Potential use of serum insulin-like growth factor 1 and E-cadherin as biomarkers of colorectal cancer.

AIM: Despite many efforts, reliable biomarkers for the prediction and diagnosis of colorectal cancer (CRC) are still missing. Insulin-like growth factor 1 (IGF-1) and E-cadherin are recognized as potential biomarkers, but their diagnostic capacity is largely unexplored in CRC. The aim of this work is to investigate IGF-1 and E-cadherin levels with respect to various characteristics of CRC and to estimate their diagnostic potential. METHOD: Seventy CRC patients and 75 healthy individuals were enrolled. IGF-1 and E-cadherin were determined using enzyme-linked immunosorbent assay. The predictive and diagnostic capacities of IGF-1 and E-cadherin were estimated by logistic regression analysis and by determination of the area under the receiver operating characteristic (ROC) curve (AUC). RESULTS: Concentrations of IGF-1 were lower (P = 0.019) while levels of E-cadherin were higher (P < 0.001) in CRC patients than in controls. IGF-1 concentration decreased in parallel with age and progression of CRC (P = 0.023). Also, IGF-1 was higher in men with CRC than in women (P = 0.003). E-cadherin levels were unaffected by variations in either anthropometric characteristics of CRC patients, or localization, grade and stage of the tumour. Both IGF-1 and E-cadherin were independently associated with CRC (P = 0.040; P < 0.001, respectively). The diagnostic accuracy of IGF-1 was estimated as acceptable (AUC = 0.757; P < 0.001), while the diagnostic accuracy of E-cadherin was outstanding (AUC = 0.954; P < 0.001). CONCLUSIONS: Decreased IGF-1 and increased E-cadherin levels were found in CRC patients. IGF-1, but not E-cadherin, concentrations differed according to age, gender and stage of CRC. Both markers were independently associated with the presence of the disease, while E-cadherin demonstrated high diagnostic accuracy.

Effects of monacolin K-containing nutraceutical on cholesterol homeostasis re-establishment and CVD risk reduction in hypercholesterolemic subjects.

OBJECTIVE: Hypercholesterolemia is caused by cholesterol homeostasis (CH) disruption, and it contributes to cardiovascular diseases pathogenesis and progression. Status of CH can be assessed by measuring serum concentrations of non-cholesterol sterols (NCS) which serve as cholesterol synthesis and absorption surrogate markers. Monacolin K, isolated from red yeast rice, influences cholesterol synthesis by inhibiting HMG-CoA reductase activity and reduces serum total cholesterol (TC) concentration. PATIENTS AND METHODS: This longitudinal study included 30 hypercholesterolemic patients, with systematic coronary risk estimation (SCORE) values <10%, who received 3-months-long supplementation with nutraceutical mixture containing monacolin K, and vitamins C, B1 and K2. Serum NCS were quantified by HPLC-MS/MS method. Atherogenic indexes were calculated from lipid status parameters concentrations. Albumin degradation inhibition test was conducted to estimate in vitro anti-inflammatory activity of the nutraceutical mixture, whereas in vitro antioxidant activity was measured in serum enriched with prooxidants and antioxidants. RESULTS: TC, LDL-cholesterol (LDL-C), and triglycerides (TG) concentrations (p<0.001), as well as atherogenic indexes and SCORE values (p<0.001, p<0.01, respectively) were lowered following the supplementation. Concentrations of cholesterol synthesis markers were decreased (p<0.001), whereas levels of cholesterol absorption markers remained unchanged after the supplementation. Reduction in cholesterol synthesis went alongside reductions in lipid status parameters and atherogenic indexes. In vitro analyses showed certain anti-inflammatory and antioxidant activity of the nutraceutical. CONCLUSIONS: These results suggest that supplementation with monacolin K containing nutraceutical favorably influences lipid status parameters and atherogenic indexes by acting on cholesterol synthesis. Anti-inflammatory and antioxidant effects of this unique nutraceutical mixture may exhibit beneficial pleiotropic effects.

Endocan and advanced oxidation protein products in adult population with hypertension.

