Analysis of the Gut Microbiome and Dietary Habits in Metastatic Melanoma Patients with a Complete and Sustained Response to Immunotherapy.

Immunotherapy has improved the prognosis of metastatic melanoma patients, although most patients do not achieve a complete response. While specific gut microbiome and dietary habits might influence treatment success, there is a lack of concordance between the studies, potentially due to dichotomizing patients only into responders and non-responders. The aim of this study was to elucidate whether metastatic melanoma patients with complete and sustained response to immunotherapy exhibit differences in gut microbiome composition among themselves, and whether those differences were associated with specific dietary habits. Shotgun metagenomic sequencing revealed that patients who exhibited a complete response after more than 9 months of treatment (late responders) exhibited a significantly higher beta-diversity (p = 0.02), with a higher abundance of Coprococcus comes (LDA 3.548, p = 0.010), Bifidobacterium pseudocatenulatum (LDA 3.392, p = 0.024), and lower abundance of Prevotellaceae (p = 0.04) compared to early responders. Furthermore, late responders exhibited a different diet profile, with a significantly lower intake of proteins and sweets and a higher intake of flavones (p < 0.05). The research showed that metastatic melanoma patients with a complete and sustained response to immunotherapy were a heterogeneous group. Patients with a late complete response exhibited microbiome and dietary habits which were previously associated with an improved response to immunotherapy.

Potential of Coupling Metaheuristics-Optimized-XGBoost and SHAP in Revealing PAHs Environmental Fate.

Polycyclic aromatic hydrocarbons (PAHs) refer to a group of several hundred compounds, among which 16 are identified as priority pollutants, due to their adverse health effects, frequency of occurrence, and potential for human exposure. This study is focused on benzo(a)pyrene, being considered an indicator of exposure to a PAH carcinogenic mixture. For this purpose, we have applied the XGBoost model to a two-year database of pollutant concentrations and meteorological parameters, with the aim to identify the factors which were mostly associated with the observed benzo(a)pyrene concentrations and to describe types of environments that supported the interactions between benzo(a)pyrene and other polluting species. The pollutant data were collected at the energy industry center in Serbia, in the vicinity of coal mining areas and power stations, where the observed benzo(a)pyrene maximum concentration for a study period reached 43.7 ngm-3. The metaheuristics algorithm has been used to optimize the XGBoost hyperparameters, and the results have been compared to the results of XGBoost models tuned by eight other cutting-edge metaheuristics algorithms. The best-produced model was later on interpreted by applying Shapley Additive exPlanations (SHAP). As indicated by mean absolute SHAP values, the temperature at the surface, arsenic, PM10, and total nitrogen oxide (NOx) concentrations appear to be the major factors affecting benzo(a)pyrene concentrations and its environmental fate.

Study on the immunopathological effect of titanium particles in peri-implantitis granulation tissue: A case-control study.

OBJECTIVES: To identify titanium particles (TPs) in biopsy specimens harvested from peri-implantitis lesions and secondarily to study the histopathological characteristics in peri-implantitis compared to periodontitis, in order to evaluate whether the presence of TPs could alter respective inflammatory patterns. MATERIAL AND METHODS: Biopsies containing granulation tissue were harvested during routine surgical treatment in 39 peri-implantitis cases and 35 periodontitis controls. Serial sections were obtained using titanium-free microtome blades. The first and last sections of the peri-implantitis specimens were used for identification of TPs by scanning electron microscopy coupled with dispersive X-ray spectrometry. Intermediate sections and periodontitis specimens were processed for descriptive histological study using haematoxylin-eosin staining and for immunohistochemical analysis using CD68, IL-6, Nf-kB and VEGF markers. RESULTS: TPs were identified in all peri-implantitis specimens as free metal bodies interspersed within granulation tissue. However, presence of macrophages or multinucleated giant cells engulfing the TPs were not identified in any specimen. Peri-implantitis granulations were characterized by a chronic inflammatory infiltrate rich in neutrophils. About half of peri-implantitis patients exhibited a subacute infiltrate characterized with lymphocytes interweaved with neutrophils and eosinophils. When compared to periodontitis, peri-implantitis tissues showed higher proportions of macrophages and a more intense neovascularization, based on significantly higher expression of CD68 and VEGF respectively. CONCLUSION: TPs were identified in all peri-implantitis specimens, but without evidencing any foreign body reaction suggestive for direct pathological effects of TPs. The peri-implantitis granulation tissue was characterized by intense neovascularization and presence of a chronic inflammatory infiltrate dominated by plasma cells, neutrophils and macrophages.

