Expression of RASSF1A, DIRAS3, and AKAP9 Genes in Thyroid Lesions: Implications for Differential Diagnosis and Prognosis of Thyroid Carcinomas.

Thyroid carcinoma is the primary endocrine malignancy worldwide. The preoperative examination of thyroid tissue lesion is often unclear. Approximately 25% of thyroid cancers cannot be diagnosed definitively without post-surgery histopathological examination. The assessment of diagnostic and differential markers of thyroid cancers is needed to improve preoperative diagnosis and reduce unnecessary treatments. Here, we assessed the expression of RASSF1A, DIRAS3, and AKAP9 genes, and the presence of BRAF V600E point mutation in benign and malignant thyroid lesions in a Polish cohort (120 patients). We have also performed a comparative analysis of gene expression using data obtained from the Gene Expression Omnibus (GEO) database (307 samples). The expression of RASSF1A and DIRAS3 was decreased, whereas AKAP9's was increased in pathologically changed thyroid compared with normal thyroid tissue, and significantly correlated with e.g., histopathological type of lesion papillary thyroid cancer (PTC) vs follicular thyroid cancer (FTC), patient's age, tumour stage, or its encapsulation. The receiver operating characteristic (ROC) analysis for the more aggressive FTC subtype differential marker suggests value in estimating RASSF1A and AKAP9 expression, with their area under curve (AUC), specificity, and sensitivity at 0.743 (95% CI: 0.548-0.938), 82.2%, and 66.7%; for RASSF1A, and 0.848 (95% CI: 0.698-0.998), 54.8%, and 100%, for AKAP9. Our research gives new insight into the basis of the aggressiveness and progression of thyroid cancers, and provides information on potential differential markers that may improve preoperative diagnosis.

Assessment of BMP7, SMAD4, and CDH1 Expression Profile and Regulatory miRNA-542-3p in Eutopic and Ectopic Endometrium of Women with Endometriosis.

Alterations in the expression of numerous genes and the miRNAs that are recognized as their regulators in the endometrial cells of women with endometriosis may disrupt the intracellular signaling pathways associated with epithelial-mesenchymal transition (EMT). So far, the functional role of BMP7 in endometrial physiology has been confirmed, especially in the context of fertility, but the role of the activation of a specific mechanism operating through the BMP-SMAD-CDH1 axis in the formation of endometrial lesions remains unexplored. The aim of this study was to evaluate the expression profile of miR-542-3p and the EMT markers (BMP7, SMAD4, CDH1) in matched eutopic endometrium (EUE) and ectopic endometrium (ECE) samples from women with endometriosis in relation to healthy women. The levels of expression of the studied genes and miRNA in peripheral blood mononuclear cells (PBMCs) obtained from women diagnosed with endometriosis and those without the disease were also evaluated. Fifty-four patients (n = 54: with endometriosis-n = 29 and without endometriosis-n = 25) were included in the study. A comparative analysis of the relative mean expression values (RQ) of the studied mRNA and miRNA assessed by RT-qPCR demonstrated downregulation of BMP7, SMAD4, and CDH1 expression in ectopic lesions and upregulation in the eutopic endometrium compared with the control group. In the eutopic tissue of women with endometriosis, miR-542-3p expression was similar to that of the control but significantly lower than in endometrial lesions. We also confirmed a trend towards a negative correlation between miR-542-3p and BMP7 in ectopic tissue, and in PBMC, a significant negative correlation of miR-542-3p with further BMP signaling genes, i.e., SMAD4 and CDH1, was observed. These results indicate that the miRNA selected by us may be a potential negative regulator of BMP7-SMAD4-CDH1 signaling associated with EMT. The different patterns of BMP7, SMAD4, and CDH1 gene expression in ECE, EUE, and the control endometrium observed by us suggests the loss of the endometrial epithelium phenotype in women with endometriosis and demonstrates their involvement in the pathogenesis and pathomechanism of this disease.

The Expression of TGF-ß1, SMAD3, ILK and miRNA-21 in the Ectopic and Eutopic Endometrium of Women with Endometriosis.

