDNEY DISEASE

Objective: Patients with chronic kidney disease (CKD) exhibit an elevated cardiovascular risk from the early stages (I-III) with an exponentially increasing risk of cardiovascular death as kidney function worsens. The hypothesis of this study is that there is a imbalance in bone morphogenetic proteins BMP-2 (procalcifying) and BMP-7 (renoprotective) at early CKD stages associated with the development of hypertension, vascular stiffness and progressive kidney damage. Design and method: BMP-2 and BMP-7 levels were determined by ELISA in samples of CKD patients (stages I-III; n = 95) and of Munich Wistar Frömter (MWF) rats (22 weeks old; n = 6), a genetic model with spontaneous nondiabetic CKD, compared to age-matched Wistar rats. Results: Plasma BMP-2 levels were significantly higher in stage III CKD patients compared to controls. BMP-7 was, however, significantly lower at any stage of CKD compared to the control group with a significant further reduction in stage III patients. MWF rats presented significantly higher BMP-2 plasma levels compared to Wistar rats, whereas BMP-7 level was lower. The MWF strain showed an elevation of both systolic (SBP) and diastolic blood pressure (DBP), as well as pulse wave velocity (PWV) compared to the Wistar group. A positive correlation was observed between the BMP-2/BMP-7 ratio and SBP, DBP, and PWV, respectively. Plasma 25-OH cholecalciferol concentration was lower in MWF showing a negative correlation with the BMP-2/BMP-7 plasma ratio. Urinary albumin excretion (UAE), tubular damage markers, Kim-1 and Ngal, as well as renal BMP-2 were significantly higher in the MWF group compared to Wistar, whereas renal BMP-7 expression was significantly lower. A positive correlation was detected between the BMP-2/BMP-7 ratio and both the UAE and plasma creatinine level. Periaortic and mesenteric perivascular adipose tissue (PVAT) from MWF rats showed an altered expression of BMP-2, BMP-7, profibrotic and calcification markers. Conclusions: Our data show an imbalance in plasma, kidney and PVAT-derived BMP-2 and BMP-7 levels associated with hypertension and arterial stiffness. This might reflect a profibrotic/pro-calcifying propensity related with progressive CKD.

DECREASED URINARY LEVELS OF SIRT1 AS NON-INVASIVE BIOMARKER OF EARLY RENAL DAMAGE IN HYPERTENSION

Objective: Hypertension and diabetes mellitus (DM) produce renal injury. Sirtuins have become important players in renal damage, but their value as biomarkers is poorly assessed. The aims of the study were to evaluate the sirtuin1 levels (SIRT1), and miRNAs miR-34a and miR-200, that regulate SIRT1 expression in urinary cell pellet from hypertensive patients with incipient renal damage with and without diabetes. In addition, to mimic disease conditions and deepen into the implications of SIRT1 in podocyte injury, employed cultured podocytes subjected to high glucose (HG) and angiotensin II (Ang II) concentrations. Design and method: We quantified urinary SIRT1 and claudin 1 (CLDN1) mRNA and miR-34a and miR-200a levels by quantitative real-time polymerase chain reaction (RT-qPCR) from urinary cell free pellet from patients and in cultured human immortalized podocytes treated with HG and ANG II. Protein levels and subcellular localization were analyzed by Western blot and fluorescence analyses by confocal microscopy. Results: We found decreased SIRT1 levels in patients with increased urinary albumin excretion (UAE), the lowest with diabetes presence, and a strong association with UAE, discriminating incipient renal damage. In vitro experiments also showed SIRT1 overall decreases in podocyte cultures under treatment conditions. In urine samples, miR-34a was reduced and miR-200a increased, both related to UAE levels. However, both miRNAs were generally increased in podocyte cultures under high glucose and angiotensin-II treatment. Conclusions: Results evidenced reduction of urinary SIRT1 from hypertensives with or without DM. The decrease of SIRT1 and increase of CLDN1 in patients and podocytes subjected to stress support the implication of both in renal injury and evidence a reflection in urinary levels. The alterations found in miR-34a and miR-200a both in patients’ urine and podocyte cultures reveal novel points to further address their implication in renal injury through SIRT1 regulation. Interestingly, SIRT1 mRNA levels and miR-200a are related to increases in UAE and discriminate the presence of UAE in patients. In summary, SIRT1 mRNA measurements is an easily accessible and non-invasive method to early characterize renal damage in patients.

