Markers of oxidative protein damage in plasma and urine of type 2 diabetic patients.

This study aims to measure selected markers of oxidative protein damage (OPD) in patients with type 2 diabetes mellitus (T2DM) in order to estimate their utility as indicators of oxidative stress (OS) and to search for possible associations between them. The concentrations of advanced oxidation protein products (AOPP) and total sulphydryl (TSH) and reactive carbonyl (RCO) groups are measured in the plasma (P) and urine (U) of 60 patients and 22 controls using spectrophotometric methods. Significantly higher plasma concentrations of AOPP (P < 0.001), RCO groups (P < 0.01) and their P/U indexes (P < 0.001) as well as urinary levels of the RCO and TSH groups (P < 0.001) were observed in the diabetic patients compared with the controls. In contrast, the plasma levels and P/U index of the TSH groups were significantly lower (P < 0.001). A progressive increase in AOPP (plasma, urine and P/U index) in the course of albuminuria was noted, but significant differences among the subgroups of patients (with normoalbuminuria, microalbuminuria and macroalbuminuria) were found only in plasma. Plasma levels of all the measured parameters of OPD showed significant changes in T2DM patients compared with the control group. The largest increase was observed for AOPP. As the urinary AOPP concentration was not significantly different to that of the controls, it cannot be recommended as a marker of oxidative stress for monitoring the development of diabetic nephropathy. The P/U indexes did not provide any more information than the plasma concentrations of the studied markers.

Changes in chicken egg white cystatin concentration and isoforms during embryogenesis.

1. A higher concentration of cystatin, measured using the ELISA technique, was found in whites from unfertilised than fertilised eggs. 2. Up to day 8 of embryogenesis a decrease in cystatin concentration was observed, then at d 10 the concentration rose reaching the maximal value at d 14. 3. Immunoblot analyses showed both phosphorylated and nonphosphorylated isoforms of chicken cystatin in unfertilised egg white as well as in egg yolk and chicken serum. 4. Significantly lower immunostaining of the phosphorylated form after the 4th d of embryogenesis in egg white suggests its preferential transport into the yolk sac.

[Cystatin C as a marker of glomerular filtration rate]. / Cystatyna C jako wskaznik przesaczania klebuszkowego nerek.

The assessment of the glomerular filtration rate (GFR) is the most commonly used test of renal function. Cystatin C, a cysteine protease inhibitor, which can be measured by light scattering immunoassay, possesses many of the attributes required of the ideal GFR marker. This paper reviews so far obtained results dealing with cystatin C measurement, reference intervals and its diagnostic significance in nephropathy. Although serum cystatin C can generally be recommended as a marker of GFR, especially to detect mild GFR reductions, further clinical studies should be performed to confirm the validate in renal transplants and cancer patients.

[Comparison of cathepsin-B-like activity and CA 125 assays use in ovarian cancer diagnosis]. / Porównanie uzytecznosci diagnostycznej oznaczania aktywnosci katepsyno B-podobnej oraz stezenia CA 125 w surowicy krwi chorych na raka jajnika.

BACKGROUND: Determination of serum cathepsin B-like activity was used as a diagnostic marker of ovarian cancer. The aim of this study was to compare its diagnostic utility with CA 125 assay. METHODS: In 50 patients with epithelial ovarian cancer and 20 with ovarian cyst cathepsin B-like activity by fluorimety and CA 125 concentration by luminiscence immunoassay, were determined. The obtained results were compared by two cut-off values computed from the data obtained in healthy controls and ovarian cysts (cathepsin B: 11 and 108 mU/ml; CA 125: 35 and 135 U/ml), respectively. RESULTS: In differential diagnosis between ovarian carcinomas and ovarian cysts cathepsin B shows similar specificity (95%) as CA 125 but higher sensitivity (63.5% vs 50%). Both cathepsin B and CA 125 are good predictors of tumour response and recurrence CONCLUSIONS: Adding CA 125 determination to cathepsin B-like activity in the laboratory diagnostics of ovarian cancer increased the sensitivity and positive predictive values as compared to the use of cathepsin B alone.

[Levels of cystatin C, activity of antipapain and antitrypsin in plasma of patients with diabetes mellitus type 2]. / Stezenie cystatyny C, aktywnosc antypapainowa i antytrypsynowa osocza krwi u chorych na cukrzyce typu 2.

In plasma of 72 patients with diabetes mellitus type 2, cystatin C concentration, the antipapain as well as antitrypsin activities were determined. Statistically significant increase of levels of all the examined parameters (higher for inhibitors of cysteine proteases) was found in comparison to healthy persons. When considering types of vascular complications, the highest concentration of cystatin C and the strongest positive correlation with antipapain activity was found in the group of patients with microangiopathy. It might be connected with glomerular damages. Only antipapain as well as antitrypsin activities showed significant correlation with glycaemic control as measurement by glycated haemoglobin concentration.

[The role of cysteine protease and cystatin in pathogenesis and laboratory diagnosis of periodontal disease]. / Udzial proteaz cysteinowych i cystatyn w patogenezie i diagnostyce laboratoryjnej chorób przyzebia.