The PM2.5-bound polycyclic aromatic hydrocarbon behavior in indoor and outdoor environments, part II: Explainable prediction of benzo[a]pyrene levels.

Among the polycyclic aromatic hydrocarbons (PAH), benzo[a]pyrene (B[a]P) has been considered more relevant than other species when estimating the potential exposure-related health effects and has been recognized as a marker of carcinogenic potency of air pollutant mixture. The current understanding of the factors which govern non-linear behavior of B[a]P and associated pollutants and environmental processes is insufficient and further research has to rely on the advanced analytical approach which averts the assumptions and avoids simplifications required by linear modeling methods. For the purpose of this study, we employed eXtreme Gradient Boosting (XGBoost), SHapley Additive exPlanations (SHAP) attribution method, and SHAP value fuzzy clustering to investigate the concentrations of inorganic gaseous pollutants, radon, PM2.5 and particle constituents including trace metals, ions, 16 US EPA priority PM2.5-bound PAHs and 31 meteorological variables, as key factors which shape indoor and outdoor PM2.5-bound B[a]P distribution in a university building located in the urban area of Belgrade (Serbia). According to the results, the indoor and outdoor B[a]P levels were shown to be highly correlated and mostly influenced by the concentrations of Chry, B[b]F, CO, B[a]A, I[cd]P, B[k]F, Flt, D[ah]A, Pyr, B[ghi]P, Cr, As, and PM2.5 in both indoor and outdoor environments. Besides, high B[a]P concentration events were recorded during the periods of low ambient temperature (<12 °C), unstable weather conditions with precipitation and increased soil humidity.

Human health risks and benefits assessment based on OCPs, PCBs, toxic elements and fatty acids in the pelagic fish species from the Adriatic Sea.

Benefits and risks of the fish consumption should be assessed to balance the ingestion of healthy omega-3 fatty acids and adverse chemicals. Persistent organic pollutants-POPs (organochlorine pesticides-OCPs and polychlorinated biphenyls-PCB), macro- and micro-elements and fatty acid contents were determined in six fish species from the Adriatic Sea to assess health risks for consumers (worst-case scenario, diseases development risks and benefit-risk). 16 element, 24 POPs and 14 fatty acid contents were determined in six pelagic species which can be used in human diet. Element concentrations were determined by inductively coupled plasma mass spectrometry (ICP-MS), POPs by high-resolution gas chromatography and fatty acid content by gas-liquid chromatography. Diet based on chub mackerel and round sardinella showed lower daily intake (DI) of POPs and highest DI of essential omega-3 fatty acids than other investigated species. Lower ingestion of toxic elements can be observed by consuming anchovy and round sardinella. Based on POP concentrations, there was not observed non-carcinogenic (HI) nor carcinogenic (CR) risks for consumers. Based on element concentrations, there was low HI (0.1 ≥ HI ≥ 1), while the maximum HIs and outlier values (horse mackerel and anchovy samples) implied the presence of HI (HI > 1). The most significant contributor to total non-carcinogenic and carcinogenic risks was inorganic arsenic (inorganic As). Acceptable CR for consumers was assessed, but maximum CR for consumers of horse mackerel and anchovy (CR ≥ 1 × 10-6) showed adverse effects on human health. There were low HIs for developing cardiovascular, nervous, and reproductive diseases, and maximum HIs were higher than 1. Acceptable (1 × 10-4≥CR ≥ 1 × 10-6) risks were observed for developing cancer of nervous system and reproductive organs. Among investigated fish samples, those with higher Æ©BR (benefit-risks) and BR for arsenic (As) than median value have a higher risk than benefits in the human diet.

The Efficacy of an Energy-Restricted Anti-Inflammatory Diet for the Management of Obesity in Younger Adults.