OBJECTIVE: Hypertension is closely related to oxidative stress and inflammation. Endocan is a new inflammation marker whose role is not completely elucidated in hypertension. The aim of this study was to explore the association between endocan and several oxidative stress markers [i.e., advanced oxidation protein products (AOPP), total protein sulfhydryl (SH-) groups and prooxidant-antioxidant balance (PAB)] in adult population with hypertension. PATIENTS AND METHODS: A total of 90 patients with hypertension were compared with 44 controls. Blood pressure, anthropometric and biochemical parameters were measured. Associations of clinical data with hypertension were tested with univariable and multivariable logistic ordinal regression analysis. RESULTS: Endocan and AOPP were significantly higher in hypertensive patients than in the controls (p=0.006 and p=0.046, respectively). In the multivariable logistic regression analysis, AOPP and endocan kept their independent positive associations with hypertension. As AOPP rose by 1 µmol/L and endocan rose by 1 pg/mL, the probability for hypertension presence rose by 4.2% and 32.2%, respectively and 39.9% of variation in hypertension could be explained with the Model. The area under the Receiver Operating Characteristic curve [(AUC) for AOPP=0.638 (0.550-0.719), p=0.01 and for endocan=0.679 (0.593-0.757), p<0.001] demonstrated sufficient clinical accuracy towards hypertension. On the contrary, the Model showed very good clinical accuracy [AUC= 0.825 (0.749-0.900), p<0.001]. CONCLUSIONS: Endocan and AOPP are independently correlated with hypertension in adult population and these tested markers together could be reliable parameters to discriminate patients with hypertension from normotensive ones.

Total oxidant status and oxidative stress index as indicators of increased Reynolds risk score in postmenopausal women.

OBJECTIVE: Considering the knowledge gap between underlying pathophysiological mechanisms of oxidative stress and increased cardiovascular risk, the present study aimed to examine the potential relationship between total oxidant status (TOS), total antioxidant status (TAS) and oxidative stress index (OSI) and the Reynolds Risk Score (RRS) in the cohort of postmenopausal women. PATIENTS AND METHODS: A total of 126 postmenopausal women participated in this cross-sectional study. Blood pressure, anthropometric and biochemical markers were determined. OSI was calculated as the TOS/TAS ratio. Associations of biochemical parameters with RRS were tested using univariable and multivariable logistic ordinal regression analysis. RESULTS: TOS and OSI were the highest in women in high RRS category compared to moderate and low risk ones (p<0.001, for both). There was no difference in TAS level across RRS categories (p=0.370). Multivariable ordinal regression analysis showed independent association of TOS and OSI with RRS when tested with other clinical variables [OR=2.45; 95% CI (1.08-5.53); p=0.031 and OR=2.84; 95% CI (1.27-6.36); p=0.011, respectively]. CONCLUSIONS: TOS and OSI are associated with the RRS in the cohort of postmenopausal women. Longitudinal studies are needed to confirm whether adding the TOS and OSI to the standard RRS algorithm could improve its potential to predict cardiovascular event.

Lipid profile and lipid oxidative modification parameters in the first trimester of high- risk pregnancies - possibilities for preeclampsia prediction.

OBJECTIVES: The goal of this study was to investigate metabolic changes in lipids and oxidative stress parameters in the first trimester of pregnancy with the more specific aim of estimating the significance and strength of researched parameters in the prediction of preeclampsia. DESIGN AND METHODS: The study included 87 high-risk pregnant (HRG) female subjects, 14 with developed preeclampsia (PEC) and 43 healthy pregnant female subjects matched for gestational age (CG). Thiobarbituric acid-reactive substances (TBARS) concentration, lipid hydroperoxides (LOOH), pro-oxidant antioxidant balance (PAB) and total oxidative status (TOS) were measured as oxidative stress markers, while total antioxidant capacity (TAC) was measured as an antioxidative defense parameter. The Atherogenic Index of Plasma (AIP) was calculated as the base 10 logarithm of the ratio of the plasma concentration of triglycerides (TG) to the plasma concentration of high-density lipoprotein cholesterol (HDL-C), with each concentration expressed in mmol/L. RESULTS: The results have shown that lipid indices, especially AIP, were significantly higher in the first trimester of HRG (p < 0.001) and PEC (p < 0.001). Oxidative stress parameters were significantlly higher, while TAC was significantly lower in HRG vs. CG [0.7 ± 0.15 vs 1.1 ± 0.16; (p < 0.001)] and in PEC [0.6 ± 0.12 vs 1.1 ± 0.16; (p < 0.001)] vs. CG. Also, in the HRG, results have shown an independent association of AIP with the preeclampsia development (p < 0.05), while placental growth factor did not show the expected level of significance (p = 0.648). Analysis of the Receiver Operating Characteristics (ROC) curves indicated that certain parameters included in the research model have very good diagnostic accuracy for preeclampsia (AUC = 0.856). CONCLUSIONS: AIP is associated with high-risk pregnancies. Furthermore, our results firmly underscored AIP as a potential marker for preeclampsia prediction.