The paper presents a review of results concerning the role of host cysteine proteases, especially cathepsin B and cystatins in periodontal pathogenesis. Special attention was paid to the potential use of those biochemical parameters as biomarkers of periodontal disease activity.

Concentration of leukocyte elastase in plasma and polymorphonuclear neutrophil extracts in type 2 diabetes.

The concentration of leukocyte elastase/alpha1-proteinase inhibitor complexes in plasma and polymorphonuclear neutrophil extracts, and plasma trypsin inhibitory capacity were determined in 88 patients with type 2 diabetes and 47 control subjects. Higher values of these variables were found in patients as compared to controls (p < 0.001). The concentration of elastase was higher in obese patients than in lean ones (p < 0.05 for plasma, p < 0.01 for polymorphonuclear leukocytes). Only leukocyte elastase levels were significantly higher in the group with both micro- and macroangiopathy in comparison to the group with microangiopathy (p < 0.01) or macroangiopathy (p < 0.05) alone. Poor short-term glycaemic control was associated with higher elastase concentration in plasma and neutrophils (p < 0.05). The present study demonstrates that measurements of plasma polymorphonuclear neutrophil elastase level can be considered as a marker of development of diabetic angiopathy.

Plasma cystatin C concentration in non-insulin-dependent diabetes mellitus: relation with nephropathy.

We determined plasma concentrations of cystatin C, beta2-microglobulin - beta2-MG (low molecular mass protein markers of glomerular filtration rate - GFR), creatinine (marker of GFR) and urinary N-acetyl-beta-D-glucosaminidase (NAG) excretion (marker of glomerular and tubular dysfunction) in 41 non-insulin-dependent diabetic patients. A significant increase of all the measured parameters (p<0.001, p<0.05, p<0.05 and p<0.001, respectively) in comparison to the control group, was observed. In the patients with microalbuminuria, only plasma cystatin C concentration and urinary NAG excretion increased significantly in comparison to patients with normoalbuminuria. At a cut-off level of 1.74 mg/l for cystatin C and 1.81 U/g creatinine for NAG (95% percentile of the normoalbuminuric group), the sensitivity of the tests for detecting microalbuminuria was 82% for cystatin C and 86% for NAG. The specificities were 88 and 92%, respectively. The present study demonstrated that determination of plasma cystatin C might be useful in the detection of incipient diabetic nephropathy and is a potentially better marker than creatinine or beta2-MG. No correlation between parameters measured in plasma or urine and glycated hemoglobin was found.

[The role of cystatin in neoplastic progression and diagnosis]. / Rola cystatyn w procesie nowotworowym i jego diagnostyce.

Possible role of the cystatins, natural and specific inhibitors of the cysteine peptidases in cancer invasion is described. The value of cystatins determinations in serum and tissue extracts as diagnostic and prognostic parameters in human cancers is also presented.

[Dependence of serous activity of cathepsin B on histological types of ovarian carcinomas]. / Zaleznosc surowiczej aktywnosci katepsyny B od typów histologicznych raka jajnika.

The preoperative activity of cathepsin B was determined in the serum of 110 patients with primary ovarian carcinomas and in 35 women with benign ovarian tumor. The significant increase of cathepsin B activity in ovarian carcinomas is independent of its types in comparison to benign ovarian tumor. The percentage of the results with value above the cut-off values was the highest for serous types of ovarian carcinomas. The dependence between the increase of cathepsin B activity and the FIGO stage of the disease was observed only for serous and endometrioid type of ovarian carcinomas.

Cathepsin B-like activity as a serum tumour marker in ovarian carcinoma.

Serum cathepsin B-like activity was determined in 75 patients with ovarian carcinomas and in control groups. Ovarian cancers were of FIGO stages I-IV. Control groups consisted of 15 healthy women, 20 patients with myomas of the uterus, and 17 with benign ovarian cysts. Preoperative results showed elevated cathepsin B-like activity in 100% of the patients with ovarian cancers in relation to healthy subjects and patients with myomas, and in 78% in relation to benign ovarian cysts. Cathepsin B activity increased progressively with the FIGO stage of the disease, but the differences among particular stages were not statistically significant. In serous tumours cathepsin activity was significantly higher only in comparison to endometrioid ones (p < 0.001). Antipapain capacity of cystatins in the sera was also determined. No significant correlation between cathepsin B-like, and antipapain activity of cystatins was found. Serum cathepsin B-like activity may be helpful in the preoperative differential diagnosis between ovarian carcinomas and benign ovarian or uterine tumours.

The activities and subcellular localization of cathepsin B and cysteine proteinase inhibitors in human ovarian carcinoma.

Ovarian carcinomas exhibited a higher cathepsin B activity than did normal ovaries. The plasma membrane fractions of ovarian cancers were enriched in the activities of both cathepsin and cysteine proteinase inhibitors. The ratio of cathepsin to the inhibitors in these fractions was higher at stage IV than at stage III of the disease, classified according to the Federation of International Gynecology and Obstetrics.

[Contents of glycogen in neutrophils in patients with diabetes]. / Zawartosc glikogenu w granulocytach obojetnochlonnych chorych na cukrzyce.