There is growing evidence of the dietary impact on obesity-induced low-grade chronic inflammation and the associated chronic non-communicable diseases modification. We determined changes in body composition and cardiometabolic and inflammatory status of participants with obesity after 24 weeks of a dietary intervention based on an energy-reduced anti-inflammatory diet and examined the relationship of these changes with changes in the inflammatory potential of the diet. The anthropometric and body composition parameters of 81 participants (average age of 43 years, 74 women) were assessed. Metabolic status was determined using the glycemic and lipid statuses, and the cardiometabolic index and inflammatory status were determined using the concentration of high-sensitivity C-reactive protein (hs-CRP), interleukin 6 (IL-6), and tumor necrosis factor α (TNF-α). The inflammatory potential of the diet was assessed using the Dietary Inflammatory Index (DII®). Intervention with an anti-inflammatory diet resulted in a significant reduction in body weight and visceral adipose tissue and caused improvements in the participants' cardiometabolic and inflammatory statuses. The anti-inflammatory diet was shown to be effective regarding obesity management. The study data could advance current scientific knowledge in the field of inflammation and diet, provide guidelines for obesity management, and find its application in routine clinical practice.

The effect of antioxidant status on overall survival in renal cell carcinoma.

INTRODUCTION: The oxidative stress contributes to all three phases of carcinogenesis and represents a concomitant condition in renal cell carcinoma (RCC). RCC is the most common type of neoplasm of the kidney, and despite numerous studies the set of predictive and prognostic markers of survival are still unknown. The aim of our study was to examine the relation between antioxidant (AO) status and overall survival (OS) in RCC patients. MATERIAL AND METHODS: Our study included 95 patients with RCC, who underwent radical nephrectomy. We analysed the prognostic role of antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase, glutathione S-transferase, glutathione reductase, glutathione, and malondialdehyde) and other clinicopathological factors (size, grade, stage, and histological subtype) on the OS of RCC patients. RESULTS: The 5-year OS was 54.6%. The survival analysis related to AO parameters showed no significant difference in survival of RCC patients. The concentration of malondialdehyde, an indicator of lipid peroxidation, also had no significant effect on the survival rate of RCC patients. Univariate and multivariate analysis confirmed the significance of clinicopathological parameters (size, p < 0.001; Fuhrman grade, p = 0.001, and stage, p < 0.001) for patients' survival. CONCLUSIONS: In our cohort of patients, different antioxidant parameters were not found to be predictors for OS of patients with RCC, who underwent radical nephrectomy.

Non-Diabetic Kidney Disease in Patients with Type 2 Diabetes Mellitus-11-Year Experience from a Single Center.

INTRODUCTION: In patients with diabetes mellitus (DM), non-diabetic renal disease (NDRD) can also occurs, as well as diabetic nephropathy. NDRD is most accurately diagnosed using kidney biopsy. AIM: The aim of the study was to investigate the incidence and type of NDRD diagnosed by kidney biopsy in patients with type 2 DM and the correlation of clinical and laboratory findings with histopathological diagnosis. MATERIAL AND METHODS: From April 2007 to October 2018, 290 kidney biopsies were performed at the Department of Nephrology, Internal Medicine Clinic in Banja Luka, out of which 18 patients (males 9, mean age 59.8 years) were with type 2 DM. The US-guided (ultrasound device: Toshiba Famio 5) kidney biopsy was performed using an automatic biopsy instrument FAST-GUN® with needle 16G. Kidney tissue samples were analyzed by light microscopy and immunofluorescence. RESULTS: In 18 patients with type 2 DM, the average duration of the disease was 5.9 years, 5 patients had a retinopathy, and 16 patients had hypertension. Biopsy indications were: nephrotic syndrome in 11 patients, asymptomatic urinary abnormalities in 3 patients, and rapid chronic renal failure progression. Unsatisfactory quality sample for pathohistological analysis was obtained in one patient, and out of the other 17, 6 (35.3%) had NDRD, 3 (17.6%) had NDRD superimposed with the diabetic nephropathy, and 8 (47.1%) had diabetic nephropathy. Of the patients who had NDRD, 3 had membranous glomerulonephritis, 1 had focal segmental glomerulosclerosis, and two had hypertensive nephroangiosclerosis. Out of patients with coexisting NDRD and diabetic nephropathy, 2 had hypertensive nephroangiosclerosis and one diabetic nephropathy and lupus nephritis. CONCLUSION: NDRD was diagnosed using kidney biopsy in 9/17 patients with type 2 DM, which confirms the significance of the kidney biopsy in patients with DM with properly indications. Accurate diagnosis provides disease specific treatment and thus significantly improves the long-term prognosis of the patient.

