Mitsumine Fukui

mRNA Expression of Growth Factors in Glomeruli From Diabetic Rats

Evaluations of glomerular mRNA levels encoding for PCNA, TNF-α, PDGF-A and -B chains, TGF-β, IGF-I, bFGF, and EGF were made at 4, 12, and 24 wk after injection of STZ in Sprague-Dawley rats. The mRNA levels for PCNA, TNF-α, PDGF-B chain, TGF-β, and bFGF increased with age in STZ-induced diabetic rats. At 24 wk after STZ injection, mRNA levels for PCNA, TNF-α, PDGF-B chain, TGF-β, and bFGF were increased 3.8-fold, (P < 0.01), 4.2-fold (P < 0.01), 4.0-fold (P < 0.01), 5.2-fold (P < 0.001), and 3.6-fold (P < 0.01), respectively, in the glomeruli of diabetic rats when compared with control rats. In contrast, mRNA levels for IGF-I, PDGF-A chain, and EGF were not altered in glomeruli from diabetic and control rats throughout the experimental period. Insulin treatment partially ameliorated the increase in mRNA levels for PCNA, TNF-α, PDGF-B chain, TGF-β, and bFGF in the glomeruli of diabetic rats. These data indicate that alterations in growth factor mRNA levels in glomeruli may be a manifestation of diabetic nephropathy, and that hyperglycemia or insulin deficiency may play a role in abnormal growth factor gene regulation.

Effect of a Specific Endothelin Receptor A Antagonist on mRNA Levels for Extracellular Matrix Components and Growth Factors in Diabetic Glomeruli

We have previously reported that the mRNA levels of extracellular matrix (ECM) components including αl(I), αl(III), and αl(IV) collagen chains, laminin Bl and B2 chains, and growth factors including tumor necrosis factor (TNF)-α, platelet-derived growth factor (PDGF)-B chain, transforming growth factor (TGF)-β, and basic fibroblast growth factor (FGF) all increase with age in diabetic glomeruli. The present study was designed to assess whether glomerular expression of these mRNAs in rat diabetic glomeruli is affected by a specific endothelin receptor A antagonist, FR139317. Diabetes was produced by streptozotocin injection, and animals were divided into four groups: untreated diabetic rats, FR139317-treated diabetic rats, control nondiabetic rats, and FR139317-treated control rats. FR139317 treatment was continued for 24 weeks. FR139317 attenuated the rise in creatinine clearance (P < 0.01) and reduced urinary protein excretion (P < 0.01) in diabetic rats, but did not affect blood pressure. FR139317 attenuated the increases in glomerular mRNA levels of αl(I) (P < 0.01), α1(III) (P < 0.01), and α1(IV) (P < 0.01) collagen chains, laminin B1 (P < 0.01) and B2 (P < 0.01) chains, TNF-α (P < 0.01), PDGF-B (P < 0.01), TGF-β (P < 0.001) and basic FGF (P < 0.01) in diabetic rats. However, FR139317 did not affect these mRNA levels in glomeruli of control rats. These findings suggest that FR139317 may be useful in the treatment of diabetic glomerulopathy by reducing mRNA levels of ECM components and growth factors.

ECM Gene Expression and Its Modulation by Insulin in Diabetic Rats

The steady-state levels of mRNA encoding for the α1(IV) collagen chain, laminin B1 and B2 chains, basement membrane HSPG, and α1(I) and a1(III) collagen chains were examined in rat glomeruli at 4, 12, and 24 wk after injection of STZ. The mRNA levels for the α1(IV) collagen chain, laminin B1 and B2 chains, and α1 (I) and α1(I) and α1(III) collagen chains increased significantly with age in the STZ-induced diabetic rats before morphological thickening of basement membrane occurred. In contrast, the mRNA levels for HSPG decreased markedly 4 wk after STZ injection and then increased with age compared with those for control rats. The mRNA levels for these ECM components showed a continuous decline with age in controls. Treating the diabetic rats with insulin for 4 wk ameliorated the abnormally regulated ECM gene expression in the glomeruli. These data suggest that the abnormal regulation of ECM gene expression in the glomeruli may contribute to the expansion of mesangial matrix and basement membrane thickening in diabetic rats, and that hyperglycemia may play a role in the abnormal ECM gene expression.

