Urinary angiotensin-converting enzyme 2 increases in diabetic nephropathy by angiotensin II type 1 receptor blocker olmesartan

Abstract

Introduction:

Angiotensin-converting enzyme 2 (ACE2) is a member of the renin–angiotensin system that degrades angiotensin (Ang) II to the seven-amino acid peptide fragment Ang-(1-7). We evaluated the changes in urinary ACE2 levels in response to treatment with the angiotensin II type 1 receptor blocker olmesartan in diabetes patients with nephropathy.

Materials and methods:

This prospective, open-label, interventional study was conducted with 31 type 2 diabetes patients with nephropathy. After initial evaluation, patients received 20 mg/day olmesartan, which was increased to 40 mg/day over a 24-week period.

Results:

In diabetes patients with chronic kidney disease, olmesartan significantly increased urinary ACE2 levels independently of blood pressure and plasma aldosterone levels and reduced albuminuria, urinary liver-type fatty acid binding protein (L-FABP), and plasma aldosterone levels. Multivariable regression analysis revealed that the change in urinary L-FABP levels was an independent predictor of increased urinary ACE2 levels.

Conclusion:

Olmesartan may have the unique effect of increasing urinary ACE2 levels. However, whether this contributes to olmesartan’s renoprotective effect must be examined further.

Keywords

Angiotensin-converting enzyme 2 angiotensin II type 1 receptor blocker chronic kidney disease diabetic nephropathy olmesartan

Introduction

Angiotensin-converting enzyme 2 (ACE2) is a member of the renin–angiotensin system (RAS) that degrades angiotensin (Ang) II to the seven-amino acid peptide fragment Ang-(1-7).^1,2 ACE2 is thought to function as a negative regulator of the RAS by degrading Ang II and producing Ang-(1-7) to counterbalance the Ang II-forming activity of ACE. Although ACE2 is reported to localize in renal glomeruli and tubules,^3–6 the differences in ACE2 expression within the kidney seem to be not only tissue-specific but also cell-specific.^6–9 Glomerular expression of ACE2 is reduced in diabetic mice,⁶ and reduced ACE2 gene and protein expression has been confirmed in renal biopsies of patients with type 2 diabetes.⁷ On the other hand, at the tubular level, increased ACE2 activity and protein expression have been found in diabetic mice.^8,9 The renal RAS, specifically Ang II, plays a pivotal role in the pathogenesis of diabetic nephropathy.¹⁰ Ang II and ACE are activated in type 2 diabetes¹¹ and are involved in the development of microvascular and macrovascular complications of diabetes such as nephropathy, retinopathy, and cardiovascular disease.¹² ACE 2 plays an integral role in protecting against renal damage and cardiovascular disease.^8,13 It was reported that deletion of ACE2 leads to the development of Ang II-dependent renal damage, suggesting ACE2 as a renoprotective target in diabetes.¹⁴ ACE2 is a homologue of ACE but is not blocked by ACE inhibitors. However, there are no clinical data showing whether Ang II type 1 receptor blockers (ARB) increase ACE2 activity. In this study, we evaluated the changes in urinary ACE2 levels resulting from ARB treatment in diabetes patients with chronic kidney disease (CKD).

Research design and methods

This prospective, open-label, interventional study was conducted with 31 outpatients after obtaining written informed consent and with the approval of the local ethics committee. The study was performed in accordance with the Declaration of Helsinki. Enrollment criteria were as follows: 1) type 2 diabetes with nephropathy under stable glycemic control, defined as a hemoglobin A1c (HbA1c) level <8.0% after administration of oral antidiabetic agents, insulin injection therapy, or diet therapy alone for 8 consecutive weeks; 2) albuminuria: urinary albumin/creatinine (Cr) ratio >30 mg/g (average of two consecutive measurements recorded during the 8-week pre-treatment period); 3) estimated glomerular filtration rate (eGFR) of 30–89 mL/min/1.73 m²; and 4) hypertension: systolic and diastolic blood pressure (BP) ≥130/80 mmHg treated with a fixed dose of antihypertensive agents other than RAS inhibitors for at least 8 weeks before the study. Exclusion criteria were as follows: 1) age <20 years or >80 years; 2) more than second-degree hypertension (BP >160/100 mmHg)¹⁵; 3) eGFR ≥90 mL/min/1.73 m² or <30 mL/min/1.73 m²; 4) severe heart failure, angina, myocardial infarction, or stroke occurring within 6 months before the trial; and 5) previous treatment with ARB for at least 8 weeks before the study.