Inverse association between serum endocan levels and small LDL and HDL particles in patients with type 2 diabetes mellitus.

OBJECTIVE: Determination of lipoprotein size and subclasses distribution can provide more significant information on cardiovascular disease risk than measurement of traditional lipid parameters alone. Accordingly, we aimed to examine their potential relationship with the novel biomarker of endothelial dysfunction, such as endocan in patients with type 2 diabetes mellitus (T2D), since there are no studies concerning this issue. PATIENTS AND METHODS: This case-control study included a total of 42 individuals with T2D and 64 diabetes-free participants. Serum endocan, lipid parameters, and lipoprotein subclasses were measured. RESULTS: Patients with T2D exhibited higher proportion of the smallest high-density lipoprotein (HDL) particles HDL 3c, as compared with diabetes-free participants (p=0.047). Higher serum endocan levels in T2D patients with low small dense low-density lipoprotein (LDL) particles (sdLDL) %, as compared with corresponding group of diabetes-free subjects was shown (p<0.01). Univariate binary logistic analysis revealed significant positive association of endocan and LDL diameter (OR=1.686, p=0.004), and negative associations of endocan with proportions of sdLDL (OR=0.928, p=0.007) and HDL3b (OR=0.789, p=0.009) particles. In a multivariate analysis, LDL diameter and proportions of sdLDL and HDL3b subclasses remained independent predictors of endocan levels in tested population. CONCLUSIONS: The results of our study showed that larger LDL diameters, but lower sdLDL and HDL3b proportions were associated with higher endocan levels in population with T2D. More studies in the future are needed to confirm the observed relationship and to examine its causal nature.

Effect of bromelain on alkaline phosphatases of intestinal and non-intestinal tissues and serum.

Human alkaline phosphatases extracted with butanol from liver, kidney and placenta, and from foetal and adult small intestine each contain fragments with molecular masses within the range of approximately 8 kDa to 20 kDa which can be removed by digestion with bromelain. However, in the case of adult intestine, this fragment (which is presumed to represent a membrane-binding domain) can only be demonstrated in tissue extracted immediately after removal at operation. Similar fragments are also present in foetal intestinal phosphatase in amniotic fluid, and in liver and bone alkaline phosphatases recovered from serum. Again, however, adult intestinal phosphatase from serum differs in the absence of the bromelain-sensitive fragment. These observations indicate differences in the ways in which intestinal and non-intestinal alkaline phosphatases gain access to the circulation, and also have implications for structural studies on intestinal phosphatase extracted post mortem from adult tissue.

Spectrophotometric assay of xanthine oxidase with 2,2'-azino-di(3-ethylbenzthiazoline-6-sulphonate) (ABTS) as chromogen.

A new method for the determination of xanthine oxidase activity, based on the oxidation of 2,2'-azino-di(3-ethylbenzthiazoline-6-sulphonate) (ABTS) by use of uricase and peroxidase, is described. The absorbance increase of the oxidized form of ABTS, measured after 10 min at 410 nm is proportional to xanthine oxidase activity. The method is sensitive, precise (CV below 8.3%), and linear up to 20 U/l. The analytical recovery of the ABTS-method was quantitative. Comparison with the UV and colorimetric NBT-method gave good correlation (r greater than or equal to 0.984). Reference values for serum xanthine oxidase activities determined with the new ABTS-method on 83 healthy persons are 0 to 1.20 U/l.

