muramidase and Diabetic-Nephropathies

Diabetic nephropathy (DN) is one of the most common and serious complications of diabetes, and is the most important cause of death for diabetic patients. Baicalin (BAI) has anti-oxidative, anti-inflammatory and anti-apoptotic activities, which play a role in attenuating insulin resistance and protecting the kidney. Moreover, cell-specific targeting of renal tubular cells is an approach to enhance drug accumulation in the kidney.. Forty-five Sprague-Dawley rats were divided into four groups. A diabetes model was created using streptozotocin (STZ) intraperitoneally injection. The four groups included: Control group (n = 10), DN (n = 15), BAI treatment (BAI; n = 10) and BAI-LZM treatment (BAI-LZM; n = 10) groups. In the current study, the renoprotection and anti-fibrotic effects of BAI-lysozyme (LZM) conjugate were further investigated in rats with DN induced by STZ compared with BAI treatment alone.. The results suggest that BAI-LZM better ameliorates renal impairment, metabolic disorder and renal fibrosis than BAI alone in rats with DN, and the potential regulatory mechanism likely involves inhibiting inflammation via the nuclear factor-κB signaling pathway, inhibiting extracellular matrix accumulation via the transforming growth factor-β/Smad3 pathway and regulating cell proliferation via the insulin-like growth factor (IGF)-1/IGF-1 receptor/p38 Mitogen-activated protein kinase (MAPK) pathway. BAI and the kidney-targeted BAI-LZM can utilize the body's cytoprotective pathways to reactivate autophagy (as indicated by the autophagy markers mechanistic target of rapamycin and sirtuin 1 to ameliorate DN outcomes.. Our data support the traditional use of S. baicalensis as an important anti-DN traditional chinese medicine (TCM), and BAI, above all BAI-LZM, is a promising source for the identification of molecules with anti-DN effects.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Blood Glucose; Cholesterol; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Drug Carriers; Drug Delivery Systems; Fibrosis; Flavonoids; Insulin; Kidney; Malondialdehyde; Muramidase; Rats; Triglycerides

Diabetic nephropathy is the leading cause of chronic renal disease and one of the major causes of cardiovascular mortality. Evidence suggests that its progression is due to the chronic hyperglycemia consequent to the production and accumulation of advanced glycation endproducts (AGEs). Lysozyme was shown to posses AGE-sequestering properties and the capacity to reduce the severity of the early stage manifestations of the diabetic nephropathy. This study was aimed to contribute to the understanding the molecular mechanisms of lysozyme effectiveness in the diabetic nephropathy, using an in-vitro cellular model, represented by the HK-2 cells, human proximal tubular epithelial cells. Lysozyme significantly reduced the AGE-induced IL-6 mRNA and an ELISA assay showed also a decreased release of the functional protein with a dose-dependent trend. In addition, lysozyme prevented macrophage recruitment, suggesting its capacity to elicit an anti-inflammatory action. We may conclude that the protective action of lysozyme on the nephrotoxic effects of AGE may depend, at least in part, on its ability to prevent the production and release of inflammatory mediators, such as IL-6 and to reduce macrophage recruitment in the inflammatory sites.

Topics: Cell Line; Cell Movement; Cell Survival; Chemokine CX3CL1; Diabetic Nephropathies; Down-Regulation; Epithelial Cells; Glycation End Products, Advanced; Humans; Inflammation Mediators; Interleukin-18; Interleukin-6; Kidney Tubules, Proximal; Macrophage Activation; Macrophages; Muramidase; Recombinant Proteins; RNA, Messenger; Tumor Necrosis Factor-alpha; U937 Cells

Diabetic nephropathy is the leading cause of end-stage kidney disease in developed countries and is related to chronic hyperglycaemia. The increased production and tissue deposition of advanced glycation end products (AGE) are known to play a major role in the pathogenesis of diabetic kidney damage. This study was undertaken to determine if lysozyme (LZ), microencapsulated in orally administrable chitosan-coated alginate microspheres (MS), is effective against the early changes seen in the initial stages of diabetic nephropathy.. LZ-containing MS (MSLZ) and an equivalent dose (equidose) of nonencapsulated LZ were given as oral treatments. LZ was administered to Wistar rats for seven weeks after diabetes induction with streptozotocin.. The results showed that microencapsulated LZ treatment significantly reduced the concentration of serum AGE in the circulation and their deposition in the kidneys. Likewise, MSLZ significantly prevented the development of microalbuminuria compared with untreated diabetic rats. Furthermore, MSLZ significantly prevented the development of glomerular and renal hypertrophy as well as overexpression of AGE receptors (RAGE). An equidose of free LZ had little or no effect whatsoever.. Our study supports a relationship between serum AGE and nephropathy in diabetes, and suggests that orally administered microencapsulated LZ can exert kidney-protective activity in a diabetic animal model.

