Dubravka Juretić

Effect of ‘antidiabetis’ herbal preparation on serum glucose and fructosamine in NOD mice

The antihyperglycemic effect of the Antidiabetis herbal preparation ((Myrtilli folium (Vaccinium myrtillus L.), Taraxaci radix (Taraxacum officinale Web.), Cichorii radix (Cichorium intybus L.), Juniperi fructus (Juniperus communis L.), Centaurii herba (Centaurium umbellatum Gilib.), Phaseoli pericarpium (Phaseolus vulgaris), Millefollii herba (Achillea millefolium L.), Morii folium (Morus nigra L.), Valeriane radix (Valleriana officinalis L.), Urticae herba et radix (Urtica dioica L.)), patent No. P-9801091 Zagreb, Croatia was investigated. Two extracts were prepared: ethanol extract (extract 1), and ethanol extract from which ethanol was evaporated on a rotatory evaporator at a temperature of 45 degrees C (extract 2). Extract 1 and extract 2 were administered (in experiment 1) to alloxan-induced non-obese diabetic (NOD) mice in the same dose of 20 mg/kg. Blood glucose was determined before, and 10, 30, 60 and 120 min after the preparation administration. Extract 1 and extract 2 decreased the level of blood glucose by 10 and 20%, respectively, of the initial value (at 0 min, mean = 22.6 +/- 8.3 mmol/l). Serum levels of glucose and fructosamine were determined in NOD mice, NOD mice administered extract 2 in a dose of 20 mg/kg of extract 2, and NOD mice administered acarbose in a dose of 25 mg/100 g chow, in order to verify the hypoglycemic action of extract 2 (in experiment 2). Extract 2 and acarbose were admixed to the chow. The duration of treatment was 7 days. Significantly lower glucose (P < 0.05) and fructosamine (P < 0.001) levels were recorded in extract 2 treated NOD mice as compared with NOD mice. Study results showed extract 2 to significantly decrease the level of glucose and fructosamine in alloxan induced NOD mice. Our future studies will be focused on the search of active principles of the extracts.

Follicular fluid contents of hyaluronic acid, follicle-stimulating hormone and steroids relative to the success of in vitro fertilization of human oocytes * †

OBJECTIVES To determine the concentrations of hyaluronic acid, FSH, P, and E2 in the follicular fluid (FF) obtained from IVF-ET patients and to assess the value of these measurements in predicting the outcome of fertilization. DESIGN One hundred eleven samples were retrospectively analyzed for the hyaluronic acid and hormone contents. SETTING University-based tertiary care center. PATIENTS Preovulatory FF samples were collected from 67 women undergoing IVF-ET treatment because of tubal absence or obstruction. MAIN OUTCOME MEASURES The FF hyaluronic acid and hormone concentrations were compared according to the type of ovulation induction, follicular development, and IVF outcome. RESULTS According to the type of ovulation induction, a significantly lower hyaluronic acid concentration was found in FF harvested from the patients treated with GnRH agonist-hMG. No significant correlation was found between FF hyaluronic acid and either morphological maturity of the oocyte-cumulus complex or fertilizability of oocytes. The level of FSH was significantly higher in FF, yielding a mature oocyte-cumulus complex and from which the oocyte obtained successfully fertilized and cleaved. A significant increase in the E2 concentration was found in FF in which mature cumuli oophori were present. The levels of hyaluronic acid significantly correlated with FSH in FF. CONCLUSIONS Expansion of the human oocyte-cumulus cell complex is an FSH-dependent phenomenon. The data are in agreement with the hypothesis that intrafollicular FSH plays an important role in the secretion of hyaluronic acid by granulosa cells and may act synergistically with E2 to enhance cytoplasmic maturation, resulting in successful fertilization.

Altered distribution of urinary glycosaminoglycans in diabetic subjects

Abstract. We studied the influence of type 1 and type 2 diabetes mellitus with similar duration on the urinary excretion of total glycosaminoglycans and alteration of urinary glycosaminoglycan distribution pattern. Investigations were performed in the 24-hour urine samples of 31 children with type 1 diabetes mellitus, 36 adults with type 2 diabetes mellitus and in 30 age-matched controls for each group. We found that type 2 diabetes mellitus also induced an increased urinary excretion of total glycosaminoglycans and that both type 1 and type 2 diabetes alter the urinary distribution of heparan sulphate and dermatan sulphate. Observed changes correlate with duration of the disease. Microalbuminuria was detected in 9 of 36 type 2 adult diabetics (25%). The microalbuminic group had a significanlty higher heparan sulphate and/or dermatan sulphate excretion rate. To clarify whether an altered urinary distribution of heparan sulphate and dermatan sulphate may precede the development of microalbuminuria, it is necessary to performed a prospective study in which urinary glycosaminoglycans and microalbuminuria are measured year by year starting from the diagnosis of diabetes.

