Corticosteroids regulate a number of physiological processes and are synthetic analogs of the natural steroid hormones produced by the adrenal cortex. As drugs, corticosteroids are non-inflammatory and are used for the treatment of plethora of conditions which include arthritis, kidney, skin, lungs or thyroid disorders, for the treatment and relief of symptoms of allergies and symptoms of some gastrointestinal disorders. In addition, glucocorticoids can regulate the effects of inflammatory disorders, including sepsis, autoimmune diseases, and allergies. These conditions are potentially fatal. Consequently, this drug class is among the most commonly prescribed globally. One representative of corticosteroid class of drugs is dexamethasone which is used to treat allergies, adrenal problems, arthritis, asthma, diseases of blood or bone marrow, inflammation, kidney diseases, different types of skin conditions, and episodes of multiple sclerosis. Virulence factors help bacteria colonize the host at the level of the cell. In their nature, these factors are secretory, associated with the membrane or present in the cytosol. Secretory factors allow bacterium to circumvent the host immune response, while membrane factors aid bacterium in adhesion to the host cell. Finally, cytosol factors help bacteria adapt metabolically, physiologically, and morphologically to their changing environment. One such factor is aspartyl proteinase, a protein that degrades other proteins and is a virulence factor in many pathogens playing a role in the host invasion process. Another important virulence factor is the ability to form biofilms, which can render bacteria resistant to antimicrobials. Despite the widespread use of corticosteroids, including dexamethasone, little is known about their possible influence on the expression of virulence factors such as aspartyl proteinase. If such a connection is to exist the use of corticosteroids could elicit pathogenesis in certain microbes. In the here-presented study we wanted to investigate the effects of dexamethasone on the growth, expression of aspartyl proteinase and biofilm formation in three E. coli strains that were previously isolated from patients suffering from urinary tract infection. To this aim, we amended the growth media with 0.5 mg/mL dexamethasone. Bacterial growth was measured over the period of 24 hours and the effect of dexamethasone was established at different time points. Administration of 0.5 mg/mL glucocorticoid drug dexamethasone did not significantly affect bacterial growth. However, it resulted in an increase in concentration of secreted E. coli virulence factor aspartyl proteinase, which increased up to 2.6-fold for some E. coli strains. In addition, we noted the increased biofilm formation in to three out of four studied strains. This study indicates dexamethasone as a possible trigger molecule for the expression of virulence factor aspartyl proteinase in E. coli.

Limited knowledge exists about the effects of commonly used diuretic medications on the human normal flora. Thus, we investigated potential stimulatory effects of diuretic drug furosemide on urogenital tract microbiota in women. Three strains of E. coli and C. albicans with different biofilm forming capacities were obtained from female patients diagnosed with urinary tract infections. All tested strains were treated with two different concentrations of furosemide drug, in comparison to non-treated strains as the negative control. At specific time intervals, samples were obtained from growing culture and analyzed for their proliferation rate, aspartyl proteinase excretion and biofilm formation ability. E. coli and C. albicans strains significantly increased their aspartyl proteinase excretion under furosemide treatment. This effect was frequently observed after 16 hours of incubation at 37oC. This drug has also increased the biofilm forming capacities of E. coli and C. albicans strains. Interestingly, both E. coli and C. albicans non-biofilm former strains, gained the capacity of biofilm formation when treated with furosemide at certain concentrations. E. coli control became a weak biofilm former after 48 hours of incubation, while non-biofilm former C. albicans strain became a weak biofilm former in dose-dependent fashion, after 48 hours incubation with furosemide in concentration of 0.1 mg/mL, and after 16 hours of incubation with furosemide in concentration of 0.5 mg/mL. Loop diuretic drug furosemide is able to increase the microbial virulence and turn commensal microbes into opportunistic pathogens. Additionally, the results suggest that enzyme aspartyl proteinase might act as a signal molecule for the biofilm formation, leading to the increased microbial pathogenicity.

Glioblastoma multiforme (GBM) is the most frequent type of primary astrocytomas. We examined the association between single nucleotide polymorphisms (SNPs) in Aurora kinase A (AURKA), Aurora kinase B (AURKB), Aurora kinase C (AURKC) and Polo-like kinase 1 (PLK1) mitotic checkpoint genes and GBM risk by qPCR genotyping. In silico analysis was performed to evaluate effects of polymorphic biological sequences on protein binding motifs. Chi-square and Fisher statistics revealed a significant difference in genotypes frequencies between GBM patients and controls for AURKB rs2289590 variant (p = 0.038). Association with decreased GBM risk was demonstrated for AURKB rs2289590 AC genotype (OR = 0.54; 95% CI = 0.33–0.88; p = 0.015). Furthermore, AURKC rs11084490 CG genotype was associated with lower GBM risk (OR = 0.57; 95% CI = 0.34–0.95; p = 0.031). Bioinformatic analysis of rs2289590 polymorphic region identified additional binding site for the Yin-Yang 1 (YY1) transcription factor in the presence of C allele. Our results indicated that rs2289590 in AURKB and rs11084490 in AURKC were associated with a reduced GBM risk. The present study was performed on a less numerous but ethnically homogeneous population. Hence, future investigations in larger and multiethnic groups are needed to strengthen these results.