Topics: Albuminuria; Animals; Blood Glucose; Body Weight; Capsules; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Glycation End Products, Advanced; Glycosuria; Muramidase; Rats; Rats, Wistar

Lysozyme (LZ), a host-defense protein, contains an 18 amino-acid domain with high affinity binding for sugar-derived proteins or lipids, called advanced glycation endproducts (AGE), that are implicated in diabetes- and age-dependent complications (DC).. A) The effects of LZ on AGE- removal were tested in vivo. LZ was injected (200 ug/day, i.p., X2 weeks) in non-obese diabetic (NOD), db/db (+/+) mice, and non-diabetic, AGE-infused Sprague-Dawley rats. B) LZ: AGE interactions with macrophage-like T1B-183 cells (Mf) and mesangial cells (MC) were tested in vitro.. A) In NOD mice, LZ reduced the elevated basal serum AGE (sAGE) (p < 0.05), enhanced urinary AGE (uAGE) excretion by approximately 2-fold (p < 0.01), while it reduced albuminuria (UA), p < 0.005. In db/db mice, LZ infusion also reduced the elevated sAGE (p < 0.05), doubled uAGE excretion (p < 0.05), and decreased UA (p < 0.01). In addition, LZ maintained normal sAGE in normal rats infused with AGE-BSA, as it doubled the urinary AGE (uAGE) clearance (p < 0.01). B) LZ stimulated the uptake and degradation of (125) I-labeled AGE-BSA and (25) I-human serum AGE by Mf, while suppressing AGE-induced TNFalpha and IGF-I production. In MC, LZ suppressed the AGE-promoted PDGF-B, alpha1 type IV collagen, and tenascin mRNA levels, and restored the AGE-suppressed expression and activity of MMP-9, but not MMP-2.. LZ may act to: a) accelerate renal in-vivo AGE clearance, b) suppress macrophage and mesangial cell- specific gene activation in vitro, and c) improve albuminuria due to diabetes. These data suggest that LZ by sequestering AGEs may protect against diabetic renal damage.

Topics: Animals; Anti-Infective Agents; Chickens; Diabetes Mellitus, Type 1; Diabetic Nephropathies; Female; Glomerular Mesangium; Glycation End Products, Advanced; Humans; In Vitro Techniques; Kidney; Macrophages; Male; Mice; Mice, Inbred NOD; Middle Aged; Muramidase; Rats

Diabetic uremic sera contain excessive amounts of reactive advanced glycation endproducts (AGEs), which accelerate the vasculopathy of diabetes and end-stage renal disease. To capture in vivo-derived toxic AGEs, high affinity AGE-binding protein lysozyme (LZ) was linked to a Sepharose 4B matrix. Initial studies showed that > 80% of 125I-AGE-BSA was retained by the LZ matrix, compared with < 10% retained by a control matrix. More than 60% of AGE-lysine was captured by the LZ matrix, and the LZ-bound fraction retained immunoreactivity and cross-linking activity, but had little intrinsic fluorescence (370/440 nm). After passage through the LZ matrix, AGE levels in diabetic sera (0.37+/-0.04 U/mg) were significantly reduced to a level (0.09+/-0.01 U/mg; n = 10; P < 0. 0001) comparable with the level of normal human serum, whereas total protein absorption was < 3%. The AGE-enriched serum fraction exhibited cross-linking activity, which was completely prevented by aminoguanidine. Among numerous LZ-bound proteins in diabetic uremic sera, three major proteins "susceptible" to AGE modification were identified: the immunoglobulin G light chain, apolipoprotein J (clusterin/SP-40,40), and the complement 3b beta chain. These findings indicate that the LZ-linked AGE affinity column may serve as an efficient method for the depletion of toxic AGEs from sera, including specific AGE-modified proteins that may be linked to altered immunity, lipoprotein metabolism, and accelerated vasculopathy in renal failure patients with or without diabetes.