After initial evaluation, patients received 20 mg/day olmesartan, which was increased to 40 mg/day over a 24-week period. Doses of other antihypertensive agents and statins were maintained during the study. BP measurements were performed at the outpatient clinic according to Japanese Society of Hypertension (JSH) 2009 guidelines at fixed times after the administration of medications.¹⁵ Target BP was <130/80 mmHg. Measurements were made using a sphygmomanometer (Nippon Colin Co. Ltd, Tokyo, Japan) and performed twice with the patient seated after a 5-min rest.

At each patient visit, safety variables and patient compliance with treatment regimens were assessed. Plasma renin activity and aldosterone concentrations were measured by radioimmunoassay at a contract laboratory (SRL, Inc., Tokyo, Japan) at baseline and at week 24 with the patient in the supine position after a 20-min rest. Urinary albumin excretion was assessed by measuring urinary concentrations of albumin and Cr (albumin/Cr ratio) in the first morning urine sample. Urinary albumin levels were measured by the immunoturbidimetric assay. Urinary liver-type fatty acid binding protein (L-FABP) was measured in the same urine sample by a specific enzyme linked immunosorbent assay (ELISA; SRL, Inc.), and the results were expressed relative to the urinary Cr concentration. Urinary ACE2 concentration was measured at baseline and at week 24 by ELISA (Human ACE2 ELISA Kit, AdipoGen, Inc., Seoul, Korea) as described previously.^16,17 The eGFR was calculated according to the following formula for Japanese patients¹⁸: eGFR (mL/min/1.73 m²) = 194 × sCr^−1.094 × age^−0.287(× 0.739 for women).

Data are expressed as means ± SEM. Changes in biochemical and urinalysis parameters were analyzed using the paired t-test. The relationships between variables were analyzed using Pearson’s correlation. Determinants of the factors associated with the changes in urinary ACE2 levels were assessed using multivariable linear regression. Age, sex, body mass index, changes in systolic and diastolic BP, urinary albumin/Cr ratio, and levels of eGFR, plasma glucose, HbA1c, plasma aldosterone, and urinary L-FABP were selected for the multivariate model. SAS 8.13 (SAS Institute, Cary, NC) was used for all analysis, and significance was set at p < 0.05.

Results

All enrolled patients (n = 31) remained in the study until the end of the trial. Baseline characteristics and concomitant medication use at baseline are shown in Table 1. Fifteen patients (48%) were using other antihypertensive agents at baseline. The final olmesartan dose was 26 ± 2 mg/day. As shown in Table 2, olmesartan treatment significantly decreased both systolic and diastolic BP values. No significant differences were seen in serum Cr levels or eGFR values during the study period. Furthermore, urinary L-FABP level was significantly decreased after olmesartan treatment. Figure 1 shows that the urinary albumin/Cr ratio was significantly decreased by olmesartan treatment from 720 ± 201 mg/gCr at baseline to 317 ± 91 mg/gCr at the end of the study (p < 0.01), whereas urinary ACE2 concentration was significantly increased from 27.8 ± 4.6 ng/mgCr at baseline to 42.9 ± 7.0 ng/mgCr at the end of the study (p < 0.05). Figure 2 shows that plasma renin activity was not significantly changed after 24 weeks of treatment, but plasma aldosterone levels were decreased from 97.6 ± 10.4 at baseline to 77.5 ± 6.7 pg/mL at the end of the study (p < 0.05).