The rs705708 A allele of the ERBB3 gene is associated with lower prevalence of diabetic retinopathy and arterial hypertension and with improved renal function in type 1 diabetic patients

IntroductionThe Erb-b2 receptor tyrosine kinase 3 (ERBB3) is involved in autoimmune processes related to type 1 diabetes mellitus (T1DM) pathogenesis. Accordingly, some studies have suggested that single nucleotide polymorphisms (SNPs) in the ERBB3 gene confer risk for T1DM. Proliferation-associated protein 2G4 (PA2G4) is another candidate gene for this disease because it regulates cell proliferation and adaptive immunity. Moreover, PA2G4 regulates ERBB3. To date, no study has evaluated the association of PA2G4 SNPs and T1DM. Aim: To evaluate the association of ERBB3 rs705708 (G/A) and PA2G4 rs773120 (C/T) SNPs with T1DM and its clinical and laboratory characteristics. MethodsThis case-control study included 976 white subjects from Southern Brazil, categorized into 501 cases with T1DM and 475 non-diabetic controls. The ERBB3 and PA2G4 SNPs were genotyped by allelic discrimination-real-time PCR. ResultsERBB3 rs705708 and PA2G4 rs773120 SNPs were not associated with T1DM considering different inheritance models and also when controlling for covariables. However, T1DM patients carrying the ERBB3 rs705708 A allele developed T1DM at an earlier age vs. G/G patients. Interestingly, in the T1DM group, the rs705708 A allele was associated with lower prevalence of diabetic retinopathy and arterial hypertension as well as with improved renal function (higher estimated glomerular filtration rate and lower urinary albumin excretion levels) compared to G/G patients. ConclusionsAlthough no association was observed between the ERBB3 rs705708 and PA2G4 rs773120 SNPs and T1DM, the rs705708 A allele was associated, for the first time in literature, with lower prevalence of diabetic retinopathy and arterial hypertension. Additionally, this SNP was associated with improved renal function.

Efficacy and safety of eplerenone treatment for patients with diabetic nephropathy: A meta-analysis.

Diabetic nephropathy (DN), which is correlated with an increased risk of cardiovascular disease, significantly elevates the morbidity and mortality of patients with diabetes. Recently, the benefits of mineralocorticoid receptor antagonists in chronic kidney disease (CKD), such as their anti-inflammatory and anti-fibrotic properties, have been discovered. Thus, the present meta-analysis aimed to systematically assess the efficacy and safety of eplerenone treatment in patients with DN. Six electronic databases—PubMed, The Cochrane Library, Embase, Web of Science, CNKI (China National Knowledge Infrastructure), and CBM(Chinese BioMedical Literature Database)—were searched to retrieve randomized controlled trials that assessed eplerenone treatment in patients with DN and were published up to July 31, 2021. Eight randomized controlled trials involving 838 patients were included. Between the eplerenone treatment groups and controls, significant differences were identified in 24-h urine protein levels (mean difference [MD], −19.63 [95% CI, −23.73 to −15.53], P < 0.00001), microalbuminuria (MD, -7.75 [95% CI, -9.75 to -5.75], P < 0.00001), urinary albumin-creatinine ratio (MD, -48.29 [95% CI, -64.45 to -32.14], P < 0.00001), systolic blood pressure (SBP) (MD, -2.49 [95% CI, -4.48 to -0.50], P = 0.01), serum potassium levels (MD, 0.19 [95% CI, 0.13 to 0.24], P < 0.00001), and levels of the renal fibrosis indicator laminin (MD, -8.84 [95% CI, -11.93 to -5.75], P < 0.00001). However, for the effect of estimated glomerular filtration rate (MD, 1.74 [95% CI, -0.87 to 4.35], P = 0.19) and diastolic blood pressure (MD, -0.51 [95% CI, -1.58 to 0.57], P = 0.36), the differences between the two groups were not significant. In addition, no noticeable difference was identified in the adverse events of hyperkalemia and cough between them. These findings suggest that eplerenone exerts beneficial effects on DN by significantly reducing urinary albumin or protein excretion, SBP, and laminin levels, without increasing the incidence of hyperkalemia and other adverse events.

Factor Xa inhibitor, edoxaban ameliorates renal injury after subtotal nephrectomy by reducing epithelial-mesenchymal transition and inflammatory response.

