GPX3 rs8177412 Polymorphism Modifies Risk of Upper Urothelial Tumors in Patients with Balkan Endemic Nephropathy.

Current data suggest that aristolochic acid (AA) exposure is a putative cause of Balkan endemic nephropathy (BEN), a chronic kidney disease strongly associated with upper tract urothelial carcinoma. The cellular metabolism of AA is associated with the production of reactive oxygen species, resulting in oxidative distress. Purpose: Therefore, the aim of this study was to analyze individual, combined and cumulative effect of antioxidant gene polymorphisms (Nrf2 rs6721961, KEAP1 rs1048290, GSTP1AB rs1695, GSTP1CD rs1138272, GPX3 rs8177412 and MDR1 rs1045642), as well as GSTP1ABCD haplotypes with the risk for BEN development and associated urothelial cell carcinoma in 209 BEN patients and 140 controls from endemic areas. Experimental method: Genotyping was performed using polymerase chain reaction (PCR) and PCR with confronting two-pair primers (PCR-CTTP) methods. Results: We found that female patients carrying both variant GPX3 rs8177412 and MDR1 rs1045642 genotypes in combination exhibited significant risk towards BEN (OR 1 = 3.34, 95% CI = 1.16-9.60, p = 0.025; OR 2 = 3.79, 95% CI = 1.27-11.24, p = 0.016). Moreover, significant association was determined between GPX3rs8174412 polymorphism and risk for urothelial carcinoma. Carriers of variant GPX3*TC + CC genotype were at eight-fold increased risk of BEN-associated urothelial tumors development. There was no individual or combined impact on BEN development and BEN-associated tumors among all examined polymorphisms. The haplotype consisting of variant alleles for both polymorphisms G and T was associated with 1.6-fold increased risk although statistically insignificant (OR = 1.64; 95% CI = 0.75-3.58; p = 0.21). Conclusions: Regarding GPX3 rs8177412 polymorphism, the gene variant that confers lower expression is associated with significant increase in upper urothelial carcinoma risk. Therefore, BEN patients carrying variant GPX3 genotype should be more frequently monitored for possible upper tract urothelial carcinoma development.

Thyroid stimulating hormone values of clinical decisions of hypothyroidism measurement by three different automated immunoassays.

Background: An accurate measurement of serum thyroid stimulating hormone (TSH) is crucial for a thyroid disorder diagnosis and management. We compared the values of TSH between three automated immunoassays with aims to provide insights into variations in TSH levels that are important for a clinical decision: 2.50 mIU/L, 4.00 mIU/L and 10.00 mIU/L.Methods: We measured TSH with three different fully automated immunoassays: Abbott (Architect ci8200), Siemens (ADVIA Centaur XP) and Roche (Cobas e411). Serum was collected from 110 patients between August 2018 and January 2019. The results were compared using the Passing-Bablok regression method. Additionally, linear regression coefficients were calculated after logarithmic transformation of data.Results: Although all three regression coefficients were high (r2 > 0.98), the slopes from Passing-Bablok plots for the correlation of Abbott with either Roche or Siemens were merely 0.66 and 0.73, respectively. The slope for the correlation of Roche and Siemens was 1.11. Consecutively, the results obtained by the Roche and Siemens methods were proportionally higher than those obtained by the Abbott method (38% and 52%, respectively) at all measured ranges.Conclusions: Although immunoassays correlated among themselves, they cannot achieve the same values for clinical decisions for hypothyroidism (clinical requirements). Clinicians should be conscious of these limitations. A harmonisation of the methods is needed to meet clinical requirements and to enable appropriate clinical decisions in cases of hypothyroidism.Likewise, we suggest the introduction of borderline and high risk values of TSH for hypothyroidism depending on immunoassays to avoid misdiagnosis.

Atorvastatin treatment increases plasma bilirubin but not HMOX1 expression in stable angina patients.