Table 1.

Patient baseline characteristics and medication use.

Number of patients	31
Male/female	20/11
Age (years old)	70.0 ± 1.5
Body mass index (kg/m²)	24.2 ± 0.3
Antihypertensive agents
Calcium channel blocker	13
Diuretics	3
β-blocker	5
α-blocker	2
Antidiabetic agents
Insulin	6
Sulfonylureas	3
DPP-4 inhibitors	12
α-glucosidase inhibitors	5
Metformin	4
Pioglitazone	1
Statins	18
Antiplatelet agent	10
Antihyperuricemic agent	7

DPP-4: dipeptidyl peptidase-IV.

Table 2.

Changes in measured parameters.

	Baseline	24 weeks	p Value
Systolic BP (mmHg)	142 ± 1	135 ± 1	< 0.0001
Diastolic BP (mmHg)	78 ± 2	74 ± 2	< 0.0001
Heart rate (bpm)	76 ± 2	76 ± 2	0.923
Serum creatinine (mg/dL)	1.18 ± 0.1	1.23 ± 0.1	0.111
eGFR (mL/min/1.73 m²)	49.4 ± 3.6	48.0 ± 3.6	0.250
Hemoglobin A1c (%)	6.4 ± 0.1	6.3 ± 0.1	0.288
Plasma glucose (mg/dL)	143 ± 4	141 ±3	0.457
Uric acid (mg/dL)	5.9 ± 0.3	6.2 ± 0.3	0.065
Sodium (mEq/L)	141 ± 0.3	142 ± 0.5	0.246
Potassium (mEq/L)	4.4 ± 0.1	4.5 ± 0.1	0.122
Urinary L-FABP (μg/gCr)	31.7 ± 7.3	15.0 ± 4.0	0.0007

BP: blood pressure; eGFR: estimated glomerular filtration rate; L-FABP: liver-type fatty acid binding protein.

Figure 1.

Changes in the urinary albumin excretion ratio and urinary angiotensin-converting enzyme 2 levels. *p < 0.05, **p < 0.01 vs. baseline.

Figure 2.

Changes in plasma renin activity and plasma aldosterone levels. *p < 0.05 vs. baseline.

In a simple regression analysis, changes in urinary ACE2 levels correlated significantly with changes in urinary L-FABP levels (r = 0.427, p = 0.016), but not with systolic and diastolic BP, urinary albumin/Cr ratio, or eGFR levels (Table 3). Multivariable regression analysis showed that urinary L-FABP levels were an independent predictor of changes in urinary ACE2 levels (R² = 0.426) (Table 4).

Table 3.

Simple regression analysis of changes in urinary angiotensin-converting enzyme 2 levels.

Parameter	r	p Value
Age	−0.160	0.391
Body mass index	0.122	0.517
Systolic blood pressure	0.184	0.321
Diastolic blood pressure	0.140	0.451
eGFR	0.168	0.370
Plasma glucose	−0.045	0.811
Hemoglobin A1c	0.135	0.473
Plasma aldosterone	−0.207	0.262
UACR	0.029	0.879
Urinary L-FABP	0.427	0.016

eGFR: estimated glomerular filtration rate; L-FABP: liver-type fatty acid binding protein; UACR: urinary albumin to creatinine ratio.

Table 4.

Multivariable regression analysis of changes in urinary angiotensin-converting enzyme 2 levels.