The identification of mutually exclusive somatic mutations shared among myeloproliferative neoplasm (MPN) subtypes has provided a powerful tool for studying disease evolution. Clinical features, gene mutations, and survival over 18 years were analyzed in MPN patients. One hundred thirty-eight MPN patients were subcategorized according to MPN subtypes: essential thrombocythemia (ET, n = 41), polycythemia vera (PV, n = 56), primary myelofibrosis (PMF, n = 10), and MPN unclassified (MPN-U, n = 31). Patient characteristics included clinical parameters, overall survival (OS), and mutational status of the Janus kinase 2 (JAK2), calreticulin (CALR), and myeloproliferative leukemia virus oncogene (MPL) genes. We compared hematologic and clinical features of JAK2V617F-ET vs. CALR-mutated ET vs. JAK2V617F-PV patients. JAK2V617F-patients had higher values of erythrocytes, hemoglobin, and hematocrit compared to CALR-mutated patients (p < 0.05). The mutant allele burden in JAK2V617F-PV and JAK2V617F-ET patients directly correlated with erythrocyte, hemoglobin, and hematocrit values, but it inversely correlated with platelet count. Thus, mutant allele burden was an indicator of the clinical phenotype in JAK2V617F-MPN patients. OS was not affected by the mutational status. In general, mutated JAK2, CALR, and MPL genes left specific hematological signatures.

Single nucleotide polymorphisms (SNPs) in genes encoding mitotic kinases could influence development and progression of gastric cancer (GC). Case-control study of nine SNPs in mitotic genes was conducted using qPCR. The study included 116 GC patients and 203 controls. In silico analysis was performed to evaluate the effects of polymorphisms on transcription factors binding sites. The AURKA rs1047972 genotypes (CT vs. CC: OR, 1.96; 95% CI, 1.05–3.65; p = 0.033; CC + TT vs. CT: OR, 1.94; 95% CI, 1.04–3.60; p = 0.036) and rs911160 (CC vs. GG: OR, 5.56; 95% CI, 1.24–24.81; p = 0.025; GG + CG vs. CC: OR, 5.26; 95% CI, 1.19–23.22; p = 0.028), were associated with increased GC risk, whereas certain rs8173 genotypes (CG vs. CC: OR, 0.60; 95% CI, 0.36–0.99; p = 0.049; GG vs. CC: OR, 0.38; 95% CI, 0.18–0.79; p = 0.010; CC + CG vs. GG: OR, 0.49; 95% CI, 0.25–0.98; p = 0.043) were protective. Association with increased GC risk was demonstrated for AURKB rs2241909 (GG + AG vs. AA: OR, 1.61; 95% CI, 1.01–2.56; p = 0.041) and rs2289590 (AC vs. AA: OR, 2.41; 95% CI, 1.47–3.98; p = 0.001; CC vs. AA: OR, 6.77; 95% CI, 2.24–20.47; p = 0.001; AA+AC vs. CC: OR, 4.23; 95% CI, 1.44–12.40; p = 0.009). Furthermore, AURKC rs11084490 (GG + CG vs. CC: OR, 1.71; 95% CI, 1.04–2.81; p = 0.033) was associated with increased GC risk. A combined analysis of five SNPs, associated with an increased GC risk, detected polymorphism profiles where all the combinations contribute to the higher GC risk, with an OR increased 1.51-fold for the rs1047972(CT)/rs11084490(CG + GG) to 2.29-fold for the rs1047972(CT)/rs911160(CC) combinations. In silico analysis for rs911160 and rs2289590 demonstrated that different transcription factors preferentially bind to polymorphic sites, indicating that AURKA and AURKB could be regulated differently depending on the presence of particular allele. Our results revealed that AURKA (rs1047972 and rs911160), AURKB (rs2241909 and rs2289590) and AURKC (rs11084490) are associated with a higher risk of GC susceptibility. Our findings also showed that the combined effect of these SNPs may influence GC risk, thus indicating the significance of assessing multiple polymorphisms, jointly. The study was conducted on a less numerous but ethnically homogeneous Bosnian population, therefore further investigations in larger and multiethnic groups and the assessment of functional impact of the results are needed to strengthen the findings.