BACKGROUND: In vitro and animal studies indicate that statins increase heme oxygenase-1 gene (HMOX1) expression, which then, presumably, increases plasma bilirubin concentration. However, clinical confirmation that statins concomitantly increase HMOX1 expression and plasma bilirubin concentration is lacking. We hypothesized that in patients with stable angina atorvastatin therapy (20 mg/day for 10 weeks) concomitantly increases total bilirubin concentration and HMOX1 expression, as assessed non-invasively by plasma analysis. METHODS: In 44 patients with stable angina plasma concentrations of total bilirubin, HMOX1 mRNA and HMOX1 protein were measured before and after the statin treatment, as well as plasma concentrations of oxidized low-density lipoprotein (OxLDL), malondialdehyde (MDA), monocyte chemoattractant protein (MCP-1) and C-reactive protein (CRP). RESULTS: Atorvastatin treatment increased total bilirubin concentration (median 6.95 µmol/L vs. 8.55, + 23.02%; p < 0.001), but did not change plasma HMOX1 mRNA and HMOX1 protein concentrations. Plasma concentrations of OxLDL (- 31.85%, p < 0.001), MCP-1 (- 16.20%, p = 0.020) and CRP (- 7.32%, p = 0.010) were decreased but MDA was not decreased (15.32%, p = 0.107). Within subjects, increment of plasma bilirubin concentration did not correlate with the changes in HMOX1 mRNA and protein concentrations, but correlated with low-density lipoprotein cholesterol decrement (r = - 0.374, p = 0.012). Bilirubin increment did not correlate with the changes in oxidative and inflammatory markers. CONCLUSION: Our prospective observational trial finally confirms that atorvastatin (20 mg/day for 10 weeks) increases plasma total bilirubin concentration. However, it seems that this effect is HMOX1-independent.

The predictive role of biochemical markers on outcomes of severe COVID-19 patients admitted to intensive care unit.

Background: The pandemic of severe acute respiratory syndrome by coronavirus 2 (SARS-CoV-2) is a multi-system disease caused by a diffuse systemic process involving a complex interaction of the inflammatory, immunological and coagulative cascades. This study aims to identify the most effective biomarkers to predict poor outcome in intensive care unit (ICU) patients with severe COVID-19 disease. Methods: A single-centre retrospective observational study enrolled 69 deceased and 20 recovered patients treated in the ICU of the General Hospital Gradiska in the period from March 1, 2021. until April 1, 2022. We evaluated the leukocytes (WBC), lymphocytes (LYM), neutrophils (NEU), platelets (PLT), haemoglobin, neutrophil-lymphocyte ratio (NLR), platelet lymphocyte ratio (PLR), and systemic immune-inflammation index (SII). In addition, we evaluated the IL-6, ferritin, CRP, D-dimer, magnesium, bilirubin and lactate dehydrogenase.

Analysis of redox status and HDL subclasses in patients with lymphoma and the associations with FDG-PET/CT findings.

Newer research points to alterations in the plasma redox status and the HDL subclass distributions in cancer. We aimed to assess the redox status and the HDL subclass distributions, lipids, and inflammatory markers in lymphoma patients in order to determine whether they were correlated with changes in FDG-PET/CT scans. At the beginning of this study, redox status, HDL subclasses, lipids, and inflammation biomarkers were determined in 58 patients with lymphoma (Hodgkin lymphoma, n=11 and non-Hodgkin lymphoma, n=47), and these same measurements were reassessed during their ensuing treatment (in 25 patients). Initially, the total oxidation status (TOS), the prooxidant-antioxidant balance (PAB), the OS index (OSI), the total protein sulfhydryl groups (SH-groups), and the advanced oxidation protein products (AOPP) were significantly higher in lymphoma patients as compared to healthy subjects, but the total antioxidant status (TAS) was significantly reduced. The PAB had a strong correlation with the CRP and interleukin-6 (rho=0.726, p<0.001; rho=0.386, p=0.003). The correlations between these parameters and the maximum standardized uptake values (SUVmax) were: PAB, rho=0.335 and p=0.010; SH-groups, rho=0.265 and p=0.044; CRP, rho=0.391 and p=0.002; HDL3b, rho=0.283 and p=0.031; HDL2b, rho= -0.294 and p=0.025; and HDL size, rho= -0.295 and p=0.024. The reductions in SUVmax between two follow-up points were associated with increases in the OSI, TOS, and SH-groups, as well as a reduction in the PAB and TAS. In conclusion, the redox parameters in patients with lymphoma were consistent with FDG-PET/CT findings. Targeting the redox status parameters and the HDL subclasses could be potential strategies in the molecular fight against lymphoma.