Parameter	Standardized coefficient (β)	SE	95% CI	p Value
Age (per additional year)	0.132	0.609	−0.873 to 1.676	0.518
Male sex	−0.241	11.08	−35.67 to 10.71	0.274
Body mass index	0.349	3.113	−0.781 to 12.25	0.081
Systolic blood pressure	−0.117	2.204	−5.791 to 3.435	0.599
Diastolic blood pressure	−0.056	1.094	−2.560 to 2.022	0.808
eGFR	−0.151	0.463	−1.233 to 0.706	0.575
Plasma glucose	−0.149	0.896	−2.464 to 1.286	0.519
Hemoglobin A1c	0.337	9.232	−3.337 to 35.30	0.099
Plasma aldosterone	−0.039	0.155	−0.349 to 0.306	0.876
UACR	0.146	0.077	−0.109 to 0.216	0.499
Urinary L-FABP	−0.551	0.235	−1.012 to −0.027	0.039

R² = 0.426. CI: confidence interval; eGFR: estimated glomerular filtration rate; L-FABP: liver-type fatty acid binding protein; UACR: urinary albumin to creatinine ratio.

During the observation period, none of the patients exhibited a significant increase in the occurrence of adverse effects such as hypotension, and none required renal replacement therapy for hyperkalemia or progressive loss of kidney function.

Discussion

Ang-(1-7) is formed from Ang I and Ang II mainly by ACE2 and acts on the receptor protein Mas.¹⁹ Based on these interactions, a new bioactive pathway in the RAS, referred to as the ACE2–Ang–(1-7)–Mas axis, has been proposed.^20,21 This axis is antagonistic to the classical RAS pathway with ACE–Ang II–Ang II type 1 receptors.^22,23 Deletion or inhibition of ACE2 has been associated with albuminuria in diabetic mice^24,25 and with the development of Ang II-dependent renal damage.¹⁴ Conversely, administration of human recombinant ACE2 was shown to reduce BP and attenuate glomerular injury in rodent models.^26,27 Furthermore, ACE2 delivery by a recombinant protein or virus ameliorated the progression of diabetes-related complications, such as nephropathy and retinopathy.^28–30 Thus, ACE2 may be an endogenous protector against the progression of CKD.

Chronic administration of either telmisartan or captopril did not change urinary ACE2 levels in mice.³¹ Previous studies in humans have shown no significant effects of ARBs or ACE inhibitors on urinary ACE2 levels,^17,32 although in these studies the effects were analyzed for ARBs in general, not for specific ARBs. Furthermore, urinary ACE2 levels in patients treated with olmesartan for >1 year were higher than in healthy controls, although other antihypertensive drugs, including calcium channel blockers and other RAS inhibitors, had no such effects on urinary ACE2 levels in an observational study.³³ However, the urine samples for ACE2 determination were collected only once, and thus the extent of changes in ACE2 levels due to antihypertensive agents could not be critically assessed. Taken together, no interventional studies have examined whether ARBs can definitely increase ACE2 activity in humans. Here, we demonstrated that the ARB olmesartan can increase urinary ACE2 levels and decrease plasma aldosterone levels in diabetic nephropathy patients with CKD. Many functional differences exist among ARBs that are not mediated by Ang II type 1 receptor blockade.³⁴ Long-term treatment of hypertensive patients with olmesartan reduces plasma Ang II levels, whereas several other types of ARBs have been reported to increase plasma Ang II levels in hypertensive patients.^35–37 These findings suggest that the increased ACE2 activity due to olmesartan contributes to reduced plasma Ang II levels by upregulating Ang-(1-7) formation.