The molecular pathogenesis of endometriosis has been associated with pathological alterations of protein expression via disturbances in homeostatic genes, miRNA expression profiles, and signaling pathways that play an essential role in the epithelial-mesenchymal transition (EMT) process. TGF-ß1 has been hypothesized to play a key role in the development and progression of endometriosis, but the activation of a specific mechanism via the TGF-ß-SMAD-ILK axis in the formation of endometriotic lesions is poorly understood. The aim of this study was to assess the expression of EMT markers (TGF-ß1, SMAD3, ILK) and miR-21 in ectopic endometrium (ECE), in its eutopic (EUE) counterpart, and in the endometrium of healthy women. The expression level of the tested genes and miRNA was also evaluated in peripheral blood mononuclear cells (PBMC) in women with and without endometriosis. Fifty-four patients (n = 54; with endometriosis, n = 29, and without endometriosis, n = 25) were enrolled in the study. The expression levels (RQ) of the studied genes and miRNA were evaluated using qPCR. Endometriosis patients manifested higher TGF-ß1, SMAD3, and ILK expression levels in the eutopic endometrium and a decreased expression level in the ectopic lesions in relation to control tissue. Compared to the endometrium of healthy participants, miR-21 expression levels did not change in the eutopic endometrium of women with endometriosis, but the RQ was higher in their endometrial implants. In PBMC, negative correlations were found between the expression level of miR-21 and the studied genes, with the strongest statistically significant correlation observed between miR-21 and TGF-ß1. Our results suggest the loss of the endometrial epithelial phenotype defined by the differential expression of the TGF-ß1, SMAD3 and ILK genes in the eutopic and ectopic endometrium. We concluded that the TGF-ß1-SMAD3-ILK signaling pathway, probably via a mechanism related to the EMT, may be important in the pathogenesis of endometriosis. We also identified miR-21 as a possible inhibitor of this TGF-ß1-SMAD3-ILK axis.

Effect of physical training on parathyroid hormone and bone turnover marker profile in relation to vitamin D supplementation in soccer players.

The aim of the study was to assess the impact of vitamin D supplementation and regular physical activity on 25-hydroxyvitamin D [25(OH)D], parathyroid hormone (PTH) and bone turnover marker concentrations in healthy male athletes. Twenty-five youth soccer players were divided into groups: non-supplemented (GN) and supplemented (GS) with a vitamin D dose of 20 000 IU twice a week for 8 weeks. The study was conducted during an 8-week preseason period, from mid-January to mid-March. At baseline (T1) and at the end of this period (T2), the serum concentrations of 25(OH)D, (PTH), osteocalcin (OC) and ß-isomerized C-terminal telopeptide of type I collagen (ß-CTx) were measured. At T2, 25(OH)D increased by 70% in GS (p = 0.004) and by 6% in GN (p > 0.05). Significant differences between GS and GN groups were observed throughout the study in the group-by-time interaction and changes of 25(OH)D (p = 0.002; η 2 p = 0.36) and OC (p = 0.008; η 2 p = 0.26). Increased OC (ES = 0.74; moderate) and ß-CTx (ES = 1.31, large) in GN athletes who had an optimal baseline vitamin D level (GO) were observed. In GN, at T2, ß-CTx positively correlated with PTH and OC (p = 0.007 and p = 0.002). In GS, ß-CTx positively correlated with OC at both time points (T1, p = 0.027 and T2, p = 0.037). A negative correlation between 25(OH)D and PTH was observed at T2 (p = 0.018). The obtained results suggest that the 20 000 IU vitamin D3 dose applied twice a week for 8 weeks is effective for vitamin D compensation and sufficient to maintain the correct PTH concentration, as revealed by changes in the bone marker concentrations. In conclusion, the results suggest that the applied vitamin D supplementation dose in athletes leads to intensive bone remodelling and has protective effects on bone under intensive physical effort.

Effect of Vitamin D3 Supplementation on Interleukin 6 and C-Reactive Protein Profile in Athletes.

Vitamin D3 has a preventive, anti-inflammatory effect. However, there are still few studies linking the effects of athlete training to vitamin D3 supplementation and the immune response. The study evaluated the impact of vitamin D3 supplementation on interleukin 6 (IL-6) release during physical exercise in relation to C-reactive protein (CRP) levels in healthy male athletes. Twenty-five soccer players were divided into two groups-with (GS) and without (GN) vitamin D3 supplementation in a dose of 20,000 IU twice a week for 8 wk (about 6,000 IU/d). At the baseline (T1) and at the end (T2) of the training cycle serum concentrations of 25-hydroxyvitamin D [25(OH)D], IL-6 and CRP were measured. In the GS group, we observed a significant increase in 25(OH)D concentration (p=0.004), and non-significantly increased levels (p>0.05) of IL-6 and CRP. At the baseline, CRP in the supplemented athletes who had suboptimal vitamin D3 concentration in T1 (GSO) was significantly higher than in those with an optimal baseline vitamin D3 level (GO) (p=0.028). However, in GO in T2, a non-significant trend of negative correlation (p=0.055) between 25(OH)D concentration and IL-6 level was found. In the total study group (TG), a statistically significant (p=0.021) negative correlation in T1 was observed between 25(OH)D and CRP. However, our results do not support the immune-modulatory effect of vitamin D3 supplementation in a dose of 6,000 IU/d in athletes, in relation to IL-6 production and its subsequent stimulatory effect on CRP releasing.