Low serum alpha-1-antitrypsin specific activity in monoclonal gammopathies.

BACKGROUND: Serum alpha 1-antitrypsin (alpha 1AT) antigen concentration is elevated in malignancies as the result of acute phase reaction. In the present study, we examined whether the alpha 1AT elevation in monoclonal gammopathies was accompanied by an adequate increase of its functional activity. METHODS: In this case-control study, serum alpha 1AT concentration was measured in 187 ambulatory patients with monoclonal gammopathies and 320 healthy blood donors matched according to sex and age. The alpha 1AT antigen concentration was assayed by immunonephelometry, whereas its functional activity was measured as trypsin inhibitory capacity (TIC). The specific alpha 1AT inhibitory activity (SIA) was calculated, defined as the TIC/antigen concentration ratio. RESULTS: The alpha 1AT antigen concentrations obtained in the patients' samples were very significantly higher as compared with the corresponding values in the control group (mean +/- SD = 134 +/- 41.9% of normal, p < 0.001). However, the TIC values were higher in the patients than in the healthy controls only by 4% (104 +/- 23.8%, p < 0.05). The specific alpha 1AT activity was very significantly lower in the patients, as compared with the controls (p < 0.001), indicating that serum alpha 1AT in monoclonal gammopathies was partially inactive. CONCLUSIONS: As poor correlation between the TIC values and the antigen concentrations was obtained in the patient group as well as the decreased specific alpha 1AT activities, the TIC values in patients with monoclonal gammopathies should be interpreted with caution.

Plasma C1 inhibitor in malignant diseases: functional activity versus concentration.

Plasma C1 inhibitor (C1-INH) was determined in 64 patients with different malignant diseases and in 58 healthy persons. C1-INH antigen concentration was measured by radial immunodiffusion (RID), whereas its functional activity was assayed with chromogenic substrate. Within-run and day-to-day precision of both methods were good, with CVs ranging from 3.6 to 5.4%. Plasma C1-INH antigen concentrations were significantly higher in the patients than in healthy controls (P = 4.0 x 10(-3)), as were their C1-INH functional activities (P = 3.5 x 10(-3)). C1-INH activities obtained in the patient plasma samples were in correlation with their antigen concentrations (r = 0.914), showing that C1-INH synthesized in malignant disease was functional. However, the specific activity of the C1-INH (functional activity/antigen concentration ratio) was significantly lower in the patient group as compared with the controls (P = 2.1 x 10(-6)), indicating partial inactivation of plasma C1-INH in malignant diseases. The C1-INH specific activity in patients was inversely proportional to its antigen concentration.

Oxidative stress and anti-oxidative defense in schoolchildren residing in a petrochemical industry environment.

OBJECTIVE: To evaluate the possible relationship between industrial air pollution and oxidative stress in schoolchildren by comparing parameters from children residing in two nearby localities with contrasting environmental conditions. PARTICIPANTS: 42 schoolchildren (12-15 years) from Pancevo (site of Serbias largest petrochemical installation) formed the exposed group. 82 schoolchildren from Kovacica village, located 30 km north of Pancevo, formed the non-exposed group. METHODS: Oxidative stress status, anti-oxidative defense parameters, paraoxonase-1 status, lipid status, glucose concentration and leukocyte counts were compared in two groups. RESULTS: The children from Pancevo showed higher level of oxidative stress demonstrated by an elevated malondialdehyde concentration (P <0.001) and decreased superoxide dismutase activity (P<0.01) in comparison to the non-exposed group. CONCLUSIONS: The results suggested a relationship between the presence of air pollutants and increased oxidative stress in schoolchildren residing in an industrial environment.

Association of oxidative stress and PON1 with LDL and HDL particle size in middle-aged subjects.