Urinary ACE2 levels are associated with hyperglycemia, and insulin treatment has been observed to reduce urinary ACE2 levels in mice.³¹ However, no significant associations were observed in the present study between changes in urinary ACE2 levels and changes in glycemic indices, including plasma glucose and HbA1c levels, since only well-controlled patients were assessed. Although it has been reported that some calcium channel blockers, including benidipine, azelnidipine, and cilnidipine, reduced albuminuria in diabetic nephropathy,^38–40 many of the patients in the present study had been treated with amlodipine for at least 8 weeks at baseline, suggesting only a slight effect of amlodipine on decreasing the urinary albumin/Cr ratio and increasing urinary ACE2 levels. However, the changes in urinary ACE2 levels correlated significantly with urinary L-FABP levels in the present study. Significant urinary ACE2 levels most likely reflect the state of the proximal tubules, and differences may exist in patients with CKD and tubular injury compared with healthy individuals.⁴¹ In patients with diabetic nephropathy, the development of progressive diabetic kidney disease was associated with changes in proximal tubules, renal function, and prognosis and showed a higher correlation with structural lesions in the tubulointerstitium than with glomerular changes.⁴² Various proximal tubule pathophysiological stresses, such as proteinuria, induce upregulation of human L-FABP gene expression, thereby resulting in increased proximal tubular L-FABP excretion and increased urinary L-FABP excretion.⁴³ Urinary L-FABP is an important marker of tubulointerstitial changes in diabetic nephropathy, while oxidative stress is a pathogenic factor underlying diabetic complications such as nephropathy.^44–46 Because multivariate regression analysis showed that urinary L-FABP is an independent variable significantly associated with urinary ACE2 levels, olmesartan may have potential against the progression of tubulointerstitial damage due to increased urinary ACE2 levels. Nevertheless, further studies are needed to identify the factors associated with the increase in ACE2 levels, as ACE2 has the potential to be a marker of renal RAS activation in these patients.

Our study design was limited by the small sample size, short treatment duration, and lack of a control group since it was a preliminary investigation. Furthermore, we did not measure plasma levels of Ang-(1-7), Ang II, and ACE2, or urinary ACE2 activity, in this study, and thus the changes in ACE2 activity and their impact on the RAS may not have been fully characterized. Additional long-term studies are needed to accurately assess the renoprotective effects of olmesartan therapy in diabetes patients. Moreover, it should be clarified whether ARBs other than olmesartan have the same potential to increase urinary ACE2 levels.

In conclusion, olmesartan treatment leads to a reduction in albuminuria, urinary L-FABP, and plasma aldosterone levels in diabetes patients with CKD; however, it may have the unique effect of increasing urinary ACE2 levels. Some previous studies in mice and humans have shown that other RAS blockers do not increase urinary ACE2 levels. Whether this unique increase contributes to olmesartan’s renoprotective effect must be examined further in a future study.

Footnotes

Conflict of interest

None declared.

Funding

This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.

References

Donoghue

Hsieh

Baronas

. A novel angiotensin-converting enzyme-related carboxypeptide (ACE2) converts angiotensin I to angiotensin 1–9. Circ Res 2000; 87: 1–9.

Tipnis

Hooper

Hyde

. A human homolog of angiotensin-converting enzyme. Cloning and functional expression as a captopril-insensitive carboxypeptidase. J Biol Chem 2000; 275: 33238–33243.

Tikellis

Johnston

Forbes

. Characterization of renal angiotensin-converting enzyme 2 in diabetic nephropathy. Hypertension 2003; 41: 392–397.

Wysocki

Naaz

. Increased ACE2 and decreased ACE protein in renal tubules from diabetic mice: A renoprotective combination? Hypertension 2004; 43: 1120–1125.

Shaltout

Westwood

Averill

. Angiotensin metabolism in renal proximal tubules, urine, and serum of sheep: Evidence for ACE2-dependent processing of angiotensin II. Am J Physiol Renal Physiol 2007; 292: F82–91.

Wysocki

William

. Glomerular localization and expression of angiotensin-converting enzyme 2 and angiotensin-converting enzyme: Implications for albuminuria in diabetes. J Am Soc Nephrol 2006; 17: 3067–3075.

Reich

Oudit

Penninger

. Decreased glomerular and tubular expression of ACE2 in patients with type 2 diabetes and kidney disease. Kidney Int 2008; 74: 1610–1616.

Wysocki

Soler

. ACE and ACE2 activity in diabetic mice. Diabetes 2006; 55: 2132–2139.