Correlation between the Positive Effect of Vitamin D Supplementation and Physical Performance in Young Male Soccer Players.

The aim of this study was to determine whether supplementation with vitamin D during eight weeks of high-intensity training influences muscle power and aerobic performance in young soccer players. A total of 25 athletes were divided into two groups: the supplemented group (GS; n = 12; vitamin D 20,000 IU, twice a week) and the non-supplemented group (GN; n = 13). A set of measurements, including sprint tests, explosive power test, maximal oxygen uptake (VO2max), and serum 25(OH)D concentration, were obtained before (T1) and after (T2) the intervention. A significant group x time interaction was found in the 25(OH)D serum levels (p = 0.002; ES = 0.36, large). A significant improvement in VO2max was found in the TG (p = 0.0004) and the GS (p = 0.031). Moreover, a positive correlation between 25(OH)D and VO2max (R = 0.4192, p = 0.0024) was calculated. The explosive power tests revealed insignificant time interactions in the average 10-jump height and average 10-jump power (p = 0.07, ES = 0.13; p = 0.10, ES = 0.11, respectively). A statistically insignificant trend was observed only in the group-by-time interaction for the sprint of 10 m (p = 0.05; ES = 0.15, large). The present study provides evidence that vitamin D supplementation has a positive but trivial impact on the explosive power and locomotor skills of young soccer players, but could significantly affect their aerobic performance.

An Analysis of IL-10, IL-17A, IL-17RA, IL-23A and IL-23R Expression and Their Correlation with Clinical Course in Patients with Psoriasis.

Being one of the most common dermatological inflammatory disorders, psoriasis is a frequent subject of research. It is considered to be a T cell-dependent immune disease whose pathogenesis is influenced by cytokines, such as IL-10, IL-17A, IL-17RA, IL-23A and IL-23R. The present study examines whether the expression of selected genes is correlated with the clinical course of psoriasis, assessed by the PASI, BSA and DLQI scales. Skin biopsies and blood from 60 patients with psoriasis and 24 healthy controls were obtained for RNA isolation. These were subjected to RT-PCR for IL-10, IL-17A, IL-17RA, IL-23A and IL-23R genes. The results were presented as an RQ value. IL-17A and IL-23R expression levels were higher in psoriatic skin compared to controls, while IL-10 expression was lower. A positive correlation was also found between RQ for IL-23A and PASI index. Psoriatic skin is characterised by elevated expression of IL-17A and IL-23R and decreased expression of IL-10. This indicates that the selected cytokines may be one of the factors involved in the pathogenesis and pathomechanism of psoriasis, but more studies need to be made before we can elucidate the exact reason for the unbalance in cytokine expression levels.

Diagnostic value of PPARδ and miRNA-17 expression levels in patients with non-small cell lung cancer.

The PPARδ gene codes protein that belongs to the peroxisome proliferator-activated receptor (PPAR) family engaged in a variety of biological processes, including carcinogenesis. Specific biological and clinical roles of PPARδ in non-small cell lung cancer (NSCLC) is not fully explained. The association of PPARα with miRNA regulators (e.g. miRNA-17) has been documented, suggesting the existence of a functional relationship of all PPARs with epigenetic regulation. The aim of the study was to determine the PPARδ and miR-17 expression profiles in NSCLC and to assess their diagnostic value in lung carcinogenesis. PPARδ and miR-17 expressions was assessed by qPCR in NSCLC tissue samples (n = 26) and corresponding macroscopically unchanged lung tissue samples adjacent to the primary lesions served as control (n = 26). PPARδ and miR-17 expression were significantly lower in NSCLC than in the control (p = 0.0001 and p = 0.0178; respectively). A receiver operating characteristic (ROC) curve analysis demonstrated the diagnostic potential in discriminating NSCLC from the control with an area under the curve (AUC) of 0.914 for PPARδ and 0.692 for miR-17. Significant increase in PPARδ expression in the control for current smokers vs. former smokers (p = 0.0200) and increase in miR-17 expression in control tissue adjacent to adenocarcinoma subtype (p = 0.0422) were observed. Overexpression of miR-17 was observed at an early stage of lung carcinogenesis, which may suggest that it acts as a putative oncomiR. PPARδ and miR-17 may be markers differentiating tumour tissue from surgical margin and miR-17 may have diagnostic role in NSCLC histotypes differentiation.