BACKGROUND: Alterations in plasma lipoprotein subclass distributions affect atherosclerosis risk. Smaller, denser low-density lipoprotein (LDL) particles (sdLDL) are more susceptible to oxidation. In contrast, most of the protective effects of high-density lipoproteins (HDL) are attributable to larger particles. This study investigates the connection between LDL and HDL particle heterogeneity and oxidative stress, antioxidative defence (AOD) and paraoxonase (PON1) status in a healthy middle-aged Serbian population. MATERIALS AND METHODS: LDL and HDL particle sizes and subclass distributions were measured by gradient gel electrophoresis in 104 men and 103 women, aged 53 +/- 9.4 years. PON1 activities and PON1(Q192R) phenotypes were determined with paraoxon and diazoxon as substrates. The oxidative stress/AOD status was estimated by measuring malondialdehyde (MDA) and superoxide-anion (O2*(-)) levels and superoxide-dismutase (SOD) activity. RESULTS: Subjects with sdLDL had significantly higher MDA (P < 0.001) and O2*(-)(P < 0.05) levels and greater diazoxonase (DZOase) activity (P < 0.05) compared to subjects with larger LDL particles. A high MDA concentration was a significant predictor of the sdLDL phenotype (P < 0.005). Increased levels of and MDA were associated with smaller HDL(3) subclass abundance. Reduced HDL particle size was associated with lower DZOase activity (P < 0.01). CONCLUSIONS: Even in the absence of symptoms of atherosclerosis, sdLDL particles are associated with increased oxidative stress, which may stimulate a compensatory rise in PON1 DZOase activity. Elevated oxidative stress may significantly affect HDL subclass distribution, resulting in the accumulation of smaller, denser HDL particles with diminished antioxidative capacity.

Some kinetic characteristics of erythrocyte alanine aminotransferase phenotypes.

The activity of alanine aminotransferase (ALT) phenotypes was determined in 148 hemolysates of the Serbian population. The highest activity was obtained for phenotype ALT 1 (0.614 U/g Hb), intermediate for ALT 2-1 (0.475 U/g Hb), and the lowest for ALT 2 (0.395 U/g Hb). To explain the differences in catalytic activity between the ALT phenotypes, some kinetic characteristics were investigated. No difference in heat stability and calculated activation energies for ALT phenotypes could be detected. Addition of pyridoxal 5'-phosphate to the reaction system did not increase the catalytic activity. For the catalytic activity of all three phenotypes, a broad pH optimum in the range 7.1 to 7.6 was found. The Tris/HCl buffer concentration of 140 mmol/liter was optimal. The Michaelis-Menten constants for L-alanine as substrate were 2.462 mmol/liter for ALT 1, 1.965 mmol/liter for ALT 2-1, and 2.698 mmol/liter for ALT 2. For another substrate, 2-oxoglutarate, Km values were 0.299, 0.208, and 0.202 mmol/liter, respectively.

Eight red cell enzymes polymorphisms in the Slovakian ethnic group of Yugoslavia.

The polymorphisms of eight red cell enzymes were studied in 211 unrelated voluntary blood donors from the ethnic group of Slovakians (Yugoslavia). Only common phenotypes were detected, which are usually present in European populations. The allele frequencies found were: GLO1*1 = 0.410, ESD*1 = 0.887, AK1*1 = 0.962, PGM1*1 = 0.780, ACP1*A = 0.315, ACP1*B = 0.637, ACP1*C = 0.047, GPT*1 = 0.535, ADA*1 = 0.952 and PGD*A = 0.940. These findings are discussed in the context of other European populations and the population of Serbia, Yugoslavia.

Polymorphism of red cell glyoxalase I in Serbia, Yugoslavia.

Red cell glyoxalase I (GLO) phenotypes were determined in 258 unrelated adults from the population of Serbia (Yugoslavia). The GLO1 gene frequency was estimated to be 0.384.

A study of several red cell enzyme markers in the Rumanian ethnic group in Yugoslavia.

The present investigation reports the polymorphism of eight red cell enzymes, studied in 308 unrelated voluntary blood donors from the Rumanian ethnic group of Yugoslavia. Only common phenotypes were detected, which are distributed as in European populations. The estimated gene frequencies were: GLO1*1 = 0.401, GPT*1 = 0.533, PGM1*1 = 0.707, ESD*1 = 0.878, AK1*1 = 0.982, PGD*A = 0.974, ADA*1 = 0.939, ACP1*A = 0.384, ACP1*B = 0.550 and ACP1*C = 0.065. The observed gene frequencies are discussed in the context of other European populations and other populations from Yugoslavia.