Abnormal Gene Expression of Matrix Metalloproteinases and Their Inhibitor in Glomeruli from Diabetic Rats

The steady state levels of mRNA encoding for metalloproteinase (MMP)-1, -2, -3, and -9 and tissue inhibitor of metalloproteinase (TIMP)-1 were examined in glomeruli at 4, 12, and 24 weeks after the injection of streptozocin (STZ) in rats. The mRNA levels for MMP-1 and MMP-3 decreased with age in STZ-induced diabetes. At 24 weeks after STZ injection, mRNA levels for MMP-1 and MMP-3 fell to 40% (p < 0.01) and 20% (p < 0.01), respectively, in the glomeruli of diabetic rats when compared with control rats. In contrast, mRNA levels for TIMP-1 increased significantly with age in the diabetic glomeruli and reached an 8-fold (p < 0.01) increased at 24 weeks after STZ injection. mRNA levels for MMP-2 were not altered in glomeruli from diabetic and control rats throughout the experimental period, whereas those for MMP-9 were not detected in glomeruli from either group of rats. Insulin treatment partially ameliorated the decrease in mRNA levels for MMP-1 and MMP-3 and the increase in those for TIMP-1 in the glomeruli of diabetic rats. These data indicate that abnormal gene regulation of MMPs and TIMP-1 in the glomeruli of diabetic rats may contribute to the progression of glomerular lesions and that hyperglycemia or insulin deficiency may be associated with abnormal MMPs and TIMP-1 gene regulation.

Serum Cystatin C Is a More Sensitive Marker of Glomerular Function than Serum Creatinine

We determined the relationship between the levels of serum cystatin C or creatinine (s-Cr) and the grade of creatinine clearance (CCr) in patients with various glomerular diseases. Serum samples from 96 patients with glomerular diseases were obtained from our hospital. The levels of serum cystatin C were measured using the Dade Behring Cystatin C assay with the automated Dade Behring Nephelometer II (BNII). CCr levels were classified into six groups according to the Guidelines of the Japanese Society of Nephrology as follows: grade 1 (normal renal function); grade 2 (slight decrease of renal function); grade 3 (moderate decrease of renal function); grade 4 (severe decrease of renal function); grade 5 (renal failure), and grade 6 (uremia). The mean levels of serum cystatin C in grade 3 patients were significantly higher than those in grade 1. The mean levels of serum cystatin C in grades 4, 5 and 6 patients were also significantly higher than those in grade 1. However, the mean levels of serum Cr in grade 3 patients were not significantly higher than those in grade 1. The levels of s-Cr in grades 4, 5 or 6 patients were significantly higher than those in grade 1. In this study, an increase of serum cystatin C levels occurred earlier than that of s-Cr in various glomerular diseases. It appears that the levels of serum cystatin C may provide early prognostic marker of patients with various glomerular diseases rather than the levels of s-Cr.

Renal Expression of mRNAs for Endothelin-1, Endothelin-3 and Endothelin Receptors in NZB/W F1 Mice

Endothelin (ET)-1, ET-3, and ET receptors are widely distributed in the vascular system and the kidney. The present study was designed to measure the renal concentrations of ET-1, ET-3, and ET receptor mRNAs in NZB/W F1 mice at 8, 24, and 48 weeks of age. The renal concentration of ET-1 mRNA increased significantly in NZB/W F1 mice as their nephritis progressed, reaching, by 48 weeks, a 10-fold higher concentration than in control NZW mice. Renal ET-3 mRNA concentrations, however, remained unchanged. The renal concentrations of ET receptor A and B mRNAs in NZB/W F1 mice increased gradually with the progression of nephritis, reaching 5- and 3-fold higher concentrations, respectively, at 48 weeks of age than found in control mice. Transforming growth factor-beta (TGF-beta) and tumor necrosis factor-alpha (TNF-alpha) have been shown to stimulate ET-1 mRNA expression in cultured mesangial cells. We therefore also examined the concentrations of TGF-beta and TNF-alpha mRNAs in the renal tissues of NZB/W F1 mice, and found that, at 48 weeks of age, they were 10- and 8-fold higher, respectively, than in control NZW mice. At 24 weeks, NZB/W F1 mice were divided into two groups receiving either methylprednisolone (MPSL) or saline injections for the following 24 weeks. The development of the histologic lesions characteristic of lupus nephritis and the increased renal concentrations of ET-1, ET receptors, TGF-beta and TNF-alpha mRNAs were suppressed by MPSL treatment. These data suggest that ET and ET receptor gene transcription is upregulated in the renal tissues of NZB/W F1 mice and that the beneficial treatment of lupus nephritis with MPSL is accompanied by a reduction in the elevated concentrations of ET-1, ET receptors, TGF-beta and TNF-alpha mRNAs.