Circulating miRNAs Related to Epithelial-Mesenchymal Transitions (EMT) as the New Molecular Markers in Endometriosis.

Endometriosis is a chronic gynecological disease defined by the presence of endometrial-like tissue found outside the uterus, most commonly in the peritoneal cavity. Endometriosis lesions are heterogenous but usually contain endometrial stromal cells and epithelial glands, immune cell infiltrates and are vascularized and innervated by nerves. The complex etiopathogenesis and heterogenity of the clinical symptoms, as well as the lack of a specific non-invasive diagnostic biomarkers, underline the need for more advanced diagnostic tools. Unfortunately, the contribution of environmental, hormonal and immunological factors in the disease etiology is insufficient, and the contribution of genetic/epigenetic factors is still fragmentary. Therefore, there is a need for more focused study on the molecular mechanisms of endometriosis and non-invasive diagnostic monitoring systems. MicroRNAs (miRNAs) demonstrate high stability and tissue specificity and play a significant role in modulating a range of molecular pathways, and hence may be suitable diagnostic biomarkers for the origin and development of endometriosis. Of these, the most frequently studied are those related to endometriosis, including those involved in epithelial-mesenchymal transition (EMT), whose expression is altered in plasma or endometriotic lesion biopsies; however, the results are ambiguous. Specific miRNAs expressed in endometriosis may serve as diagnostics markers with prognostic value, and they have been proposed as molecular targets for treatment. The aim of this review is to present selected miRNAs associated with EMT known to have experimentally confirmed significance, and discuss their utility as biomarkers in endometriosis.

Serum Extracellular Vesicle-Derived miRNAs in Patients with Non-Small Cell Lung Cancer-Search for Non-Invasive Diagnostic Biomarkers.

The aim of the study was a search for diagnostic and/or prognostic biomarkers in patients with non-small cell lung cancer (NSCLC) patients, based on circulating microRNAs (miRs: miR-23a, miR-361, miR-1228 and miR-let7i) in extracellular vesicles (EVs). Serum EVs were isolated from NSCLC patients (n = 31) and control subjects (n = 21). RNA was isolated from EVs and reverse transcription reaction was performed. Relative levels of miR-23a, miR-361, miR-1228 and miR-let7i were assessed in real-time qPCR using TaqMan probes. Analysis was based on the 2-ΔΔCT method. Statistically significant lower levels of miR-23a and miR-let7i were observed among NSCLC patients vs. control group: miR-23a, 0.054 vs. 0.107; miR-let7i, 0.193 vs. 0.369 (p = 0.003, p = 0.005, respectively). A receiver operating characteristic (ROC) curve analysis demonstrated the diagnostic potential of each individual serum EV-derived miRNA with an area under the curve AUC = 0.744 for miR-23a (p = 0.0003), 0.733 for miR-let7i (p = 0.0007). The decreased level of miR-23a in patients correlated with metastasis to lymph nodes and with AJCC tumor staging system. The results demonstrate that miR-23a and miR-let7i may prove clinically useful as significant, non-invasive markers in NSCLC diagnosis. Additionally, changing profile level of miR-23a that correlates with cancer development may be considered as an NSCLC progression marker.

Assessment of Wnt pathway selected gene expression levels in peripheral blood mononuclear cells (PBMCs) of postmenopausal patients with low bone mass.