Indirect estimation of reference intervals for thyroid parameters using advia centaur XP analyzer.

Background: The aim of this study was to determine the reference intervals (RIs) for thyroid stimulating hormone (TSH), free thyroxine (FT4), free triiodothyronine (FT3) and FT3/FT4 ratio using indirect methods. Methods: We analyzed 1256 results TSH, FT4 and FT3 collected from a laboratory information system between 2017 and 2021. All measurements were performed on a Siemens ADVIA Centaur XP analyzer using the chemiluminescent immunoassay. We calculated the values of the 2.5th and 97.5th percentiles as recommended by the IFCC (CLSI C28-A3). Results: The RIs derived for TSH, FT4, FT3 and FT3/FT4 ratio were 0.34-4.10 mIU/L, 11.3-20.6 pmol/L, 3.5-6.32 pmol/L and 0.21-0.47, respectively. We found a significant difference between calculated RIs for the TSH and FT4 and those recommended by the manufacturer. Also, FT3 values were significantly higher in the group younger than 30 years relative to the fourth decade (5.26 vs. 5.02, p=0.005), the fifth decade (5.26 vs. 4.94, p=0.001), the sixth decade (5.26 vs. 4.87, p<0.001), the seventh decade (5.26 vs. 4.79, p<0.001) and the group older than 70 years old (5.26 vs. 4.55, p<0.001). Likewise, we found for TSH values and FT3/FT4 ratio a significant difference (p <0.001) between different age groups. Conclusions: The establishing RIs for the population of the Republic of Srpska were significantly differed from the recommended RIs by the manufacturer for TSH and FT4. Our results encourage other laboratories to develop their own RIs for thyroid parameters by applying CLSI recommendations.

Plasma Cathepsin S is Associated with High-density Lipoprotein Cholesterol and Bilirubin in Patients with Abdominal Aortic Aneurysms.

BACKGROUND: Cathepsin S (CTSS) is a cysteine protease involved in atherogenesis. We compared the plasma CTSS as well as other biomarkers of atherosclerosis in patients with abdominal aortic aneurysms (AAA) and aortoiliac occlusive disease (AOD), aiming to identify the underlying pathogenic mechanisms of the disease development. Also, we hypothesised that the level of plasma CTSS simultaneously increases with a decrease of plasma high-density lipoprotein cholesterol (HDL-C) values. METHODS: 33 patients with AAA and 34 patients with AOD were included in this study. RESULTS: There was no difference in the level of plasma CTSS between the two analysed groups (p=0.833). In the patients with AAA, the plasma CTSS was correlated with HDL-C (r = -0.377, p = 0.034) and total bilirubin (r =0.500, p = 0.003) while, unexpectedly, it was not correlated with cystatin C (Cys C) (r =0.083, p = 0.652). In the patients with AOD, the plasma CTSS correlated with triglycerides (r = 0.597, p< 0.001), only. When the patients were divided according to HDL-C (with HDL-C ≤0.90 and HDL-C >0.90 mmol/L), the plasma CTSS values differed among these groups (31.27 vs.25.61 µg/L, respectively, p<0.001). CONCLUSIONS: These results provide the first evidence that CTSS negatively correlated with HDL-C and bilirubin in patients with AAA. It is possible that differences in the association of the CTSS and other markers of atherosclerosis can determine whether atherosclerotic aorta will develop dilatation or stenosis.

Direct Estimation of Reference Intervals for Thyroid Parameters in the Republic of Srpska.