Topics: Adult; Aged; Amino Acid Sequence; Blood Proteins; Diabetic Nephropathies; Glycation End Products, Advanced; Humans; Immunoblotting; Middle Aged; Molecular Sequence Data; Muramidase; Serum Albumin, Bovine; Uremia

3-Deoxyglucosone (3-DG) has been identified as an intermediate of the Maillard reaction in vitro. We measured serum 3-DG levels using gas chromatography/mass spectrometry and found a marked elevation in serum 3-DG levels in uremic patients compared with healthy subjects. The uremic patients with diabetes showed significantly higher serum concentrations of 3-DG than those without diabetes. 3-DG was demonstrated to be a potent protein-cross-linking agent in the reaction with lysozyme, leading to browning, fluorescence formation and polymerization of the protein by formation of advanced glycation end products (AGE). The increase in serum 3-DG levels in the uremic patients suggests that 3-DG may be responsible for the development of uremic complications by promoting the formation of AGE.

Topics: Adult; Aged; Blood Proteins; Cross-Linking Reagents; Deoxyglucose; Diabetic Nephropathies; Female; Gas Chromatography-Mass Spectrometry; Humans; In Vitro Techniques; Maillard Reaction; Male; Middle Aged; Muramidase; Uremia

Early morning urine specimens were obtained from two groups of non-insulin dependent diabetic patients and a group (43 subjects) of normal controls. The diabetic patients were divided into two subgroups according to the degree of diabetic control as judged by their glycosylated haemoglobin (HbA1) levels (well-controlled, 47 subjects; poorly controlled, 51 subjects). The concentration of the low-molecular-weight enzyme (lysozyme) was determined in each urine specimen and related to the concentration of creatinine (lysozyme/creatinine). The mean urinary lysozyme concentration was higher in each of the two diabetic groups as compared with the control group. However, it was not significantly different between the two diabetic groups. These result suggest that there is no association between the degree of glycaemic control and tubular proteinuria.

Topics: Adult; Aged; Blood Glucose; Creatinine; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Female; Glycated Hemoglobin; Humans; Male; Middle Aged; Muramidase; Proteinuria

Renal tubular function was investigated in 98 non-insulin-dependent and 18 insulin-dependent diabetics under conditions of standard glycemic control. Mean urinary excretion of lysozyme, beta 2-microglobulin and N-acetyl-beta-D-glucosaminidase (NAG) in both Albustix-negative and positive patients were significantly elevated above the control range. The increased excretion of lysozyme, beta 2-microglobulin and NAG was found in 21, 55 and 62% of the normoalbuminuric patients, and in 40, 57 and 74% of the microalbuminuric patients, respectively. Besides the parameters cited above, urinary acid-soluble glycoprotein (ASP) was measured to assess its potential as an indicator of early renal dysfunction. Mean urinary ASP excretion was also elevated in both Albustix-negative and positive patients. The albumin/ASP ratio increased as nephropathy advanced. Such a mode of excretion was similar to those of low-molecular-weight proteins (lysozyme and beta 2-microglobulin). The results of multiple regression analysis showed that serum creatinine most highly correlated with the excretion of the urinary proteins except for NAG.

Topics: Acetylglucosaminidase; Adult; Albuminuria; beta 2-Microglobulin; Biomarkers; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Female; Humans; Kidney Tubules; Male; Middle Aged; Muramidase; Proteinuria; Regression Analysis

We measured the excretion rates of six urinary enzymes that either originate from the proximal renal tubule, like alanine aminopeptidase (EC 3.4.11.2), alkaline phosphatase (EC 3.1.3.1), gamma-glutamyltransferase (EC 2.3.2.2), and N-acetyl-beta-D-glucosaminidase (EC 3.2.1.30), or that are typical low-molecular-mass proteins, like lysozyme (EC 3.2.1.17) and pancreatic ribonuclease (EC 3.1.27.5). These rates were compared with those of total protein and albumin in urine of 36 insulin-dependent diabetic men and 30 healthy men. Seventeen of the diabetics had "clinical proteinuria," defined as excretion of more than 7.5 g of protein per mole of urinary creatinine (group B). Group A comprised the 19 diabetics without proteinuria. Except for gamma-glutamyltransferase, the excretions of enzymes and proteins were significantly higher in diabetics than in controls and were greater in group B than in group A. N-Acetyl-beta-D-glucosaminidase was the analyte most often increased in group A (89%), followed by albumin and alkaline phosphatase (each 32%). All patients in group B showed increased excretion of N-acetyl-beta-D-glucosaminidase. We conclude from the comparative data that this enzyme may be useful as an early predictor of diabetic nephropathy.