The purpose of the study was to assess the expression of selected genes of the Wnt pathway: APC, AXIN1, CTNNB1, DKK1, GSK3ß, KREMEN1, SFRP1, and WNT1 in peripheral blood mononuclear cells (PBMC) of patients, selected in consideration of their bone mineral density (BMD), and the occurrence of low-energy fractures. The study involved 45 postmenopausal women, divided into four groups, according to BMD and fracture history. Measurements of laboratory parameters and RNA expression in PBMC cells were carried out in material, collected once at the inclusion visit. The densitometric examination was performed on all participants. In the analysis of the relative expression levels (RELs) of the studied genes in the entire population, we observed an overexpression for SFRP1 in 100% of samples and WNT1. In addition, the REL of DKK1, APC, and GSK3ß genes were slightly elevated versus the calibrator. In contrast, CTNNB1 and AXIN1 presented with a slightly decreased RELs. Analysis did not show any significant differences among the groups in the relative gene expression levels (p < 0.05) of particular genes. However, we have observed quite numerous interesting correlations between the expression of the studied genes and BMD, the presence of fractures, and laboratory parameters, both in the whole studied population as well as in selected groups. In conclusion, the high level of CTNNB1 expression maintains normal BMD and/or protects against fractures. It also appears that the changes in expression levels of the Wnt pathway genes in PBMCs reflect the expected changes in bone tissue.

Type of training has a significant influence on the GH/IGF-1 axis but not on regulating miRNAs.

The growth hormone (GH)/insulin-like growth factor-1 axis is responsible for glucose homeostasis. In the present study we assessed the expression levels of miRNA-124, miRNA-210 and miRNA-375 and immunoexpression of IGFBP-3 in relation to the concentrations of IGF-1 and glucose in athletes performing different types of effort. Sixty-six young male athletes (age 25.4±4.1 years) were divided into: group EN (33 male athletes; age 25.6±4.4 years) with endurance-type efforts (disciplines: triathlon, long distance running, cycling) and group ST (33 male athletes; age 25.2±3.9 years) with strength-type efforts (disciplines: weightlifting, body building, CrossFit). The control group consisted of 28 non-training men (age 29.1±4.7 years). Statistically significantly higher IGF-1 concentration and lower glucose concentration (P<0.05) in serum were observed in the group of athletes (vs. controls). Immunoexpression of IGFBP-3 was higher in athletes (vs. controls), and a higher value of immunoexpression was obtained in athlete group ST vs. group EN (P>0.05). Levels of expression of miRNA-210 and miRNA-375 were higher in athletes vs. controls (P>0.05). The obtained data confirmed the importance of the somatotropic axis in the regulation of metabolic adaptation to physical exercise. The detected variation in the concentrations and expression levels of the studied molecules involved in the somatotropic axis in athletes confirmed the role of the somatotropic axis in adaptation to physical effort. Statistically significant reduction of glucose concentration and the highest expression of IGF-1in serum in athletes suggest the anabolic effect of IGF-1 through insulin receptors on many tissues under the influence of moderate physical exercises (mainly during resistance training).

Analysis of molecular markers as IL-12, IL-22 and IFN-γ in correlation with a clinical course in patients with psoriasis.

OBJECTIVES: As a chronic, recurrent, immunologically mediated systemic disease and a common cause of dermatological problems, psoriasis is often a subject of scientific research. Skin changes located on the hands can cause difficulties and limitations in the performance of professional activities, especially manual ones. The main role in pathogenesis is played by immunological factors - improper functioning of the components of the immune system, among others, T lymphocytes and cytokines like interleukin-12 (IL-12), interleukin-22 (IL-22) and interferon gamma (IFN-γ). MATERIAL AND METHODS: The obtained tissue and blood were destined for RNA isolation. The RNA was then subjected to a reverse transcription reaction. The relative gene expression level was evaluated by the real-time polymerase chain reaction for IL-12B, IL-22 and IFN-γ genes, and presented as the relative quantification (RQ) value, relative to the reference gene GAPDH. In addition, a correlation analysis of the expression level of selected genes with the clinical course of the disease, as assessed by the Psoriasis Area and Severity Index (PASI), the Body Surface Area (BSA) and the Dermatology Life Quality Index (DLQI) scores was performed. RESULTS: Statistical analysis confirmed a significant increase in RQ values for IL-12B, IL-22 and IFN-γ in the group of psoriatic patients vs. the control group. A positive correlation was also found between BSA and PASI and RQ for the IL-12B gene. CONCLUSIONS: Increased expression levels of IL-12B, IL-22 and IFN-γ genes in psoriatic skin confirm that selected cytokines play an important role in the initiation and sustenance of psoriasis. Int J Occup Med Environ Health. 2020;33(5):635-47.

Evaluation of the relationship between the IL-17A gene expression level and regulatory miRNA-9 in relation to tumor progression in patients with non-small cell lung cancer: a pilot study.