Effects of Antihypertensive Drugs or Glycemic Control on Antioxidant Enzyme Activities in Spontaneously Hypertensive Rats with Diabetes

The activities of glomerular intrinsic antioxidant enzymes (AOEs) were measured in a diabetic spontaneously hypertensive rat (SHR) model. The effects of antihypertensive drugs, i.e. captopril or triple therapy (hydralazine, reserpine, and hydrochlorothiazide), on glomerular intrinsic AOE activities in this model were evaluated. The effects of blood glucose control on the AOE activities were also determined. The aim of the present study was to determine whether activities of glomerular intrinsic AOEs might correlate with disease activity in diabetic SHR. This study showed a decrease of glomerular intrinsic AOE, i.e. Cu/Zn-SOD and Mn-SOD (SOD = superoxide dismutase), glutathione peroxidase, and catalase, activities in diabetic SHR. Glomerular Cu/Zn-SOD or Mn-SOD, glutathione peroxidase, and catalase activities in nondiabetic SHR were slightly lower than those in nondiabetic WKY rats. These activities in diabetic SHR were significantly improved after captopril or triple therapy or blood glucose control. The levels of urinary albumin excretion, creatinine clearance, and glomerular tuft areas in diabetic SHR were also improved after the therapy. It appears that hypertension and hyperglycemia may influence the glomerular intrinsic AOE activities, albuminuria, creatinine clearance, and glomerular tuft areas in diabetic SHR. Thus, it is indicated that control of blood pressure or blood glucose is a very important factor for preventing renal injuries in the diabetic SHR model.

Effects of the new hydroxy-3-methylglutaryl coenzyme a reductase inhibitor fluvastatin on anti-oxidant enzyme activities and renal function in streptozotocin-induced diabetic rats.

1. The effects of 11 week treatments with the new hydroxy‐3‐methylglutaryl coenzyme A (HMG‐CoA) reductase inhibitor fluvastatin on renal intrinsic anti‐oxidant enzyme (AOE) activities and renal function were evaluated in streptozotocin (STZ)‐induced diabetic rats.

Increase of Urinary Type IV Collagen in Normoalbuminuric Patients with Impaired Glucose Tolerance

Accessible online at: http://BioMedNet.com/karger Dear Sir, Microalbuminuria is the only early clinical sign of the subsequent diabetic nephropathy. However, it is also known that significant structural changes have already appeared even at the stage of microalbuminuria in non-insulin-dependent diabetes mellitus (NIDDM) patients. Thus, it is necessary to develop a more sensitive measurement for detecting the early stage of renal injury in patients with diabetic nephropathy. Since type IV collagen is the principal component of glomerular basement membrane and mesangial matrix, the levels of type IV collagen in sera and urinary samples may reflect the rate of its turnover, such as the balance of production by intrinsic renal cells and degradation by matrix proteinases in diseased kidneys. To investigate the alteration of renal turnover of type IV collagen in patients with impaired glucose tolerance (IGT), urinary type IV collagen (uIV) was measured by a highly sensitive one-step sandwich enzyme immunoassay (EIA) (Fuji Chemical Industries, Co. Ltd, Takaoka, Toyama, Japan) [1]. Diagnosis of NIDDM was made according to the criteria of the 75-gram oral glucose tol-

Effects of Anti hypertensive Drugs on Antioxidant Enzyme Activities and Renal Function in Stroke-Prone Spontaneously Hypertensive Rats

Abstract The reactive oxygen species has been proposed as a key mediator of the progression of renal injury associated with essential hypertension. Among the defense systems operating against the reactive oxygen species, superoxide dismutase, glutathione peroxidase, and catalase are the most important antioxidant enzymes (AOEs). In the present study, systolic blood pressure, renal function (creatinine clearance, urinary albumin, and N-acetyl-β D-glucosaminidase excretion), renal intrinsic AOE activities, and renal histopathology were determined in stroke-prone spontaneously hypertensive rats and Wistar Kyoto rats. The effects of a 20-week treatment using three antihypertensive drug regimens—captopril, a sulfhydryl-containing angiotensin-converting enzyme inhibitor; temocapril, a potent, non-sulfhydryl-containing angiotensin-converting enzyme inhibitor prodrug; and a conventional triple drug combination that includes a vasodilator (hydralazine, hydrochlorothiazide and reserpine)—on renal function, renal tissue, AOE activities, and renal histopathologic abnormalities were evaluated in stroke-prone spontaneously hypertensive rats. Renal function and AOE activities were lower in the stroke-prone spontaneously hypertensive rats than in the Wistar Kyoto rats. Normalization of systolic blood pressure using the antihypertensive drugs improved renal function and produced a nonuniform alteration in renal AOEs; only glutathione peroxidase activity increased significantly with the use of all three drug regimens. The mild renal histopathologic abnormality in stroke-prone spontaneously hypertensive rats was not altered by drug treatment. The improvement in renal function may be related to an increase in glutathione peroxidase activity, but no correlation was seen between renal function changes and alteration in activities of superoxide dismutase or catalase.