Effect of non-genetic factors on paraoxonase 1 activity in patients undergoing hemodialysis

OBJECTIVES Hemodialyzed patients have lower paraoxonase 1 (PON1) activity. Higher mortality risk from cardiovascular disease observed in these patients could be due to the low antiathetrogenic activity of PON1. Understanding the mechanism that causes lower PON1 activity could provide the possibility for modulation of enzyme activity in purpose of preventing and/or decreasing development of atherosclerosis. DESIGN AND METHODS 87 healthy individuals and 71 hemodialyzed patients were enrolled in this study. RESULTS Hemodialyzed patients had reduced PON1 paraoxonase and arylesterase activity, concentrations of HDL, HDL(3) and HDL(2) and concentrations of free thiol groups. Distribution of HDL subfractions and distribution of PON1 phenotypes as well as concentrations of MDA were not different between two study groups. In the in vitro experiment high concentrations of urea, creatinine, uric acid and addition of patients sera ultrafiltrate did not significantly affect PON1 paraoxonase activity. CONCLUSION Decreased HDL concentration as well as lower PON1 concentration (shown indirectly by the enzyme arylesterase activity) might contribute, at least partly, to the reduced PON1 activity observed in hemodialyzed patients. Decreased concentration of free thiol groups in sera suggest that free thiol group (Cys284) in PON1 might also be oxidized, which can affect PON1 activity.

Tryptophan content in serum and brain of long-term insulin-treated diabetic rats.

SummaryThe metabolism of tryptophan (TRP) was studied in diabetic and insulin-treated diabetic rats throughout a five-month period. In alloxan diabetic rats the serum and brain TRP levels were decreased (serum: 38 to 48 mmol/l, brain: 8.6 to 9.2 mmol/g) in comparison to the values of control rats (serum: 59 to 64 mmol/l, brain: 11.3 to 12.6 mmol/g). Daily long-term (for 45, 75, 90 or 135 days) treatment with intermediatly acting insulin (4 IU/rat, s.c.) was not able to restore brain concentration of TRP. On the contrary, the serum TRP concentrations were totally or partially restored. The concentrations of branched chain amino acids (BCAA) were increased in serum (valine=361.2 to 461.0 μmol/l or leucine + isoleucine=431.0 to 520.3 μmol/l) through-out the entire five-month examination period. Insulin treatment did not return serum concentration of BCAA to normal level in the observation period either.

Lipid status, paraoxonase-1 activity and metabolic parameters in serum of heifers and lactating cows related to oxidative stress

The objective of this study was to investigate serum lipids, metabolic parameters and activity of the anti-oxidative enzyme paraoxonase-1 (PON1). The study was conducted on non-pregnant heifers with optimal health status and on healthy dairy cows in the period of intensive lactation, assuming that the energy and metabolic demands during lactation reduce anti-oxidative protection. Total cholesterol and HDL-cholesterol concentrations were significantly higher (P<0.05) in lactating cows than in heifers. Bilirubin concentration and γ-glutamyltransferase (GGT) activity were also significantly higher in lactating cows (P<0.05), indicating increased hepatic efforts of cows to meet energy requirements for lactation. Significantly lower PON1 activity and PON1/HDL ratio in lactating cows compared to heifers (P<0.05) showed that metabolic efforts during pregnancy, parturition and lactation influence PON1 activity due to oxidative stress. Concurrent increase in total and HDL-cholesterol during lactation indicated that the HDL particle is a major carrier of cholesterol in cows.

A liver urocanase deficiency.

Two sisters with a rare inborn error of histidine metabolism resulting from urocanase deficiency are being presented. The more common form of familial histidinemia due to histidase deficiency is excluded. The urocanase deficiency is proven by demonstrating increased excretion of metabolites of the product of the urocanase enzyme action. Further, the strongest evidence for the urocanase defect rests on the demonstration of urocanase deficiency and normal histidase activity in liver.