A pro-inflammatory cytokine, IL-17A, is associated with increased risk of developing numerous cancers, including non-small cell lung cancer (NSCLC). IL-17A is a target gene for miR-9. This encouraged us to analyze these two genes in terms of their usefulness as prognostic markers in NSCLC. The expression levels of IL-17A gene and miR-9 was assessed in 26 NSCLC tissue samples and 26 unchanged lung tissue adjacent to lung tumors (control tissue), using qPCR. In both tissue groups, a decreased expression of IL-17A was observed in 100% of samples. Increased expression of miRNA-9 was observed in 92% of tumor samples, and in 100% of control samples. Neither statistical differences in the level of expression IL-17A depending on the patient's age, gender, smoking status, nor histopathology of the cancer was found. Regarding the presence of nodule metastasis ('N' value in TNM classification), significantly lower expression level of IL-17A was observed in cN2 as compared with cN1 group. Additionally, statistically lower IL-17A expression was found in III versus II tumor stage (cAJCC classification). Significant negative correlation between both studied genes was revealed in SCC subgroup. This leads to the conclusion that miRNA-9 can regulate the expression of IL-17A as an IL-17A mRNA antagonistic mediator. Inhibition of proinflammatory action of IL-17A in correlation with tumor progression can be related to various activity of Th17 cells on cancer development according to its immunogenicity, and also may suggest suppressive role of IL-17A in tumor progression. However, because of low number of analyzed samples, further studies on the functional role of IL-17A in development and/or progression NSCLC seem warranted.

Investigation of Cervical Tumor Biopsies for Chromosomal Loss of Heterozygosity (LOH) and Microsatellite Instability (MSI) at the HLA II Locus in HIV-1/HPV Co-infected Women.

Background: A subgroup of women who are co-infected with human immunodeficiency virus type 1 (HIV-1) and human papillomavirus (HPV) progress rapidly to cervical disease regardless of high CD4 counts. Chromosomal loss of heterozygosity (LOH) and microsatellite instability (MSI) are early frequent genetic alterations occurring in solid tumors. Loss of an allele or part of a chromosome can have multiple functional effects on immune response genes, oncogenes, DNA damage-repair genes, and tumor-suppressor genes. To characterize the genetic alterations that may influence rapid tumor progression in some HIV-1-positive women, the extent of LOH and MSI at the HLA II locus on chromosome 6p in cervical tumor biopsy DNA samples with regard to HIV-1/HPV co-infection in South African women was investigated. Methods: A total of 164 women with cervical disease were recruited for this study, of which 74 were HIV-1-positive and 90 were HIV-1-seronegative. DNA from cervical tumors and matched buccal swabs were used for analyses. Six fluorescently-labeled oligonucleotide primer pairs in a multiplex PCR amplification were used to study LOH and MSI. Pearson chi-squared test for homogeneity of proportions using an exact p value, a two-proportion Z-score test, ROC curves and a logistic regression model were used for statistical analyses. All p-values were corrected for false discovery rate (FDR) using the Benjamini-Hochberg test and the adjusted p-values (q-values) were reported. All tests were significant when both p and q < 0.05. Results: Tumor DNA from HIV-1/HPV co-infected women demonstrated a higher frequency of LOH/MSI at the HLA II locus on chromosome 6p21.21 than tumor DNA from HIV-1-seronegative women (D6S2447, 74.2 vs. 42.6%; p = 0.001, q = 0.003), D6S2881 at 6p21.31 (78.3 vs. 42.9%; p = 0.002, q = 0.004), D6S2666 at 6p21.32 (79 vs. 57.1%; p = 0.035, q = 0.052), and D6S2746, at 6p21.33 (64.3 vs. 29.4%; p < 0.001, q < 0.001), respectively. Conclusions: HPV infection alone can induce LOH/MSI at the HLA II locus in cervical tumor DNA, whereas HIV-1 co-infection exacerbates it, suggesting that this may accelerate cervical disease progression in a subgroup of HIV-1-positive women.

An assessment of the relationship between the expression of CCR7/CCL19 axis and selected regulatory miRNAs in non-small cell lung cancer.