BACKGROUND: The aim of this study was to determine the reference values for thyrotropin (TSH), thyroid hormones (total and free thyroxine, T4 and fT4; total and free triiodothyronine, T3 and fT3), thyroglobulin (Tg) and thyroid antibodies (thyroid peroxidase, TPOAb and thyroglobulin antibody, TgAb) in the population of the Republic of Srpska. METHODS: A total of 250 euthyroid subjects were enrolled in this study. A direct method for choosing reference subjects was used to establish reference intervals. The hormones and thyroid antibodies were measured by an electrochemiluminescence immunoassay method (ECLIA, Roche Diagnostics, Mannheim, Germany). We calculated the reference intervals by MedCalc, version 12.1.4.0 (MedCalc software, Belgium) as recommended by the IFCC (CLSI C28-A3). RESULTS: Using guidelines recommended by the National Academy of Clinical Biochemistry (NACB) and based on standard statistical approaches, the reference intervals derived for TSH, fT4, T4, fT3, T3 were 0.75-5.32 mIU/L, 12.29-20.03 pmol/L, 73.49-126,30 nmol/L, 4.11-6.32 pmol/L, 1.15-2.32 nmol/L and for Tg, TPOAb, TgAb were 3.63-26.00 µg/L, <18.02 mIU/L, < 98.00 mIU/L, respectively. We found a significant difference (p<0.05) in TSH and fT3 values between different age groups as well as in T4, fT4 and fT3 values between ge nder groups. CONCLUSIONS: The established reference values for the population of the Republic of Srpska were significantly different from the values recommended by the manufacturer of reagents (Roche Diagnostics). Our results showed that a laboratory needs to establish its own reference values in order to set up a proper diagnosis, as well as to treat patients successfully.

The impact of time of sample collection on the measurement of thyroid stimulating hormone values in the serum.

OBJECTIVE: The aim of our research is to determine whether the time of blood sampling and fasting of patients have an impact on TSH values. DESIGN AND METHODS: A total of 198 participants were enrolled in this study and classified into five groups: A--the first sample collection for TSH measurement was taken between 7:00 and 8:00 a.m. at fasting and the second after 140 min without food intake; B--between 7:00 and 8:00 a.m. at fasting and the second after 140 min with food intake; C--between 7:00 and 8:00 a.m. at fasting the previous day and the second one between 7:00 and 8:00 a.m. at fasting the following day; D--between 9:00 and 10:00 a.m. at fasting the previous day and the second one between 9:00 and 10:00 a.m. at fasting the following day, and E--between 9:00 and 10:00 a.m. at fasting the previous day and the second one between 7:00 and 8:00 a.m. on the following day. Serum TSH concentration was measured by electrochemiluminescence immunoassay (ECLIA, Roche Diagnostics, Mannheim, Germany). RESULTS: TSH values (mIU/L) were in group A: 2.50 (2.20-2.81) first samples, 1.74 (1.52-1.96) second samples, p<0.001; B: 2.11 (1.52-2.72) first samples, 1.56 (1.13-1.81) second samples, p<0.001; C: 2.60 (2.28-2.91) first samples, 2.23 (1.92-2.53) second samples, p<0.001; D: 1.80 (1.48-2.11) first samples, 1.77 (1.44-2.09) second samples, p<0.597; and E: 1.32 (1.11-2.16) first samples, 1.67 (1.48-2.93) second samples, p<0.001. CONCLUSION: The time of sample collection must be standardised for the purpose of standardisation and harmonisation of TSH measurements.

The Pleiotropic Effects of Atorvastatin on Stable Angina Patients: Evidence by Analysis of High-Density Lipoprotein Size and Subclasses, and Plasma mRNA.

BACKGROUND: High-density lipoproteins (HDL) have athero-protective biological properties: antioxidative, anti-apoptotic, anti-inflammatory, and they have the efflux capacity of cellular cholesterol. Plasma mRNA analysis can be used to investigate statin pleiotropy in vivo as a new analytical tool for non-invasive assessment of gene expression in vascular beds. The aim of this study was to assess the pleiotropic effects of atorvastatin in stable angina patients with high-risk values (group A) as compared with patients who had borderline and desirable HDL-cholesterol (HDL-C) values (group B). METHODS: The atorvastatin therapy (20 mg/day) was given to forty-three patients with stable angina for 10 weeks. We investigated three statin pleiotropy-targeted genes: inter-cellular adhesion molecule-1, chemokine (C-C motif) ligand 2 and cathepsin S and assessed by gel electrophoresis gradient the effects of atorvastatin on HDL size and subclasses. RESULTS: In group A, after therapy, HDL-C concentration was significantly increased but not in group B. Atorvastatin lowered plasma chemokine (C-C motif) ligand 2 and intercellular adhesion molecule-1 mRNA levels in both groups, but did not change the plasma cathepsin S mRNA levels. In group A only, baseline total bilirubin showed negative correlations with the genes of cathepsin S (r=-0.506; p=0.023) and significantly increased after therapy. CONCLUSIONS: HDL-C and bilirubin can be promising therapeutic targets in the treatment of cardiovascular diseases. Analysis of cell-free mRNA in plasma might become a useful tool for estimating statin pleiotropy.