Chronic kidney disease (CKD) is an increasing and life‐threatening disease worldwide. Recent evidence indicates that blood coagulation factors promote renal dysfunction in CKD patients. Activated factor X (FXa) inhibitors are safe and first‐line drugs for the prevention of thrombosis in patients with atrial fibrillation. Here, we investigated the therapeutic effects of edoxaban on CKD using the mouse 5/6 nephrectomy model. Eight‐week‐old wild‐type mice were subjected to 5/6 nephrectomy surgery and randomly assigned to two groups, edoxaban or vehicle admixture diet. Edoxaban treatment led to reduction of urinary albumin excretion and plasma UN levels compared with vehicle group, which was accompanied with reduced glomerular cross‐sectional area and cell number. Edoxaban treatment also attenuated fibrinogen positive area in the remnant kidneys after subtotal nephrectomy. Moreover, edoxaban treatment resulted in attenuated tubulointerstitial fibrosis after 5/6 nephrectomy, which was accompanied by reduced expression levels of epithelial‐mesenchymal transition (EMT) markers, inflammatory mediators, and oxidative stress markers in the remnant kidneys. Treatment of cultured proximal tubular cells, HK‐2 cells, with FXa protein led to increased expression levels of EMT markers, inflammatory mediators, and oxidative stress markers, which were abolished by pretreatment with edoxaban. Treatment of HK‐2 cells with edoxaban attenuated FXa‐stimulated phosphorylation levels of extracellular signal‐regulated kinase (ERK) and NF‐κB. Our findings indicate that edoxaban can improve renal injury after subtotal nephrectomy by reducing EMT and inflammatory response, suggesting that FXa inhibition could be a novel therapeutic target for CKD patients with atrial fibrillation.

Investigation of angiotensin-1 converting enzyme 2 gene (G8790A) polymorphism in patients of type 2 diabetes mellitus with diabetic nephropathy in Pakistani population.

BackgroundType 2 diabetes mellitus is a multifactorial disease that escalates the risk of other associated complications such as diabetic neuropathy, retinopathy, and nephropathy. Diabetic nephropathy is a microvascular condition that leads to end-stage renal disease (ESRD). There are several genes involved in disease development and it is a challenging task to investigate all of these. Nonetheless, identifying individual gene roles can assist in evaluating the combinatorial effects with other genes. Angiotensin-1 converting enzyme 2 (ACE2), is the key regulator of blood pressure in the Renin-Angiotensin-Aldosterone System that hydrolyzes angiotensin II (vasoconstrictor) into angiotensin 1–7 (vasodilator). The association of different variants of the ACE2 with the risk of type 2 diabetes mellitus has been determined in various populations with susceptibility to other complications. This study was aimed to investigate the association of Angiotensin-1 converting enzyme 2 polymorphism, G8790A, with the increased risk of type 2 diabetes mellitus (T2DM) development with the complication of diabetic nephropathy (DN) in the Pakistani population.MethodsIn this case-control study, a total of 100 healthy controls and 100 patients of type 2 diabetes mellitus aged > 40 years, having disease duration ≥ 10 years were compared. The G8790A polymorphism in ACE2 was analyzed by allele-specific polymerase chain reaction (AS-PCR). The urinary albumin excretion (UAE), urinary creatinine, and albumin to creatinine ratios (ACR) were determined to assess renal function status. Pearson bivariate correlation coefficients were calculated to investigate the relationship among all the parameters. Crude and adjusted odds ratios were found to determine any risk association between ACE2 G8790A polymorphisms and disease development. The p-values < 0.05 were considered significant.ResultsA homogeneity was obtained regarding the distribution of data by sex, BMI, diastolic blood pressure, pulse rate and urinary creatinine levels between case and control groups. The ACR showed a significant correlation with UAE (r = 0.524, p = 0.001), urinary creatinine (r = -0.375, p = 0.001) and random blood sugar levels (r = 0.323, p = 0.005) with the complication of diabetic nephropathy in T2DM patient. Females with the AA genotype had a 10-fold increased risk for the development of type 2 Diabetes (OR = 9.5 [95% CI = 2.00–21.63] p<0.002). Males having A allele showed a significant association for susceptibility of type 2 Diabetes (OR = 3.807 [95% CI = 1.657–8.747] p<0.002). However, none of the genotypes or alleles revealed an association for diabetic nephropathy in male and female patients. Urinary ACR was also found to be positively correlated with UAE (r = 0.642 p = 0.001 & 0.524, p = 0.001) and random blood sugar levels (r = 0.302, p = 0.002 & r = 0.323, p = 0.005) in T2DM and T2DM+DN groups, respectively.ConclusionThe study finding indicated that female AG/AA genotype and male A genotype of G8790A polymorphism in the ACE2 gene were associated with type 2 diabetes mellitus as a genetic risk factor but are not associated with diabetic nephropathy in the Pakistani population.