The association between the Mediterranean diet and high physical activity among the working population in Croatia.

BACKGROUND: Unhealthy eating habits and physical inactivity constitute an emerging public health problem. The working population is of special interest for public health monitoring and evaluation because workers' unhealthy lifestyles may lead to reduced work ability. The aim of this study was to determine diet quality and adherence to the Mediterranean diet (MD), according to the level of physical activity, and to detect variables associated with the working population's being highly physically active. MATERIAL AND METHODS: At the Institute for Occupational Medicine 400 full-time workers were examined for obesity factors, filled in the short version of International Physical Activity Questionnaire (IPAQ-short) and a validated food frequency questionnaire for adherence to the Mediterranean diet using Mediterranean Diet Score (MDS). The workers were divided into low, moderate and high physical activity groups according to the IPAQ-short scoring protocol. Hierarchical linear regression was performed to determine the variables associated with being highly active. RESULTS: One-third of the participants were highly physically active and their diet adhered to the MD (MeMDS = 7). Significant variables associated with a high level of physical activity were gender (p < 0.001), age (p = 0.02), waist-to-hip ratio (WHR) (p < 0.001), sitting level (p = 0.044) and occupational type (p < 0.001). CONCLUSIONS: It was found that the participants displaying a high level of physical activity had a better quality diet that adhered to the Mediterranean diet but not to a significant degree. The variables associated with a high level of physical activity were male gender, younger age, normal WHR, non-sedentary occupation and reduced sitting time. The study findings could serve the purpose of improving future public health promotion of physical activity and the Mediterranean diet. Med Pr. 2019;70(2):169-76.

Enhanced Recovery in Surgical Intensive Care: A Review.

Patients are admitted to the surgical intensive care (SICU) unit after emergency and elective surgery. After elective surgery, for further support, or to manage coexisting comorbidities. The implementation of the ERAS (Enhanced recovery after surgery) protocols in surgery should decrease the need for ICU beds, but there will always be unpredicted complications after surgery. These will require individual management. What we can do for our surgical patients in ICU to further enhance their recovery? To promote early enhanced recovery in surgical intensive care-SICU, three areas need to be addressed, sedation, analgesia, and delirium. Tools for measurement and protocols for management in these three areas should be developed to ensure best practice in each SICU. The fourth important area is Nutrition. Preoperative screening and post-operative measurement of the state of nutrition also need to be developed in the SICU. The fifth important area is early mobilization. ERAS protocols encourage early mobilization of the critically ill patients, even if on mechanical ventilation. Early mobilization is possible and should be implemented by special multidisciplinary ICU team. All team members must be familiar with protocols to be able to implement them in their field of expertise. Personal and professional attitudes are critical for implementation. In the core of all our efforts should be the patient and his well-being.

Immunohistochemical expression of proliferative markers in renal cell carcinoma.

PURPOSE: The purpose of this study was to investigate into the expression of cyclin A and telomerase in renal cell carcinoma (RCC) and to analyze the relationship between expression and the clinicopathological characteristics of the tumor and their impact on survival. METHODS: The overall material included 74 samples of RCC and 4 of normal renal tissue. Primary cyclin A antibody from Santa Cruz Biotechnology and TERT MA5-16034 antibody from Thermo Fisher Scientific Inc were used. Staining was performed by streptavidin-biotin technique using DAKO LSAB+ kit. Statistical analyses were performed using of SPSS 23 Statistics software from IBM. RESULTS: No differences in cyclin A and telomerase expression among gender and age groups were found, nor did the tumor dimensions have any significant impact on expression. Also, tumor grades and stages did not differ. However, histological types differed in favor of the papillary type. A significant positive correlation between both markers, as well as between the expression and tumor stage and grade was noticed. Only the tumor stage had negative impact on survival. CONCLUSIONS: Although not affecting survival, the expression of cyclin A and telomerase increased with tumor stage and grade, suggesting that cyclin A and telomerase could be potential proliferative immunohistochemical markers of RCC.

Concomitance of Polymorphisms in Glutathione Transferase Omega Genes Is Associated with Risk of Clear Cell Renal Cell Carcinoma.