Interrelated cathepsin S-lowering and LDL subclass profile improvements induced by atorvastatin in the plasma of stable angina patients.

AIM: We hypothesized that, in stable angina patients, atorvastatin therapy lowers the cathepsin S (CTSS) concentrations, as assessed non-invasively according to a plasma analysis. In addition, the low-density lipoprotein (LDL) and high-density lipoprotein (HDL) size and subclasses in the plasma were analysed to establish the association between CTSS and lipoprotein metabolism and determine whether this association is atorvastatin-sensitive. METHODS: A total of 43 patients with stable angina received atorvastatin therapy (20 mg/day, 10 weeks). The plasma CTSS mRNA levels, CTSS protein concentrations and CTSS activity, as well as LDL and HDL size and subclasses, were analysed before and after treatment. RESULTS: Atorvastatin treatment did not change the plasma CTSS mRNA levels, although it lowered the plasma CTSS concentrations and activity. An increased plasma CTSS concentration and activity were found to be associated with a more atherogenic LDL subclass profile (a decreased dominant LDL size and increased percentage of small, dense LDL particles). The atorvastatin-induced CTSS-loweringeffect was concomitant with an improvement in the LDL subclass profile, and the changes were found to be interrelated. Concomitant, interrelated changes in the CTSS levels and LDL subclass profiles were found in the LDL phenotype B patients only (a dominant LDL diameter of ≤ 25.5 nm at the start of the study). In this subgroup, lowering of the plasma CTSS mRNA level also correlated with lowering of the proportion of small, dense LDL particles. CONCLUSIONS: Atorvastatin-induced CTSS-lowering and LDL subclass profile improvements in the plasma of LDL phenotype B patients with stable angina are concomitant and interrelated.

Atorvastatin in stable angina patients lowers CCL2 and ICAM1 expression: pleiotropic evidence from plasma mRNA analyses.

OBJECTIVE: Statin pleiotropy is still an evolving concept, and the lack of clarity on this subject is due at least in part to the lack of a definitive biomarker for statin pleiotropy. Using plasma mRNA analysis as a novel research tool for the non-invasive in vivo assessment of gene expression in vascular beds, we hypothesised that atorvastatin lowers the plasma mRNA level from statin pleiotropy-target genes, and the reduction is independent of the reduction of low-density lipoprotein cholesterol (LDL-C). DESIGN AND METHODS: Forty-four patients with stable angina received atorvastatin therapy (20 mg/day, 10 weeks). Plasma chemokine (C-C motif) ligand 2 (CCL2) and intercellular adhesion molecule-1 (ICAM1) mRNA levels and their protein concentrations (MCP-1, sICAM-1) were analysed before and after the treatment. Plasma vascular adhesion molecule-1 (sVCAM-1) concentrations were also analysed. RESULTS: Atorvastatin lowered plasma mRNA levels (CCL2: -31.76%, p=0.037; ICAM1: -34.09%, p<0.001) and MCP-1 protein concentration (-18.88%, p=0.008) but did not lower sICAM-1 and sVCAM-1 protein concentrations, and the decreases appeared to be independent from the lowering of LDL-C. The plasma mRNA levels correlated with their protein concentrations following statin treatment only. CONCLUSION: Our results significantly strengthen the clinical evidence in support of statin pleiotropy. Furthermore, this unique simultaneous measurement of plasma mRNAs and their protein concentrations offers an advanced non-invasive in vivo assessment of the circulation pathology.

[Correlation between C-reactive protein levels with leading risk factors for cardiovascular disease in men].