Sex Differences in Glomerular Protein Expression and Effects of Soy-Based Diet on Podocyte Signaling.

Background:Kidney disease is a major public health issue arising from loss of glomerular podocyte function, and there are considerable sex differences in its prognosis. Evidence suggests a renoprotective effect of estrogen and soy diet-derived phytoestrogens, although the molecular basis for this is poorly understood.Objective:Here, we aim to assess sex differences in expression of key proteins associated with podocyte survival and determine the effects of dietary soy on glomerular and podocyte signaling.Methods:Male and female FVB mice were fed control, low (1%), and high (20%) doses of isolated soy protein (ISP) in utero and until 100 days of age. Spot urine was collected to measure proteinuria and isolated glomeruli were used to quantify activated and total levels of nephrin, Akt, and ERK1/2. To investigate protective effects of specific soy phytoestrogens, cultured podocytes were treated with or without daidzein and subject to control or high glucose as a model of podocyte injury.Results:Nephrin and Akt were elevated at baseline in glomeruli from females compared to males. Both sexes that were fed 1% and 20% ISP displayed robust increases in total glomerular Akt compared to controls, and these effects were more prominent in females. A similar trend at both doses in both sexes was observed with activated Akt and total nephrin. Notably, males exclusively showed increased phosphorylation of nephrin and extracellular signal-regulated kinase (ERK) at the 1% ISP dose; however, no overt changes in urinary albumin excretion or podocin levels were observed, suggesting that the soy diets did not impair podocyte function. Finally, in cultured male and female podocytes, daidzein treatment suppressed high glucose-induced ERK activation.Conclusions:Together, our findings reveal a putative mechanism to explain the protective influence of sex on kidney disease progression, and they provide further evidence to support a beneficial role for dietary soy in preserving glomerular function.

Selective decrease in alpha1-antitrypsin levels in diabetic retinopathy: Could the levels of it be playing a role in the pathophysiology of diabetic retinopathy?

Type 2 diabetes mellitus (T2DM) is known to induce inflammation and activation of neutrophils causing the release of neutrophil elastase (NE), a pro-inflammatory proteinase. The activity of NE is regulated by endogenous inhibitors alpha1-antitrypsin (α1-AT) and alpha2-macroglobulin (α2-MG). Disrupted proteolytic homeostasis in T2DM patients is one of the causes for vascular complications. This study was carried out for evaluating the levels of plasma NE, α1-AT, α2-MG and NE-α1-AT complex to understand their roles in the pathophysiology of diabetic nephropathy (DN) and diabetic retinopathy (DR). A total of 240 subjects (Control n=60, T2DM n=60, DN n=60 and DR n=60) were recruited after recording history, clinical examination and laboratory investigations. Retinopathy was confirmed by fundoscopy and nephropathy by urinary albumin excretion and serum creatinine levels. NE was measured using STANA. α1-AT, α2-MG and NE-α1-AT complex were estimated by ELISA. Baseline clinical and laboratory findings were confirmatory to the study groups. The mean elastase activity was higher (P<0.0005) in diabetes groups (T2DM=0.73±0.31, DN=0.87±0.35, DR=0.76±0.41) than controls (0.35±0.20). The levels of α1-AT were lower in DR (8.77±2.85) than DN (26.26±6.16) and T2DM (41.13±14.06) when juxtaposed with controls (122.95±25.71). The approximate fold decrease of α1-AT levels was 15 for DR and 4 for DN compared to controls. The levels of α2-MG were lowered in T2DM (167.29±30.45), DN (144.66±13.72) and DR (104.67±11.47) than controls (208.87±31.16). The NE-α1-AT complex levels were: controls (215.83±13.61), T2DM (98.85±23.85), DN (129.26±20.40) and DR (153.25±17.11). Homeostasis of NE, α1-AT and α2-MG is disrupted in T2DM, DN and DR. Strikingly reduced levels of α1-AT observed in DR are indicative of its possible role in the pathophysiology of retinopathy and emphasizes α1-AT as a plausible therapeutic target.

Associations of Homocysteine with B Vitamins and Zinc in Serum Levels of Patients with Type 2 Diabetes Mellitus: A Cross-Sectional Study.