Glutathione S-transferases (GSTs), a superfamily of multifunctional enzymes, play an important role in the onset and progression of renal cell carcinoma (RCC). However, novel GST omega class (GSTO), consisting of GSTO1-1 and GSTO2-2 isoenzymes, has not been studied in RCC yet. Two coding single nucleotide polymorphisms (SNPs) supposedly affect their functions: GSTO1*C419A (rs4925) causing alanine to aspartate substitution (*A140D) and GSTO2*A424G (rs156697) causing asparagine to aspartate substitution (*N142D), and have been associated with several neurodegenerative diseases and cancers. Functional relevance of yet another GSTO2 polymorphism, identified at the 5' untranslated (5'UTR) gene region (GSTO2*A183G, rs2297235), has not been clearly discerned so far. Therefore, we aimed to assess the effect of specific GSTO1 and GSTO2 gene variants, independently and in interaction with established risk factors (smoking, obesity and hypertension) on the risk for the most aggressive RCC subtype, the clear cell RCC (ccRCC). Genotyping was performed in 239 ccRCC patients and 350 matched controls, while plasma levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG), a biomarker of oxidative DNA damage, were determined by ELISA. As a result, combined effect of all three variant genotypes exhibited almost 3-fold risk of RCC development. Additionally, this association was confirmed at the haplotype level [variant GSTO1*A/GSTO2*G (rs156697)/GSTO2*G (rs2297235) haplotype], suggesting a potential role of those variants in propensity to RCC. Regarding the gene-environment interactions, variant GSTO2*G (rs156697) homozygous smokers are at higher ccRCC risk. Association in terms of oxidative DNA damage was found for GSTO2 polymorphism in 5'UTR and 8-OHdG. In conclusion, the concomitance of GSTO polymorphisms may influence ccRCC risk.

Explosive muscle power assessment in elite athletes using wingate anaerobic test / Avaliação da força muscular explosiva em atletas de elite usando o teste anaeróbico de wingate / Evaluación de la fuerza muscular explosiva en atletas de élite usando la prueba anaeróbica de wingate

ABSTRACT Introduction: Maximal effort physiological tests provide information about the current functional capacity of athletes. Objective: The aim of this study was to evaluate anaerobic performance parameters in elite athletes and to compare them in terms of the specific demands of each sport. We also created and applied the new software which enables us to quantify a new parameter -explosive muscle power (EP), a major component in sports requiring explosive bursts of movement lasting from a few seconds to 1 or 2 minutes. This new parameter reflects the velocity of energy transformation from intramuscular ATP and high-energy phosphates into mechanical power. Methods: All Wingate test parameters (standard parameters) - anaerobic power (AP), anaerobic capacity (AC), and explosive power (EP) as the new parameter were recorded in 104 subjects: 30 non-athletes and 74 athletes divided into different groups depending on their sport specialty (20 rowers, 28 wrestlers and 26 soccer players). Results: Anaerobic power (AP), anaerobic capacity (AC) and explosive power (EP) were significantly higher in the group of athletes compared to non-athletes. Among athletes, significant differences were observed in some parameters according to the type of activities they are involved in. The highest values were recorded in the group of wrestlers (AP=836W; AC=16.6kJ; EP=139W/s). The values of AP (absolute values) and EP (absolute and relative values) were significantly higher in wrestlers than in soccer players and rowers, but there was no significant difference in AC among these groups. The EP variable had a distribution similar to AP. Conclusions: Alongside anaerobic power and anaerobic capacity, the assessment of explosive power may complement the anaerobic profile of athletes. Experts in the field of sports medicine and exercise physiology could find these results useful in improving test variables, which are more important for specific sports, and for evaluating and monitoring training progress. Level of Evidence I; Diagnostic studies - Investigating a diagnostic test.

