Ewa Brzeziańska

The significance of epigenetic alterations in lung carcinogenesis

Lung cancer is recognized as a leading cause of cancer-related death worldwide and its frequency is still increasing. The prognosis in lung cancer is poor and limited by the difficulties of diagnosis at early stage of disease, when it is amenable to surgery treatment. Therefore, the advance in identification of lung cancer genetic and epigenetic markers with diagnostic and/or prognostic values becomes an important tool for future molecular oncology and personalized therapy. As in case of other tumors, aberrant epigenetic landscape has been documented also in lung cancer, both at early and late stage of carcinogenesis. Hypermethylation of specific genes, mainly tumor suppressor genes, as well as hypomethylation of oncogenes and retrotransposons, associated with histopathological subtypes of lung cancer, has been found. Epigenetic aberrations of histone proteins and, especially, the lower global levels of histone modifications have been associated with poorer clinical outcome in lung cancer. The recently discovered role of epigenetic modifications of microRNA expression in tumors has been also proven in lung carcinogenesis. The identified epigenetic events in lung cancer contribute to its specific epigenotype and correlated phenotypic features. So far, some of them have been suggested to be cancer biomarkers for early detection, disease monitoring, prognosis, and risk assessment. As epigenetic aberrations are reversible, their correction has emerged as a promising therapeutic target.

CTLA-4 Expression and Polymorphisms in Lung Tissue of Patients with Diagnosed Non-Small-Cell Lung Cancer

Cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) is a potent immunoregulatory molecule that downregulates T-cell activation and thus influences the antitumor immune response. CTLA-4 polymorphisms are associated with various cancers, and CTLA-4 mRNA/protein increased expression is found in several tumor types. However, most of the studies are based on peripheral blood mononuclear cells, and much less is known about the relationship between CTLA-4 expression, especially gene expression, and its polymorphic variants in cancer tissue. In our study we assessed the distribution of CTLA-4 two polymorphisms (+49A/G and −318C/T), using TaqMan probes (rs231775 and rs5742909, resp.), and CTLA-4 gene expression in real-time PCR assay in non-small-cell lung cancer (NSCLC) tissue samples. The increased CTLA-4 expression was observed in the majority of NSCLC patients, and it was significantly correlated with TT genotype (−318C/T) and with tumor size (T2 versus T3 + T4). The presence of G allele and GG genotype in cancer tissue (+49A/G) was significantly associated with the increased NSCLC risk. Additionally, we compared genotype distributions in the corresponding tumor and blood samples and found statistically significant differences. The shift from one genotype in the blood to another in the tumor may confirm the complexity of gene functionality in cancer tissue.

GENE DOPING IN SPORT – PERSPECTIVES AND RISKS

In the past few years considerable progress regarding the knowledge of the human genome map has been achieved. As a result, attempts to use gene therapy in patients’ management are more and more often undertaken. The aim of gene therapy is to replace defective genes in vivo and/or to promote the long-term endogenous synthesis of deficient protein. In vitro studies improve the production of human recombinant proteins, such as insulin (INS), growth hormone (GH), insulin-like growth factor-1 (IGF-1) and erythropoietin (EPO), which could have therapeutic application. Unfortunately, genetic methods developed for therapeutic purposes are increasingly being used in competitive sports. Some new substances (e.g., antibodies against myostatin or myostatin blockers) might be used in gene doping in athletes. The use of these substances may cause an increase of body weight and muscle mass and a significant improvement of muscle strength. Although it is proven that uncontrolled manipulation of genetic material and/or the introduction of recombinant proteins may be associated with health risks, athletes are increasingly turning to banned gene doping. At the same time, anti-doping research is undertaken in many laboratories around the world to try to develop and refine ever newer techniques for gene doping detection in sport. Thanks to the World Anti-Doping Agency (WADA) and other sports organizations there is a hope for real protection of athletes from adverse health effects of gene doping, which at the same time gives a chance to sustain the idea of fair play in sport.

Expression of Insulin‐like Growth Factor I (IGF‐I) Gene and of Genes for IGF‐Binding Proteins 1, 2, 3, 4 (IGFBP‐1—IGFBP‐4) in Non‐neoplastic Human Thyroid Cells and in Certain Human Thyroid Cancers. Effect of Exogenous IGF‐I on This Expression

Aim: The aim of this study was to examine the expression of the IGF‐I gene and of genes for IGFBP‐1, ‐2, ‐3, and ‐4 in cells from nodular goiters (NG), and from different human thyroid carcinomas (papillary—PTC, anaplastic—ATC, and medullary—MTC), cultured in monolayers. The influence, exerted by exogenous IGF‐I on the expression of these genes, was also investigated. Methods: Thyroid tissue specimens were obtained from 65 patients during subtotal or total thyroidectomies. After approximately 2–3 weeks of culture, thyroid cells were incubated for 24 hours with IGF‐I in concentrations of: 0, 1, 10 and 100 ng/ml. The total mRNA was isolated according to the method described by Chomczyński and Sacchi with our own modifications. Afterwards, mRNA encoding IGF‐I, IGFBP‐1—IGFBP‐4 and glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH), were amplified, using the reverse transcription‐polymerase chain reaction (RT‐PCR); GAPDH gene served as a control gene. PCR products were electrophoresed and then submitted to densitometric analysis. Results and conclusions: Our study has shown that in carcinoma cells (ATC, PTC, MTC), IGF‐I reveals a stimulatory influence on the expression of its own gene, that effect being most distinctive in ATC cells. These facts indicate an important role of IGF‐I in the pathogenesis and invasiveness of the analyzed malignant neoplasms.