CC chemokine receptor type 7 (CCR7) and its ligands has been implicated in the occurrence and progression of NSCLC. Previous studies have revealed that the diagnostic value of CCR7/CCL19 axis in lung tumorigenesis remains controversial. The present study evaluates the relationship between the mRNA expression of CCR7/CCL19 axis and selected regulatory miRNAs in NSCLC patients. It analyzes the expression level of CCR7 mRNA and its ligand in tumor tissue in relation to expression level of two miRNAs: miR let-7a and miR-335, as transcriptional regulators of study genes. Twenty-seven patients (n = 27) were enrolled. The expression of the studied genes and miRNAs was evaluated by qPCR. Tumour tissue fragments, adjacent macroscopically-unchanged lung tissue (control) and patient serum were used as biological material for study. Elevated expression of CCR7 and CCL19 mRNA was observed in patients with metastasis to lymph nodes. We noticed upregulated miR-335 expression and downregulated miR let-7a expression in patient serum with regard to AJCC tumor staging. Higher miR-335 expression and lower miR let-7a expression level was observed in patients with metastasis to lymph node. The presence of changes observed in the expression level of miR-335 and miR let-7a in the serum of NSCLC patients in relation to lymph node metastases and tumor stage may serve as a non-invasive molecular biomarker of tumor progression; however, this observation requires further investigation.

Clinicopathological Significance of Overall Frequency of Allelic Loss (OFAL) in Lesions Derived from Thyroid Follicular Cell.

BACKGROUND: Loss of heterozygosity (LOH) and microsatellite instability (MSI) are frequent molecular events in thyroid tumor etiopathogenesis occurring in several chromosomal critical areas, including 3p12-25.3, 7q21-31, 10q22-24, and 15q11-13, with loci of tumor suppressor genes. OBJECTIVE: We evaluated the usefulness of LOH/MSI as a diagnostic/prognostic biomarker in lesions derived from thyroid follicular cells: follicular thyroid carcinoma (FTC); follicular adenoma (FA), papillary thyroid carcinoma (PTC), and nodular goiter (NG). METHODS: We performed allelotyping (GeneMapper Software v. 4.0.) of ten microsatellite markers linked to the 1p31.2, 3p21.3, 3p24.2, 9p21.3, 11p15.5, and 16q22.1 region on DNA from 93 primary thyroid lesions then evaluated the LOH/MSI frequency and overall frequency of allelic loss (OFAL). RESULTS: We found regions with significantly increased frequency of LOH/MSI for specific histotypes: the 3p24.2 region for FA and 1p31.2 for FTC. LOH/MSI in 3p21.3 was significantly elevated in PTC and FTC. LOH/MSI in 3p21.3 was increased for small size tumors (T1a + T1b), tumors with no regional lymph node involvement (N0 + Nx), American Joint Committee on Cancer (AJCC) stage I tumors, and tumor diameter (Td) < 10 mm; in 1p31.2 for T2-3, N1, stage II-IV, and Td 10-30 mm; in 11p15.5 for T2-3, N1, stage II-IV, and Td > 30 mm. OFAL values were significantly higher in younger patients (< 40 years), in men, in those with T2-3 stage tumors, in those with increased Td, and in FA and FTC compared with NG and PTC. CONCLUSIONS: We confirmed the occurrence of LOH/MSI in 3p21.3 at an early stage of tumorigenesis and mapped 1p31.2 and 11p15.5 as characteristic for advanced-stage tumors. The results of our study may enable consideration of OFAL, defined as LOH/MSI coincidence in various chromosomal regions, as a tumor progression marker. OFAL values were significantly higher in follicular neoplasms (FA and FTC) than in PTC or NG; hence, increased OFAL values can be regarded as a characteristic feature of the follicular phenotype.

The Expression Levels of IL-4/IL-13/STAT6 Signaling Pathway Genes and SOCS3 Could Help to Differentiate the Histopathological Subtypes of Non-Small Cell Lung Carcinoma.