The association of homocysteine metabolism-related nutrients along with renal function to homocysteine levels is not well known in patients with type 2 diabetes mellitus (T2DM). We investigated the relevance of kidney function, albuminuria, and nutritional factors to serum homocysteine in T2DM patients. This cross-sectional study enrolled 149 T2DM patients (96 men and 53 postmenopausal women), and patient characteristics and laboratory data including kidney-related data [glomerular filtration rate (eGFR), urinary albumin excretion (UACR), uric acid] and metabolism parameters (hemoglobin A1c and lipids) were collected from the medical record and serum levels of vitamin B12, folic acid, zinc, homocysteine and UACR were also acquired. In total subjects, serum levels of homocysteine, vitamin B12, and folic acid were within reference intervals, but zinc levels were close to lower limits of its reference interval. A multivariate-adjusted analysis showed that gender (β=-0.259, p<0.001), uric acid (β=0.267, p<0.001), eGFR (β=-0.188, p=0.001), log UACR (β=0.190, p=0.002), log folic acid (β=-0.259, p<0.001), log vitamin B12 (β=-0.224, p<0.001) and zinc (β=-0.169, p=0.006) were correlated to log homocysteine. In multiple regression analysis by gender, these correlations were found similarly in men, but neither log folic acid nor zinc showed correlations with log homocysteine in women. The present study suggests that renal function parameters and the certain nutritional factors have a possible influence on serum homocysteine, in T2DM patients including diabetes kidney disease.

Glucose control, diabetic retinopathy, and hemodialysis induction in subjects with normo-microalbuminuric type 2 diabetic patients with normal renal function followed for 15 years

AimsA high urinary albumin excretion (UAE) and low glomerular filtration rate (GFR) are risk factors for progressive renal function loss in type 2 diabetic patients. In addition, diabetic retinopathy (DR) is also a risk factor for progressive renal function decline in microalbuminuric type 2 diabetic patients. We aimed to elucidate the factors, including DR, associated with a more severe situation of diabetic nephropathy, i.e., hemodialysis (HD) induction in normo- and microalbuminuric type 2 diabetic patients without renal dysfunction. MethodsNormo- and microalbuminuric type 2 diabetic patients with normal renal function whose GFRs had been measured by iohexol injection in 1995–1997 and had been followed for over 5 years were analyzed (n = 199). HbA1c levels was divided into HbA1c ≥ 7.0 (n = 146) and <7.0 (n = 53) groups. The UAE levels were classified as normoalbuminuria (NA, n = 114) and microalbuminuria (MA, n = 85). Seventy-two patients had DR, and 96 had hypertension. Patients were followed up for 15.7 ± 6.0 years and frequency of and duration to the HD induction were evaluated. ResultsDuring the study period, 8 patients received HD induction. There were no remarkable differences in the rates of HD induction between patients with and without HbA1c ≥7.0, microalbuminuria, DR or hypertension. A Kaplan-Meier analysis revealed that HbA1c ≥7.0 (p = 0.037) and DR (p = 0.037) were associated with a significantly higher risk of HD induction than HbA1c <7.0 and no DR, respectively while albuminuria grade and hypertension were not associated with the risk of HD induction. There was significant negative correlation between HbA1c and annual decline rate of eGFR and annual decline rate of eGFR in the patients with prepro-proliferative DR (PDR) was significantly higher than that in the patients without DR. In the multivariate analysis, HbA1c and PDR showed significant relationships with the annual decline rate of eGFR. ConclusionsIt was reasonable that poorer glycemic control affected HD induction for 16 years follow-up. However, DR, especially PDR, should also be considered a substantial risk factor for HD induction although microalbuminuria and hypertension did not predict it at the early stage of diabetic nephropathy in type 2 diabetic patients with normal renal function.

403-P: Plasma Renin Level and Aldosterone-to-Renin Ratio Are Associated with Diabetic Kidney Disease