RESUMO Introdução: Os testes fisiológicos de esforço máximo fornecem informações sobre a capacidade funcional atual dos atletas. Objetivo: O objetivo deste estudo foi avaliar os parâmetros de desempenho anaeróbico em atletas de elite e compará-los em relação às demandas específicas de cada esporte. Além disso, criamos e aplicamos o novo software que possibilita a quantificação de um novo parâmetro - força muscular explosiva (FE), um componente importante em esportes que requerem explosões de movimento que duram de alguns segundos a 1 ou 2 minutos. Este novo parâmetro reflete a velocidade de transformação de energia a partir de ATP e fosfatos de alta energia intramusculares em potência mecânica. Métodos: Todos os parâmetros de teste de Wingate (parâmetros padrão) - potência anaeróbica (PA), capacidade anaeróbica (CA) e força explosiva (FE) como um novo parâmetro foram registrados em 104 indivíduos: 30 não atletas e 74 atletas divididos em diferentes grupos, dependendo da sua especialidade esportiva (20 remadores, 28 lutadores e 26 jogadores de futebol). Resultados: A potência anaeróbica (PA), a capacidade anaeróbica (CA) e a força explosiva (FE) foram significativamente maiores no grupo de atletas em comparação com não atletas. Entre os atletas, diferenças significativas foram observadas em alguns parâmetros, de acordo com o tipo de atividades nas quais eles estão envolvidos. Os valores mais altos foram registrados no grupo de lutadores (PA = 836 W, CA = 16,6 kJ, FE = 139 W/s). Os valores de PA (valores absolutos) e FE (valores absolutos e relativos) foram significativamente maiores em lutadores do que em jogadores de futebol e remadores, mas não houve diferenças significativas na CA entre esses grupos. A variável FE mostrou uma distribuição similar à da PA. Conclusões: Juntamente com o poder anaeróbico e a capacidade anaeróbica, a avaliação da força explosiva pode complementar o perfil anaeróbico dos atletas. Especialistas no campo da medicina esportiva e da fisiologia do exercício poderiam achar esses resultados úteis para melhorar as variáveis de teste que são mais importantes para esportes específicos e para avaliar e monitorar o progresso do treinamento. Nível de Evidência I; Estudo diagnóstico - Investigação de um exame para diagnóstico.

RESUMEN Introducción: Las pruebas fisiológicas de esfuerzo máximo proporcionan información sobre la capacidad funcional actual de los atletas. Objetivo: El objetivo de este estudio fue evaluar los parámetros de rendimiento anaeróbico en atletas de élite y compararlos en relación con las demandas específicas de cada deporte. Además, creamos y aplicamos el nuevo software que permite la cuantificación de un nuevo parámetro - fuerza muscular explosiva (FE), un componente grande en deportes que requieren explosiones de movimiento que duran de unos segundos a 1 o 2 minutos. Este nuevo parámetro refleja la velocidad de transformación de energía a partir de ATP y fosfatos de alta energía intramusculares en potencia mecánica. Métodos: Todos los parámetros de prueba de Wingate (parámetros estándar) - potencia anaeróbica (PA), capacidad anaeróbica (CA) y fuerza explosiva (FE) como un nuevo parámetro fueron registrados en 104 sujetos: 30 no atletas y 74 atletas divididos en diferentes grupos dependiendo de su especialidad deportiva (20 remeros, 28 luchadores y 26 jugadores de fútbol). Resultados: La potencia anaeróbica (PA), la capacidad anaeróbica (CA) y la fuerza explosiva (FE) fueron significativamente mayores en el grupo de atletas en comparación con los no atletas. Entre los atletas, se observaron diferencias significativas en algunos parámetros según el tipo de actividades en las que están involucrados. Los valores más altos fueron registrados en el grupo de luchadores (PA =836 W; CA = 16,6 kJ; FE=139 W/s). Los valores de PA (valores absolutos) y FE (valores absolutos y relativos) fueron significativamente mayores en luchadores que en jugadores de fútbol y remeros, pero no hubo diferencias significativas en CA entre estos grupos. La variable FE mostró una distribución similar a la de la PA. Conclusiones: Junto con la potencia anaeróbica y la capacidad anaeróbica, la evaluación de la fuerza explosiva puede complementar el perfil anaeróbico de los atletas. Los expertos en el campo de la medicina deportiva y la fisiología del ejercicio podrían encontrar estos resultados útiles para mejorar las variables de prueba que son más importantes para deportes específicos y para evaluar y monitorear el progreso del entrenamiento. Nivel de Evidencia I; Estudio diagnóstico - Investigación de un examen para diagnóstico.

HTERT promoter methylation and single nucleotide polymorphism (-245 T>C) affect renal cell carcinoma behavior in Serbian population.