Topics: Acetylglucosaminidase; Adult; Albuminuria; Alkaline Phosphatase; Aminopeptidases; CD13 Antigens; Diabetes Mellitus, Type 1; Diabetic Nephropathies; gamma-Glutamyltransferase; Humans; Male; Muramidase; Proteinuria; Ribonuclease, Pancreatic

Seeking to study whether measurement of lysozyme (EC 3.2.1.17) in urine by a reliable radioimmunoassay can provide a suitable index of renal tubular function and how lysozymuria develops in temporal relation to proteinuria in diabetic nephropathy, we have compared the urinary excretion of lysozyme and beta 2-microglobulin with the 15-min excretion rate of phenolsulfonphthalein in 39 patients with Type 2 (non-insulin-dependent) diabetes and investigated the temporal relation between the onset of lysozymuria and proteinuria in 15 patients with Type 1 (insulin-dependent) diabetes. The concentrations of lysozyme and beta 2-microglobulin in urine increased in proportion to the decrease in the rate of excretion of phenolsulfonphthalein in these patients. The coefficient of correlation between lysozyme concentration and the 15-min excretion rate of phenolsulfonphthalein (r = -0.70) was higher than that between beta 2-microglobulin concentration and the 15-min excretion rate of phenolsulfonphthalein (r = -0.46). Abnormally high lysozymuria, suggesting the existence of tubular dysfunction, was demonstrated in six of the patients with Type 1 diabetes who showed no proteinuria or only a slight increase in urinary protein excretion. Lysozymuria may thus be added to a list of the indicators for diabetic nephropathy.

Topics: Adolescent; Adult; Aged; beta 2-Microglobulin; Child; Child, Preschool; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Humans; Kidney Tubules; Middle Aged; Muramidase; Phenolsulfonphthalein; Proteinuria

Currently, there is no effective treatment for diabetic microangiopathy. We have been examining, therefore, the effects of lysozyme on the renal conditions of spontaneous diabetic mice (NSY mice). We assessed the changes in glomerulus following the administration of lysozyme, using the thickness of the glomerular capillary basement membrane as an indicator. Littermate, male F19 NSY mice were divided into two groups. One of the groups (Group L) was treated with lysozyme; the other control group (Group C) with physiological saline. Group L received a daily intramuscular injection of lysozyme solution for 4 or 8 weeks. The thickness of the glomerular capillary basement membrane was measured in order to assess the effect of lysozyme administration. In the 4-week series, the thickness was 4783 +/- 1760 A in Group C and 3266 +/- 777 A in Group L, while in the 8-week series it was 6011 +/- 2043 A in Group C and 3540 +/- 431 A in Group L. In both series of Group L, a distinct inhibitory effect on the basement membrane thickening was found. The present findings suggest that lysozyme may be effective in human diabetic nephropathy. However its clinical usefulness must be confirmed in future studies.

Topics: Animals; Basement Membrane; Body Weight; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Extracellular Matrix; Kidney Glomerulus; Mice; Mice, Mutant Strains; Muramidase

Topics: Aged; Diabetic Nephropathies; Humans; Kidney Failure, Chronic; Kidney Function Tests; Leukocyte Count; Muramidase; Reagent Kits, Diagnostic; Urinary Tract Infections

A reversible lysozymuria indicative of proximal tubular damage to the kidney was found in three out of five patients with diabetic ketosis, and a persistent lysozymuria was found in many patients with diabetic nephropathy. There was no relation between lysozymuria and the degree of proteinuria, and lysozymuria was not due to urinary tract infection. The degree of lysozymuria could be used to assess the severity of diabetic nephropathy.

Topics: Creatinine; Diabetic Nephropathies; Glomerulonephritis; Multiple Myeloma; Muramidase; Proteinuria; Pyelonephritis

The excretion of alanine aminopeptidase in the urine is changed by several factors influencing the tubuli. The diabetics increasedly excrete alanine aminopeptidase in the urine, apparantly as an expression of an increased cell moulting of the tubuli. Whether the increased alanine aminopeptidase activity of the urine may indicate a diabetic nephropathy, is to be clarified only by means of bioptic investigations of the kidneys. Streptomycin can also like a nephrostasis as a sequel of a decompensated cor pulmonale increase the activity of alanine aminopeptidase of the urine.

Topics: Aminopeptidases; Diabetes Mellitus; Diabetic Nephropathies; Diabetic Retinopathy; Female; Humans; Male; Muramidase; Sex Factors

Topics: Adult; Aged; Albuminuria; Antibody Formation; Blood Glucose; Creatinine; Diabetes Mellitus; Diabetic Nephropathies; Female; Growth Hormone; Humans; Insulin; Kidney; Male; Metabolic Clearance Rate; Middle Aged; Muramidase; Proteinuria