INTRODUCTION: The aim of this study was to estimate the correlation between C-reactive protein levels and leading risk factors for cardiovascular disease in men. MATERIAL AND METHODS: The study included 183 working capable men chosen randomly from the regular systematical check-up in Health Centre Banja Luka in 2006. Standard laboratory methods were used to establish the following: total cholesterol, triglyceride and HDL-cholestreol level and LDL-cholesterol level was calculated. High sensitive C-reactive protein level was measured by immunuturbidimetric method CRP (Latex) HS Roche Diagnostic. RESULTS: Average values of high sensitive C-reactive protein for the whole group was 1.69 mg/L, total cholesterol 5.73 mmol/L, HDL-cholesterol 1.38 mmol/L, LDL-cholesterol 3.40 mmol/L. The average value for the systolic blood pressure was 132.9 mmHg, diastolic blood pressure 85.4 mmHg, and body mass index 28.47 kg/m2. Out of the overall number of examinees, 74 were smokers (40.4%) and 109 (59.6%) nonsmokers. The statistical analysis showed that there was a statistically significant difference between C-reactive protein level in the group with diastolic blood pressure below 90 mmHg and above (p < 0.05); as well as statistically significant difference between the group with desirable body mass index and the group with increased BMI (p < 0.05). DISCUSSION: The results of our study show that there is a significant correlation between CRP levels and high blood pressure, and in persons with increased body mass index. However, there was no correlation between CRP levels and total cholesterol HDL and LDL cholesterol levels. CONCLUSION: High sensitive CRP screening is useful in early detection and prevention of cardiovascular diseases

[The correlation between lifestyle and lipid profile].

INTRODUCTION: The aim of this study was to estimate if negative lifestyle habits such as alcohol consumption, smoking and physical inactivity affect the lipid profile values. MATERIAL AND METHODS: The study included 250 workers on regular examination in the Gradiska Health Center in the period from 2001 to 2002. There were 113 (45.2%) men and 137 (54.8%) women. The examinees were divided into three groups according age (25-39, 40-49 and 50-60 respectively). Standard laboratory methods were used to establish the following: total cholesterol, triglyceride and HDL-cholesterol level, LDL cholesterol, atherosclerosis index (AI) and total cholesterol/HDL cholesterol. RESULTS: Using a questionnaire, we have found out that out of 250 examinees 48.80% consume alcohol regularly, 50.80% are smokers and 36% are physically. The mean total cholesterol was high in all groups and it was 6.41 mmol/l. The mean triglyceride level was 1.88 mmol/l and mean HDL cholesterol was 1.48 mmol/l, IA was 2.99 and total cholesterol/HDL cholesterol ratio was 4.69. Statistical analysis showed that there was a statistically significant relationship between triglyceride values and alcohol consumption, smoking and physical activity (p<0.05). Also, we showed that there was a statistically high relationship between HDL cholesterol values, AJ, total cholesterol/HDL cholesterol and smoking in the examined groups (p<0.01). DISCUSSION: In our study the lipid profile parameters were above the desired levels, probably due to unhealthy lifestyle, including smoking, alcohol consumption and insufficient physical activity. Our results are in concordance with the results of similar studies. CONCLUSION: It is of utmost importance to take steps to improve lifestyle habits of our population.

[Analysis of lipid status parameters in relation to gender and age in the working population of district of Gradiska]. / Analiza vrednosti parametara lipidskog statusa radno sposobne populacije opstine Gradiska u zavisnosti od pola i starosti.

INTRODUCTION: The aim of this investigation was to establish the frequency of hyperlipoproteinemias, as well as the distribution of desired, elevated and high risk values of certain lipid status parameters among working population of the Gradiska municipality with age and sex distribution. MATERIAL AND METHODS: This investigation included 250 workers, 109 male and 141 female, 25-60 years of age of the Gradiska municipality on a regular checkup. Standard biochemical methods were used to determine values of total serum cholesterol, triglycerides and HDL cholesterol. LDL cholesterol and LDL to HDL cholesterol ratio were calculated. RESULTS: Total serum cholesterol was elevated in 44.04% of males and 44.68% of females, triglycerides in 20.02% of males and 19.15% of females and LDL cholesterol in 31.96% of males and 21.43% of females. High risk values of total cholesterol were established in 43.12% of males and 39.01% of females, and of triglycerides in 37.61% of males and 9.93% of females. HDL cholesterol was decreased in 55.96% of males and 41.84% of females, while highly decreased values were established only in 5.5% of males and 1.42% of females. Increase of total cholesterol and LDL cholesterol correlated with workers' age, but values of triglycerides did not. Hyperlipoproteinemia was evident in 76.4% of examinees. DISCUSSION: All tested parameters vary dramatically from desired levels. These results are probably associated with unhealthy food habits and lifestyle. CONCLUSION: Our results point to the need to perform regular laboratory diagnostic procedures and routine check-ups in the working population.