Glycogen neutrophils level was evaluated in 54 patients with non-insulin dependent diabetes mellitus (NIDDM) and 10 patients with insulin dependent diabetes mellitus (IDDM). Glycogen concentration estimated by histochemical method was lower in diabetics than in control group. Patients with NIDDM were divided in the groups according to: sex, duration of disease, a kind of complications and a way of treatment. The glycogen contents in neutrophils, defined in "score"-unit was not different in isolated groups. There was found significant correlation between glycogen contents in neutrophils and the metabolic control in patients with IDDM (r = 0.72) and less significant in patients with NIDDM (r = 0.29).

Clearance of chicken cystatin from the rat circulation.

Chicken cystatin (mixed form) was prepared from egg white. Radioactively labeled preparations were administered intravenously to rats. 125I-cystatin disappeared from the rat circulation with a half-life of approximately 73 min. The radiolabeled inhibitor was rapidly taken up by the kidneys. Percoll density gradient analysis showed that it was incorporated into lysosomes. Within 24 hr after the injection of 125I-cystatin, 25% of the administered radioactivity was recovered in the urine, but only 2% was in the protein-bound form.

Purification and characterization of cystatin from duck egg white.

It is the second peptidase inhibitor, after ovostatin, which showing the same antipapain activity in egg white in different avian species implies differences in amino-acid sequences. Cystatin from duck egg white was purified by carboxymethylpapain affinity chromatography and size-exclusion HPLC. The purified inhibitor which showed partial identity in the immunodiffusion test with chicken egg white cystatin, had an apparent molecular mass of 9.3 kDa as determined by SDS/PAGE. IEF analysis revealed five molecular forms of pI in the range 7.8-8.4. The obtained cystatin was neither glycosylated nor phosphorylated as it is in the case of chicken cystatin. The determined Ki (0.005 +/- 0.001 nM) was similar to that reported for human and chicken cystatin C.

[Cathepsin D in diagnosis of neoplastic diseases]. / Znaczenie katepsyny D w diagnostyce choroby nowotworowej.

The paper presents a review of results concerning the aspartic proteinase cathepsin D and its role in tumor invasion and metastasis. Special attention was paid to the clinical prognostic value of cathepsin D determination in the breast cancer cytosol.

[Glycosylated proteins and various acute phase factors in patients with type II diabetes]. / Glikozylowane bialka i niektóre czynniki ostrej fazy u chorych na cukrzyce typu II.

In the plasma of patients with type II diabetes a statistically significant increase was found of the concentration of glycosylated proteins, haptoglobin and sialic acid, in relation to controls. The concentration of glycosylated proteins was determined by the fructosamine method and by a method based on reaction with thiobarbituric acid. The "r" correlation coefficient between both methods in the diabetics was 0.58. No significant correlation was found between fructosamine concentration and the determined acute phase factors. The statistical analysis of the obtained results related to the coexistence of macroangiopathy or microangiopathy, diabetes duration or treatment method showed only a significantly higher fructosamine concentration expressed as mM of fructose in relation to one gram of albumin when the group of diabetics with vascular complications was compared with that without these complications.

[Diagnostic value of determining alpha1 protease inhibitor and alpha2 macroglobulin in the serum of women with malignant and non-malignant ovarian tumors and uterine myomas]. / Wartosc diagnostyczna oznaczania alpha 1-inhibitora proteaz oraz alpha 2-makroglobuliny w surowicy krwi kobiet z guzami zlosliwymi i niezlosliwymi jajników oraz miesniakami macicy.

In the serum of 21 women with malignant ovarian tumours, 13 with uterine myomas and 16 with cysts the concentrations were determined of alpha 1 inhibitor of proteases and alpha 2 macroglobulin by immunodiffusion, and the functional activity of both inhibitors which made possible calculation of the specific activity (units of inhibitory activity per one gram of inhibitor protein). In women with malignant tumours the specific activity of alpha 1 inhibitor was decreased, suggesting that a part of the inhibitor was functionally inactive. This decrease was statistically significant in relation to the control group and the groups of myomas and cysts, but was independent of the degree of disease progression. The specific activity of alpha 2 macroglobulin was significantly decreased only in patients with grade IV degrees of clinical progression (according to FIGO) in relation to the control group and to the group with cysts.

Cathepsin H from human placenta.

Cathepsin H was isolated from human placenta by autolysis, acetone fractionation, and chromatography on DEAE-cellulose, Sephadex G-75, hydroxyapatite and concanavalin A-Sepharose. The enzyme gave on SDS-polyacrylamide gel electrophoresis two bands of Mr 25,500 and 28,500. Two active forms of the enzyme, with pI of 6.0 and 6.45, were obtained by isoelectric focusing. The enzyme is stable over the pH range 5-7.5, whereas it becomes inactive on heating to 50 degrees C. Cathepsin H of human placenta, like the enzyme from other sources, hydrolyses protein and naphthylamide substrates, showing within the latter group the strongest preference towards arginine-beta-naphthylamide (pH optimum 6.8). The enzyme is inhibited by the known inhibitors of cysteine proteases and by placental cystatins.