BACKGROUND: The interleukin (IL)-4/IL-13/signal transducer and activator of transcription (STAT) 6 signaling pathway and the SOCS3 gene, one of its main regulators, constitute an important link between the inflammation process in the epithelial cells and inflammatory-related tumorigenesis. The present study is the first to evaluate IL-4, IL-13, STAT6, and SOCS3 mRNA expression in non-small cell lung carcinoma (NSCLC) histopathological subtypes. METHODS: Gene expression levels were assessed using TaqMan® probes by quantitative reverse transcription PCR (qRT-PCR) in lung tumor samples and unchanged lung tissue samples. RESULTS: Increased expression of IL-4, IL-13, and STAT6 was observed in all histopathological NSCLC subtypes (squamous cell carcinoma [SCC], adenocarcinoma [AC], and large cell carcinoma [LCC]). Significantly higher expression of IL-13 and STAT6 (p = 0.019 and p = 0.008, respectively) was found in SCC than in LCC. No statistically significant differences were found for IL-4. Significantly higher SOCS3 expression was found in LCC than in AC (p = 0.027). A negative correlation (rho = -0.519) was observed for the STAT6 and SOCS3 genes in SCC (p = 0.005). No associations were found between gene expression and tumor staging (post-operative Tumor Node Metastasis [pTNM], American Joint Committee on Cancer [AJCC]), patients' age, sex, or history of smoking. CONCLUSIONS: As the number of LCC cases in our study was quite low, the statistically significant results obtained should be confirmed in a larger group of patients, particularly as the relationships identified between increased IL-4, IL-13, and STAT6 mRNA expression and decreased SOCS3 expression suggest that these genes may serve as potential diagnostic markers for differentiating between NSCLC histopathological subtypes.

Immunoexpression analysis of selected JAK/STAT pathway molecules in patients with non- small-cell lung cancer.

INTRODUCTION    Signal transducer and activator of transcription (STAT) proteins are critically involved in tumorigenesis in various cancers, including lung cancer. OBJECTIVES    The aim of the study was to analyze the immunoexpression levels of 3 STAT proteins: STAT3, STAT5, and STAT6 in their phosphorylated forms (pSTATs), STAT inhibitors PIAS3 and SOCS3, and additionally cyclooxygenase 2 (COX­2), as potential diagnostic and prognostic markers in lung cancer. PATIENTS AND METHODS    The study included 71 patients diagnosed with non- small-cell lung cancer (NSCLC). The immunoexpression levels of the proteins were assessed in lung tissue samples, using an enzyme­linked immunosorbent assay. Tumors were staged using the postoperative TNM classification. RESULTS    All studied STATs were overexpressed in 54% to 55% of NSCLC specimens. Significantly higher STAT3 and STAT6 immunoexpression levels were observed in squamous cell carcinoma. Significant differences between NSCLC samples and controls were found for STAT5. Significantly higher STAT5 levels were observed in pT2 tumors. The COX­2 overexpression was observed in 55% of NSCLC specimens and was significantly higher in T2 tumors. STAT inhibitors were underexpressed in 56% to 58% of NSCLC specimens. The PIAS3 immunoexpression was significantly lower in non-squamous cell carcinoma. The SOCS3 level was significantly lower in smaller tumors (pT1). Negative correlations between STAT5 and PIAS3 levels, as well as between STAT6 and SOCS levels, and a positive correlation between STAT5 and COX-2 levels were observed. CONCLUSIONS    The deregulated expression of the studied pSTATs and their inhibitors may be involved in the development and progression of lung cancer. The observed differences between the histotypes suggest the potential usefulness of STAT proteins as diagnostic markers. Our results may contribute to the search for targets in lung cancer therapy.

Evaluation of Selected MicroRNAs Expression in Remission Phase of Multiple Sclerosis and Their Potential Link to Cognition, Depression, and Disability.

Accumulating data suggests that miRNAs might play a major role in neuroinflammatory processes. Therefore, our study aimed to first estimate the levels of miR-155, miR-326, and miR-301a in serum of RR-MS patients in the remission phase and then compare the levels of the examined miRNAs at different times after relapse. In this study, 36 RR-MS patients in the remission phase took part. We analyzed two subgroups of RR-MS: one, 1 to 2 months after completing steroid treatment during relapse (post-acute; n = 13) and the other, over 2 years without any relapse (stable; n = 23). Moreover, we made correlations between these biochemical results and clinical parameters of cognitive impairment, depression, and disability. The obtained results presented downregulation of miR-155 and miR-301a (in 94% and 51% samples, respectively) and overexpression of miR-326 (in 72% samples) in RR-MS patients. Moreover, we observed a positive correlation between the relative expression of miRNAs and BDI (Beck Depression Index) for miR-326 (rho = 0.385459, p = 0.022210; Spearman's rank correlation) and miR-301a (rho = 0.435131, p = 0.008991; Spearman rank correlation). We also observed the differences in expression levels between the post-acute and stable phases of RR-MS. The expression levels of miR-301a and miR155 were higher in the post-acute vs. stable phase of remission (2.385 vs. 0.524 and 0.594 vs. 0.147; respectively). Our study, for the first time, presents miRNA expression differences in two stages of remission: post-acute and stable.