PURPOSE: Renal cell carcinoma (RCC) is the most common renal cancer in adults and includes several subtypes that may be distinguished by their histology, genetic background, clinical course and responses to treatment. Human telomerase reverse transcriptase (hTERT), a crucial enzyme for telomere maintenance, has been linked to RCC development. The purpose of this study was to search for genetic and epigenetic alterations in hTERT (promoter mutations and methylation and gene amplification), and to establish a possible association between molecular and clinico-pathological characteristics of RCC. METHODS: DNA was extracted from 31 formalin-fixed, paraffin-embedded tumor samples and 23 blood samples from 54 patients with RCC. Polymerase chain reaction (PCR) products were sequenced and analyzed using the Sequencher software. HTERT amplification was determined by quantitative PCR, while the promoter methylation status was assessed by methylation specific PCR. Statistical analysis was performed using SPSS. RESULTS: No mutations could be detected in the hTERT promoter but only a single nucleotide polymorphism (SNP) (-245 T>C). In 54 analyzed RCC cases, the variant allele C was present in homozygous or heterozygous form in 48% of the patients. The C allele was significantly more frequent in low grade tumors (p=0.046). Gene amplification was detected in 19.4% of the 31 RCCs and hTERT methylation in 54.8% of the 31 samples. An association was established between methylation and histological type of RCC (p=0.047). CONCLUSIONS: HTERT seems to be implicated in RCC pathogenesis since the promoter polymorphism exerts a modulation effect on tumor behavior. In addition, hTERT promoter methylation status is related to RCC histology.

Genome-wide association study identifies multiple risk loci for renal cell carcinoma.

Previous genome-wide association studies (GWAS) have identified six risk loci for renal cell carcinoma (RCC). We conducted a meta-analysis of two new scans of 5,198 cases and 7,331 controls together with four existing scans, totalling 10,784 cases and 20,406 controls of European ancestry. Twenty-four loci were tested in an additional 3,182 cases and 6,301 controls. We confirm the six known RCC risk loci and identify seven new loci at 1p32.3 (rs4381241, P=3.1 × 10-10), 3p22.1 (rs67311347, P=2.5 × 10-8), 3q26.2 (rs10936602, P=8.8 × 10-9), 8p21.3 (rs2241261, P=5.8 × 10-9), 10q24.33-q25.1 (rs11813268, P=3.9 × 10-8), 11q22.3 (rs74911261, P=2.1 × 10-10) and 14q24.2 (rs4903064, P=2.2 × 10-24). Expression quantitative trait analyses suggest plausible candidate genes at these regions that may contribute to RCC susceptibility.

Polymorphisms of survivin -31 G/C gene are associated with risk of urothelial carcinoma in Serbian population.

PURPOSE: Survivin is thought to play an important role in carcinogenesis and is found to be associated with poor clinical outcome in various malignancies. Gene -31 G/C polymorphism has been identified as a risk factor for the development of several types of tumors. The purpose of this study was to investigate the association between survivin gene promoter -31C/G polymorphism and urothelial carcinoma (UC) risk in Serbian population and to compare the different expressions of survivin in UC of different disease stages, histological grades and tumor location in the upper or lower urinary tract. METHODS: DNA from 94 patients with primary UC and from 82 healthy subjects was subjected to PCR restriction fragment length polymorphism analysis (PCR-RFLP) to identify individual genotypes. UC samples were subjected to immunohistochemical analysis to assess survivin expression in these lesions. RESULTS: It was observed that the frequency of G/G genotype was greater in patients with UC (58.7%) than in controls (32%). Compared with study subjects carrying the C/G or C/C genotypes, significantly increased UC risk was found for individuals carrying the G/G genotype. Those carrying the G/G genotype had a significantly increased UC risk compared with those with C/G or C/C genotypes. Patients with UC carrying the G/G genotype had a greater prevalence of muscle-invading (stage T2-T4), high-grade (G2) tumor and immunohistochemicaly overexpressed survivin compared with those carrying the C/G or C/C genotypes. CONCLUSIONS: G/G genotype of the -31C/G polymorphism might be a risk factor for UC development.

GSTM1 genotype is an independent prognostic factor in clear cell renal cell carcinoma.