Chodavarapu

Grobe

Somineni

. Rosiglitazone treatment of type 2 diabetic db/db mice attenuates urinary albumin and angiotensin converting enzyme 2 excretion. PLoS One 2013; 8: e62833.

10.

Kobori

Nangaku

Navar

. The intrarenal renin-angiotensin system: From physiology to the pathobiology of hypertension and kidney disease. Pharmacol Rev 2007; 59: 251–287.

11.

Senador

Kanakamedala

Irigoyen

. Cardiovascular and autonomic phenotype of db/db diabetic mice. Exp Physiol 2009; 94: 648–658.

12.

Riberio-Oliveria

Jr Nogueira

Pereira

. The renin-angiotensin-aldosterone system and diabetes: an update. Vasc Health Risk Manag 2008; 4: 787–803.

13.

Alghamri

Weir

Anstadt

. Enhanced angiotensin II-induced cardiac and aortic remodeling in ACE2 knockout mice. J Cardiovasc Pharmacol Ther 2012; 18: 138–151.

14.

Oudit

Herzenberg

Kassiri

. Loss of angiotensin-converting enzyme-2 leads to the late development of angiotensin II-dependent glomerulosclerosis. Am J Pathol 2006; 168: 1808–1820.

15.

Japanese Society of Hypertension: Japanese Society of Hypertension Guidelines for the Management of Hypertension (JSH 2009). Hypertens Res 2009; 32: 4–107.

16.

Xiao

Hiremath

Knoll

. Increased urinary angiotensin-converting enzyme 2 in renal transplant patients with diabetes. PLos One 2012; 7: e37649.

17.

Mizuiri

Aoki

Hemmi

. Urinary ACE2 in patients with CKD. Nephrology 2011; 16: 567–572.

18.

Matsuo

Imai

Horio

. Revised equations for estimated GFR from serum creatinine in Japan. Am J Kidney Dis 2009; 53: 982–992.

19.

Santos

Simoes

Silva

Maric

. Angiotensin-(1–7) is an endogenous ligand for the G protein-coupled receptor Mas. Proc Natl Acad Sci USA 2003; 100: 8258–8263.

20.

Ferrario

Varagic

The ANG-(1–7)/ACE2/mas axis in the regulation of nephron function. Am J Physiol Renal Physiol 2010; 298: F1297–1305.

21.

Iwai

Nakaoka

Senba

. Possible involvement of angiotensin-converting enzyme 2 and mas activation in inhibitory effects of angiotensin II type 1 receptor blockade on vascular remodeling. Hypertension 2012; 60: 137–144.

22.

Grobe

Mecca

Lingis

. Prevention of angiotensin II-induced cardiac remodeling by angiotensin-(1–7). Am J Physiol Heart Circ Physiol 2007; 292: H736–H742.

23.

Zhong

Basu

Guo

. Angiotensin-converting enzyme 2 suppresses pathological hypertrophy, myocardial fibrosis, and cardiac dysfunction. Circulation 2010; 122: 717–728.

24.

Wong

Oudit

Reich

. Loss of angiotensin-converting enzyme-2 (Ace2) accelerates diabetic kidney injury. Am J Pathol 2007; 171: 438–451.

25.

Soler

Wysocki

. ACE2 inhibition worsens glomerular injury in association with increased ACE expression in streptozotocin-induced diabetic mice. Kidney Int 2007; 72: 614–623.

26.

Wysocki

Rodriguez

. Targeting the degradation of angiotensin II-dependent hypertension. Hypertension 2010; 55: 90–98.

27.

Oudit

Liu

Zhong

. Human recombinant ACE2 reduces the progression of diabetic nephropathy. Diabetes 2010; 59: 529–538.

28.

Verma

Shan

Lei

. ACE2 and Ang-(1–7) confer protection against development of diabetic retinopathy. Mol Ther 2012; 20: 28–36.

29.