Polymorphisms of pon1 and pon2 genes in hemodialyzed patients

OBJECTIVES Q192R, L55M and -108C>T polymorphisms of pon1 gene affect PON1 paraoxonase activity while S311C polymorphism of pon2 gene might be associated with coronary heart disease. The aims of this study were to determine the frequencies of Q192R, L55M, -108C>T and S311C polymorphisms in hemodialyzed patients and to examine the relationship between pon1 gene polymorphisms and PON1 paraoxonase activity in those patients. DESIGN AND METHODS The study included 238 control subjects and 263 hemodialyzed patients. RESULTS PON1 paraoxonase activity was lower in patients. Genotype frequencies were different between two compared groups only for L55M polymorphism, with control group having higher frequency of MM genotype. Polymorphisms of pon1 gene were associated with significant variation in PON1 paraoxonase activity in both study groups. CONCLUSION Our results suggest that Q192R, L55M and -108C>T polymorphisms are not by itself the causal factors leading to the lower PON1 paraoxonase activity in hemodialyzed patients.

Applicability of common reference intervals for serum creatinine concentrations to the Croatian population

Abstract Background: In accordance with an ongoing activity for worldwide harmonization based on traceability in laboratory methods, the goal of this study was to validate the applicability of recommended “common” reference intervals for serum creatinine concentrations using a specific enzymatic method to the Croatian population. Methods: The reference group consisted of 240 healthy subjects (120 males and 120 females), between 18 and 74 years of age (median 57 years), who were selected in accordance with the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) recommendations. Creatinine in serum was measured using the creatinine enzymatic assay (Olympus OSR61204) that was standardized to the isotopic dilution mass spectrometry (IDMS) method and National Institute of Standards and Technology (NIST) Standard Reference Material (SRM) 967. In addition, creatinine was measured using a kinetic Jaffe method (Olympus OSR6178) standardized to NIST SRM 909b level 2 standard. Results: Method comparison between enzymatic creatinine (x) and the Jaffe kinetic method (y) gave the following P/B equation for the entire group (n=240): y=1.00x+17.00; r=0.968. Reference intervals for serum creatinine (central 95th percentiles) obtained using the enzymatic creatinine method ranged from 54 to 107 μmol/L for males and from 50 to 93 μmol/L for females. The IFCC recommended common reference intervals for global applications are 64–104 μmol/L and 49–90 μmol/L for males and females, respectively. Conclusions: Comparability of obtained results confirmed the applicability of recently recommended “common” reference intervals to the Croatian population for all laboratories measuring serum creatinine concentrations using enzymatic methods traceable to the IDMS method and NIST SRM 967. Clin Chem Lab Med 2010;48:231–5.

Effect of different dialysis membranes on erythrocyte antioxidant enzyme levels and scavenger systems related to free hemoglobin in serum of hemodialysis patients

Abstract The main objective of the present study was to analyze some aspects of the function and structure of erythrocytes with respect to hemodialysis membrane biocompatibility throughout the erythrocyte and serum antioxidant levels. The study included 36 hemodialysis patients (14 female and 22 male, age 22–79 years, median 55) treated at the Department of Nephrology and Dialysis, Split Clinical Hospital in Split, and 30 control subjects matched for age and sex. Hemodialysis was performed three times a week for 4 hours with cellulose diacetate (n = 17; 6 females and 11 males) or polysulfone (n = 19; 8 females and 11 males) membranes. The aim of the study was to assess the level of oxidative stress in these patients by measuring catalytic concentrations of superoxide dismutase and glutathione peroxidase in erythrocyte lysate and scavenger systems related to free hemoglobin in serum (haptoglobin, hemopexin and bilirubin). In comparison with control values, the mean catalytic concentrations of superoxide dismutase were increased and catalytic concentrations of glutathione peroxidase decreased in patients before hemodialysis irrespective of the membrane used. Immediately after hemodialysis with either membrane, the mean catalytic concentrations of superoxide dismutase returned to the control range, while those of glutathione peroxidase were still decreased compared to control values, without any significant difference between the cellulose diacetate and polysulfone membranes. The predialysis and postdialysis values of haptoglobin, hemopexin and bilirubin in patient sera were within the range of control values. Comparison of the cellulose diacetate and polysulfone membranes showed no significant differences in the erythrocyte content of antioxidant enzymes and the scavenger system related to free hemoglobin in serum before and after hemodialysis.