Objectives This study aims to investigate the low-resolution human leukocyte antigen (HLA)-B locus polymorphisms between unrelated healthy individuals and patients with diagnosis of seronegative spondyloarthropathies and determine risky and protective allelic groups and genotypes. Patients and methods The study included 104 healthy control individuals (52 males, 52 females; median age 43 years; range 2 to 76 years) and 96 patients (43 males, 53 females; median age 28.5 years; range 2 to 67 years) diagnosed with: ankylosing spondylitis (AS) (n=19), reactive arthritis (n=19), psoriatic arthritis (n=28) and undifferentiated spondyloarthropathies (n=30). Genomic deoxyribonucleic acid was extracted from peripheral blood to detect allelic groups of HLA class I and II. Single-specific-primer polymerase chain reaction was used for HLA genotyping and visualization of products after their separation on 1.5% agarose gel for horizontal gel electrophoresis. Results Significantly increased frequency was found for HLA-A*02 and HLA-B*27 allelic variants in all groups of patients. The increased frequency of the HLA-B*35 allelic group in the control group represents the protective gene variant for the occurrence of AS. The predisposing genotype (HLA-B*27/B*44 and B*27/B*51) for the onset of disease was only found in AS patients. Conclusion This study shows the strong association of HLA-B*27 antigen with spondyloarthropathies, which is considered a risk variant of the gene for the onset of disease. Protective and risky allelic variants and genotypes are rare and their detection as well as increased frequency are possible if larger numbers of patients are involved.

Background: The gene for 5,10-methylenetetrahydrofolate reductase (NAD(P)H) or MTHFR gene encodes protein methylenetetrahydrofolate reductase (MTHFR), an enzyme important in folate metabolism. Aim: The aim of this study was to determine the frequencies of 677C>T and 1298A>C polymorphisms in the MTHFR gene of healthy subjects from the population. Material and methods: The blood samples were collected from 164 unrelated and healthy donors from population consisted of 98 females and 66 males. Both the MTHFR 677C>T and 1298A>C single nucleotide polymorphisms (SNPs) were analyzed by Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Linkage disequilibrium (LD) between pair of SNPs was calculated through Haploview analysis. Results: The frequency of MTHFR 677T allele in the population (32.62%) was in agreement with the frequency of this allele in most other populations, however, the frequency of MTHFR 1298C allele (38.41%) was higher than that reported for most other populations in the world. Haploview analysis showed a relatively strong LD between 677C>T and 1298A>C SNPs with D′ values of 0.87. Conclusion: Regarding the two MTHFR polymorphisms, three of the nine combined genotypes were present in 87.2% of the population. 33.54% subjects were complex heterozygous (677CT/1298AC genotype), 34.15% subjects had 677CC/1298AC and 19.51% of 677CT/1298AA genotype. The subjects with 677TT genotype had a 1298AA or 1298AC genotype while subjects with 1298CC genotype had only 677CC genotype. The subjects with 677CC/1298AA genotype were only 3.05%. We were not found triple 677CT/1298CC and quadruple 677TT/1298CC mutation suggesting decreased viability of embryos with increased numbers of mutant alleles.

AIM: The research was conducted by genotyping two Human Leukocyte Antigen (HLA) gene classes. The main objective of this research was to investigate distribution and frequency of the allelic groups, genotypes and haplotypes in the gene loci of HLA class I (HLA-A*, -B*, -C*) and HLA class II (HLA-DRB1*, -DQB1*) in patients included in the program of cadaveric renal transplantation. MATERIAL AND METHODS: Our study covered 186 blood samples of patients who are registered on the list for cadaveric renal transplantation in Federation of Bosnia and Herzegovina and included 59 control, healthy unrelated individuals. For the HLA typing, we have used three different methods: micro lymphocyte cytotoxicity test (MLCT), Polymerase Chain Reaction (PCR) – Sequence Specific Primers (SSP) and PCR – Sequence-Specific Oligonucleotides (SSO) or Luminex technology. All patients and cadaveric donors were tested using the three methods because the system is polymorphic. RESULTS: Analysis of the results of genotyping HLA class I gene loci identified dominant HLA-A*02, HLA-B*35, HLA-C*07 allelic groups. Analysis of the HLA class II gene loci genotyping showed that HLA-DRB1*11 and HLA-DQB1*03 loci had the highest incidence in HLA class II. CONCLUSION: Based on our results and previous research, there were no observed differences between allelic frequencies and genotypes of healthy people and people with ESRD. Differences between allelic groups occurred, but they were not statistically significant, except HLA-C*01 (p = 0.020).