Aim: A close association between primary aldosteronism (PA) and atherosclerosis has been well established. We aimed to determine the association between plasma renin activity (PRA) and aldosterone levels (PAC) as well as their ratios (PAC-to-PRA ratio: ARR) with urinary albumin excretion (UAE) in type 2 diabetic patients who did not fulfill PA confirmatory criteria. Methods: The subjects were 70 type2 diabetic patients (age: 53.3±11.0 years, male: 51, female: 19) with UAE ≤100 mg/gCr. They did not take ACE inhibitors, angiotensin receptor blockers (ARB) or diuretic agents. Patients were selected for this study based on various criteria: PRA level ˂10 ng/ml/hr, PAC ˂20 ng/dl, and ARR ˂20 (ng/dl per ng/ml/hr). They were divided into three groups according to UAE levels: NL group (N=33) ˂ 10mg/gCr, N group (N=22) 10-29 mg/gCr, and M group (N=15) 30-100 mg/gCr. Results: There were no significant differences in age, BMI, HbA1c, and eGFR levels among the three groups. The systolic blood pressure in the M group was significantly higher than those in the NL and N groups. The ARR in the M group was 10.1±4.6, which was significantly higher than those in NL (6.5±0.3) or N (7.0±2.7) groups. In the M group, the levels of PRA and PAC were significantly lower than those in NL or N groups. Log UAE was positively correlated with ARR (r=0.34, p&amp;lt;0.0001) and negatively correlated with PRA (r=-0.32, p&amp;lt;0.001). Systolic blood pressure showed a significant positive correlation with ARR and negative correlation with PRA. Conclusion: These results suggest that a high ARR level, which did not fulfill confirmatory criteria for PA, was associated with diabetic nephropathy. The relative increase in PAC to PRA might contribute to the development of diabetic nephropathy, even if PAC remains normal level. Disclosure M. Higa: None. T. Suzuki: None. T. Kido: None. M. Zenri: None. K. Nao: None. M. Hijikata: None. K. Yamashita: None. K. Ikehara: None. T. Ichijo: None.

MO235EFFECT OF OMEGA-3 POLYUNSATURATED FATTY ACID SUPPLEMENTATION ON GLYSEMIC CONTROL AND RENAL FUNCTION IN TYPE 2 DIABETIC PATIENTS WITH CHRONIC KIDNEY DISEASE

Abstract Background and Aims The aim of this study was to evaluate the short-term effects of omega-3 polyunsaturated fatty acids (n-3 PUFA) supplementation on glycemic control and renal function in type 2 diabetic patients with chronic kidney disease. Method Twenty-five diabetic patients received medication containing 2 g/day n-3 PUFA orally in addition to standard treatments. Their estimated glomerular filtration rates (eGFR) were &amp;lt;80 mL/min/1.73 m2. Biochemical values were evaluated before and 3 months after treatment. Results After three months of supplementation, the changes in serum creatinine, uric acid, eGFR and urinary albumin excretion levels did not reach statistical significance. There was no difference between serum glucose, HbA1C and lipid profile values before and after the n-3 PUFA supplementation in patients. Only serum albumin significantly increased from 4.10±0.26 to 4.28±0.31 g/dL (p=0.016), and systolic blood pressure decreased from 121.4±14.5 to 116.6±14.9 mmHg (p=0.001). Conclusion Short-term n-3 PUFA supplementation did not affect renal function and glycemic control in patients with type 2 diabetes with chronic kidney disease.

Triple combination therapy with telmisartan, amlodipine, and hydrochlorothiazide ameliorates albuminuria in a normotensive rat remnant kidney model

BackgroundSome types of antihypertensive drugs may have pleiotropic effects in patients with chronic kidney disease (CKD). However, whether triple combination therapy with angiotensin II receptor blockers (ARBs), calcium channel blockers (CCBs), and thiazide diuretics (TZD) confer renoprotective effects in normotensive CKD remains unknown. Thus, we explored this issue using a normotensive rat remnant kidney model.MethodsSprague-Dawley rats were randomly allocated into four groups: sham (n = 10), 5/6 nephrectomy (NTx) (n = 9), NTx treated with telmisartan and amlodipine (dual) (n = 8), and NTx treated with telmisartan, amlodipine, and hydrochlorothiazide (triple) (n = 7), and followed for 4 weeks. Blood pressure (BP), blood chemistry including renal function, urinary albumin excretion (UAE), and renal pathology were evaluated in all groups.ResultsThere was no significant change in systolic BP among the four groups during the study period. Serum blood urea nitrogen (BUN) was significantly higher, and 24-h creatinine clearance (Ccr) was lower in all NTx groups (p < 0.001). Dual therapy further increased the glomerular diameter in NTx rats (p < 0.001), which was significantly ameliorated by triple therapy (p < 0.001). Triple therapy, but not dual therapy, significantly reduced NTx-induced UAE levels (p < 0.05), whereas BUN, 24-h Ccr, and tubulointerstitial injury scores were comparable among all the NTx groups.ConclusionsOur results suggest that triple combination therapy with telmisartan, amlodipine, and hydrochlorothiazide could ameliorate glomerular hypertrophy and albuminuria in normotensive CKD rats in a BP-lowering independent manner.