PURPOSE: Owing to dual functionality of cytosolic glutathione S-transferases (GSTs), they might affect both the development and the progression of renal cell carcinoma (RCC). However, the data on the prognostic value of GST polymorphism in patients with RCC are scarce. Hence, we evaluated the effect of GST gene variants on both the risk of RCC development and the postoperative prognosis in patients with clear cell RCC (ccRCC). METHODS: GST genotypes were determined in 305 patients with RCC and 326 matched controls, whereas the overall survival was evaluated in patients with ccRCC only. The presence of GSTM1:ASK1 protein-protein interaction in ccRCC tissue samples was analyzed by methods of immunoprecipitation and immunoblot. RESULTS: We noted an increased risk of RCC development in carriers of GSTM1-null and GSTP1-variant genotype (P<0.05). On the contrary, survival analysis indicated shorter overall survival for patients with ccRCC with GSTM1-active genotype (P = 0.026). Furthermore, patients with ccRCC with GSTM1-active genotype had significantly higher hazard ratio (P<0.05), in analyzed regression models, compared with the carriers of GSTM1-null genotype. Finally, the presence of GSTM1:ASK1 protein-protein interaction was found in all RCC tissue samples studied. CONCLUSIONS: Carriers of GSTM1-null and GSTP1-variant genotypes are in increased risk of RCC development. On the contrary, GSTM1-null genotype is associated with favorable postoperative prognosis in ccRCC. The possible molecular mechanism underlying the role of GSTM1 protein in RCC progression might be the presence of GSTM1:ASK1 protein-protein interaction. Hence, determination of GSTM1-genotype might serve as a valuable indicator in both RCC risk assessment and postoperative prognosis.

Clinicopathological characteristics and survival in Serbian patients with renal cell carcinoma: a retrospective analysis.

PURPOSE: Indications of kidney cancer outcome in lowerincome countries are based on an incidence/mortality ratio due to lack of survival information. This study was conducted to provide outcome data in Serbian patients with renal cell carcinoma (RCC) and to identify prognostic factors that could affect their overall survival (OS). METHODS: This retrospective study included 185 patients who underwent nephrectomy. We assessed certain clinicopathological data including age, gender, tumor size, grade, stage and histological subtypes for their possible impact on OS. RESULTS: The 5-year OS was 63.2%. Significant association was found between OS and age (log-rank 12.455, p=0.006), tumor size (log-rank 26.425, p=0.000), grade (log-rank 13.249, p=0.000) and stage (log-rank 43.235, p=0.000). Univariate analysis indicated size (p=0.000), grade (p=0.001) and stage (p=0.000) as prognostic factors for OS. In multivariate analysis, grade (p=0.014) and stage (p=0.000) remained significant predictors of OS. CONCLUSION: Tumor grade and stage were identified as independent prognostic factors of OS survival in Serbian patients with RCC.

Combined GSTM1-Null, GSTT1-Active, GSTA1 Low-Activity and GSTP1-Variant Genotype Is Associated with Increased Risk of Clear Cell Renal Cell Carcinoma.

The aim of this study was to evaluate specific glutathione S-transferase (GST) gene variants as determinants of risk in patients with clear cell renal cell carcinoma (cRCC), independently or simultaneously with established RCC risk factors, as well as to discern whether phenotype changes reflect genotype-associated risk. GSTA1, GSTM1, GSTP1 and GSTT1 genotypes were determined in 199 cRCC patients and 274 matched controls. Benzo(a)pyrene diolepoxide (BPDE)-DNA adducts were determined in DNA samples obtained from cRCC patients by ELISA method. Significant association between GST genotype and risk of cRCC development was found for the GSTM1-null and GSTP1-variant genotype (p = 0.02 and p<0.001, respectively). Furthermore, 22% of all recruited cRCC patients were carriers of combined GSTM1-null, GSTT1-active, GSTA1-low activity and GSTP1-variant genotype, exhibiting 9.32-fold elevated cRCC risk compared to the reference genotype combination (p = 0.04). Significant association between GST genotype and cRCC risk in smokers was found only for the GSTP1 genotype, while GSTM1-null/GSTP1-variant/GSTA1 low-activity genotype combination was present in 94% of smokers with cRCC, increasing the risk of cRCC up to 7.57 (p = 0.02). Furthermore, cRCC smokers with GSTM1-null genotype had significantly higher concentration of BPDE-DNA adducts in comparison with GSTM1-active cRCC smokers (p = 0.05). GSTM1, GSTT1, GSTA1 and GSTP1 polymorphisms might be associated with the risk of cRCC, with special emphasis on GSTM1-null and GSTP1-variant genotypes. Combined GSTM1-null, GSTT1-active, GSTA1 low activity and GSTP1-variant genotypes might be considered as "risk-carrying genotype combination" in cRCC.