Assessment of OPG/RANK/RANKL Gene Expression Levels in Peripheral Blood Mononuclear Cells (PBMC) After Treatment With Strontium Ranelate and Ibandronate in Patients With Postmenopausal Osteoporosis

CONTEXT Recent research results have confirmed the high significance of the OPG/RANK/RANKL system in the development of bone diseases. AIM The aim of the reported study was to assess gene expression levels of the OPG/RANK/RANKL system in peripheral blood mononuclear cells (PBMCs) after strontium ranelate (SR) and ibandronate administered to patients with postmenopausal osteoporosis. PATIENTS AND METHODS A total of 89 postmenopausal women, aged 51 to 85 years, patients of the Outpatient Clinic of Osteoporosis of the Military Teaching Hospital in Lodz, were enrolled into the study. The patients were randomly assigned to different medical therapies: ibandronate and SR. Patients of the control group received only calcium and vitamin D₃ supplements. Patient visits were repeated after 3 and 6 months. Measurements of serum alkaline phosphatase concentrations and of RNA expression in PBMCs as well as of total serum calcium and phosphate levels and of their 24-hour urine excretion rates were carried out in material, collected at baseline and after 3 and 6 months of the therapy. Densitometry of the left hip and of the lumbar spine was done at the baseline visit and after 6 months. RESULTS The differences in gene expressions of RANKL and RANK were not significant during the study period and did not differ between the groups in a statistically significant manner. No OPG gene expression was observed in PBMCs of patients in any of the studied groups and at any time point. The tendency of correlation (P = .07) was observed between decreasing RANK gene expression and increasing bone mineral density in the patients treated with SR. CONCLUSIONS Both ibandronate and SR do not seem to cause any significant changes in gene expression levels of OPG/RANK/RANKL in PBMCs during the first 6 months of treatment.

Single nucleotide polymorphisms in the STAT3 gene influence AITD susceptibility, thyroid autoantibody levels, and IL6 and IL17 secretion

Abstract STAT3 (signal transducer and activator of transcription 3) is an important cellular effector in the Jak/STAT signaling pathway, which plays a pivotal role in human immune system regulation, mediating the effect of different cytokines. In the present study, we assessed the correlation between STAT3 polymorphisms (rs3816769 C>T and rs744166 A>G) and risk of the autoimmune thyroid diseases (AITDs) Hashimoto’s thyroiditis (HT) and Graves’ disease (GD) in the Polish population. Moreover, we evaluated the association of polymorphisms with the thyroid autoantibody levels (TPOAb, TgAb, TRAb) and the correlation between circulating proinflammatory IL6 and IL17 cytokines and thyroid autoantibody levels. The study included 71 AITD patients with HT (n = 39) or GD (n = 32) and a control group (n = 40). DNA SNP genotyping was performed using TaqMan probes. Serum levels of thyroid autoantibodies, IL6 and IL17 were measured according to enhanced chemiluminescence (ECL) assay. Allele A of STAT3 SNP rs744166 A>G was significantly more frequent in both HT and GD patients, while allele G was significantly more frequent in the control group. Similarly, allele C and CC genotype of STAT3 SNP rs3816769 C>T were significantly more frequent in the control group in comparison to HT and GD patients. Significantly higher TgAb median values were associated with CT rs3816769 genotype in HT patients. Serum levels of IL6 and IL17 positively correlated with TPOAb in the HT group. Serum level of IL6 positively correlated with TPOAb in the AITD group. Both studied polymorphisms seem to play a significant role in susceptibility to AITD (HT and GD). STAT3 SNPs may influence TAb level in AITD patients.

Association of Trp64Arg Polymorphism of β3-Adrenergic Receptor with Insulin Resistance in Polish Children with Obesity

AIM To establish the influence of the Trp64Arg variant of the beta3-adrenergic receptor (Trp64Arg- beta3AR) on body mass index (BMI) and insulin resistance (IR) in obese children. METHODS BMI, presence of the Trp64Arg mutation, plasma glucose and insulin concentrations during an oral glucose tolerance test (OGTT) and IR were determined in 60 obese and 33 normal weight children. RESULTS The frequency of Trp64Arg was similar in normal weight and obese children. BMI, glucose and insulin concentrations during an OGTT in children with Trp64Argbeta3AR were not different from those with Trp64Trpbeta3AR. IR was confirmed in 42.8% of children with Trp64Argbeta3AR and in 45.6% of children with Trp64Trpbeta3AR (NS). CONCLUSIONS 1. The similar frequency of the Trp64Argbeta3AR variant in normal weight and obese children suggests that it is not a susceptibility gene for obesity in Polish children. 2. The presence of the Trp64Argbeta3AR variant does not have an unfavourable influence on BMI, glucose or insulin concentrations during OGTT or on IR frequency in Polish obese children.