Liu

Wang

. Angiotensin-converting enzyme (ACE) 2 overexpression ameliorates glomerular injury in a rat model of diabetic nephropathy: a comparison with ACE inhibition. Mol Med 2011; 17: 59–69.

30.

Nadarajah

Milagres

Dilauro

. Podocyte-specific overexpression of human angiotensin-converting enzyme 2 attenuates diabetic nephropathy in mice. Kidney Int 2012; 82: 292–303.

31.

Wysocki

Garcia-Halpin

. Regulation of urinary ACE2 in diabetic mice. Am J Physiol Renal Physiol 2013; 305: F600–F611.

32.

Park

Kim

Park

. High urinary ACE2 concentrations are associated with severity of glucose intolerance and microalbuminuria. Eur J Endocrinol 2013; 168: 203–210.

33.

Furuhashi

Moniwa

Mita

. Urinary angiotensin-converting enzyme 2 in hypertensive patients may be increased by olmesartan, an angiotensin II receptor blocker. Am J Hypertens. Epub ahead of print 18 May 2014. DOI:10.1093/aih/hpu086.

34.

Kurtz

Pravenec

Molecule-specific effects of angiotensin II-receptor blockers independent of the renin-angiotensin system. Am J Hypertens 2008; 21: 852–859.

35.

Ichikawa

Takayama

Long-term effects of olmesartan, an Ang II receptor antagonist, on blood pressure and the renin-angiotensin-aldosterone system in hypertensive patients. Hypertens Res 2001; 24: 641–646.

36.

Gavras

Effects of eprosartan versus enalapril in hypertensive patients on the renin-angiotensin-aldosterone system and safety parameters: results from a 26-week, double-blind, multicenter study. Eprosartan Multinational Study Group. Curr Med Res Opin 1999; 15: 15–24.

37.

Bauer

Reams

. Effects of losartan on the renin-angiotensin-aldosterone axis in essential hypertension. J Hum Hypertens 1995; 9: 237–243.

38.

Abe

Maruyama

Suzuki

. L/N-type calcium channel blocker cilnidipine reduces plasma aldosterone, albuminuria, and urinary liver-type fatty acid binding protein in patients with chronic kidney disease. Heart and Vessels 2013; 28: 480–489.

39.

Abe

Okada

Suzuki

. T/L-type calcium channel blocker reduces the composite ranking of relative risk according to new KDIGO guidelines in patients with chronic kidney disease. BMC Nephrology 2013; 14: 135–144.

40.

Abe

Maruyama

Okada

. Additive anti-oxidative effects of azelnidipine on angiotensin receptor blocker olmesartan treatment for type 2 diabetic patients with albuminuria. Hypertens Res 2011; 34: 935–941.

41.

Mizuiri

Hemmi

Arita

. Expression of ACE and ACE2 in individuals with diabetic kidney disease and healthy controls. Am J Kidney Dis 2008; 51: 613–623.

42.

Thomas

Burns

Cooper

ME.

Tubular changes in early diabetic nephropathy. Adv Chronic Kidney Dis 2005; 12: 177–186.

43.

Kamijo

Kimura

Sugaya

. Urinary fatty acid-binding protein as a new clinical marker of the progression of chronic renal disease. J Lab Clin Med 2004; 143: 23–30.

44.

Sheetz

King

GL.

Molecular understanding of hyperglycemia’s adverse effects for diabetic complications. JAMA 2002; 288: 2579–2588.

45.

Nakamura

Sugaya

Kawagoe

. Effect of pitavastatin on urinary liver-type fatty acid-binding protein levels in patients with early diabetic nephropathy. Diabetes Care 2005; 28: 2728–2732.

46.

Abe

Matuyama

Okada

. Effects of lipid-lowering therapy with rosuvastatin on kidney function and oxidative stress in patients with diabetic nephropathy. J Atheroscler Thromb 2011; 18: 1018–1028.