Red cell enzyme polymorphisms of the Hungarian ethnic group in Yugoslavia.

A genetic study was carried out on phenotype and gene frequencies of the genetic markers in eight red cell enzymes: glyoxalase I (GLO1), glutamic pyruvate transaminase (GPT), phosphoglucomutase (PGM1), esterase D (ESD), adenylate kinase (AK1), 6-phosphogluconate dehydrogenase (6-PGD), adenosine deaminase (ADA), acid phosphatase (ACP1), in the Hungarian ethnic group living in Yugoslavia. The gene frequencies obtained were: GPT*1 = 0.542, PGM1*1 = 0.760, ESD*1 = 0.909, AK*1 = 0.971, PGD*A = 0.971, ADA*1 = 0.939, GLO1*1 = 0.417, ACP1*A = 0.329, ACP1*B = 0.591 and ACP1*C = 0.080. The distribution of these phenotype and gene frequencies was examined and compared with the phenotype and gene frequencies found for the Hungarian population living in Hungary and for other populations living in the northeast of Yugoslavia.

[Use of cyclosporin A for remission induction in newly-detected insulin-dependent diabetes]. / Primena ciklosporina A u indukciji remisije u novootkrivenom insulini-zavisnom dijabetesu.

It has been postulated that some of the recent-onset insulin-dependent diabetics, after the initial use of insulin therapy, might develop the "honey moon period", i.e., a spontaneous remission of the disease, defined as the state of normal metabolic control maintained without insulin therapy. However, it has also been shown that spontaneous remission appears only in 5% of the patients treated with conventional insulin therapy and lasts, most frequently, not more than a few weeks. Different therapeutic regimens of immunosuppression and immunomodulation have been used worldwide in order to induce the remission, based on the findings that an autoimmune process underlies the pathogenesis of this type of diabetes. In this study, we have shown the results of the follow-up analysis of the effects of the treatment with cyclosporin A in 21 recent-onset insulin-dependent diabetics. In 15 of those patients insulin treatment was applied as bi-daily doses of monocomponent insulin preparations, and in 6 of them intensified insulin therapy with human insulin was used. In the first group, the remission was achieved in 46.66% and in the second group in 66.66%, which is a significantly higher incidence than in control groups treated only with insulin, without cyclosporin. Moreover, the duration of remission was longer in the patients treated with cyclosporin. The analysis of the residual beta cell secretory capacity has shown that C-peptide levels (taken as a marker for insulin secretion) were slightly higher in patients with the spontaneous remission than in those with the cyclosporin-induced remission both in basal conditions and after stimulation with 1 mg of glucagon. In the patients with cyclosporin A-induced remission we found an improved basal C-peptide secretion and, even more, we detected a significant improvement in beta cell response to the glucagon stimulation. The analysis of the first-phase insulin secretory response (the insulin response to rapidly injected glucose during the intravenous glucose tolerance test) which has been shown to be impaired very early during the development of diabetes, has demonstrated the lack of its recovery both in the spontaneous and in cyclosporin A-induced remissions. The analysis of the molar insulin/C-peptide ratio has detected the impairments of this ratio which remains decreased both in spontaneous and cyclosporin-induced remissions.

Polymorphism of human red cell galactose-1-phosphate uridyl transferase in Serbia, Yugoslavia.

Phenotypes of human red cell galactose-1-phosphate uridyl transferase (GALT) were determined in 283 unrelated adults from Serbia (Yugoslavia). The gene frequencies were 0.959 for GALT N, 0.018 for GALT D and 0.023 for GALT N.

Esterase D polymorphism in Serbia (Yugoslavia).

Phenotypes of human red cell esterase D (EsD) were determined in 351 unrelated adults from Serbia (Yugoslavia). The calculated allele frequencies were 0.911 for EsD1 and 0.089 for EsD2. The phenotype distribution was in good agreement with the Hardy-Weinberg equilibrium.