Analysis of the Level of Adiponectin and Selected Cytokines in Patients with Knee Osteoarthritis.

Background and Objectives: Knee osteoarthritis (KOA) is a degenerative disease that is continuously targeting people of different ages, but especially the elderly population, the number of which tends to increase continuously at the global level. Apart from age, excess weight can influence the evolution of the disease, with obesity being associated with a weak inflammation stage and an imbalance between pro-inflammatory and anti-inflammatory cytokines. The present work aimed to analyze specific biomarkers, namely ACRP-30, IL-10, TNF-α, and IL-6, in knee synovial fluid, and correlate them with KOA patients' clinical data, radiographic changes, and functional and pain scores. Materials and Methods: 24 subjects with KOA and over 50 years of age participate in the present study. Synovial fluid was harvested using ultrasound guidance from the target knees of the enrolled KOA patients, and the levels of ACRP-30, IL-10, TNF-α, and IL-6 were measured using enzyme-linked immunosorbent assays (ELISA). All patients underwent a supine X-ray at the target knee and were classified using Kellgren-Lawrence (K-L) grading. The Western Ontario and McMaster University Osteoarthritis Index (WOMAC) was used to assess self-reported physical function, pain, and stiffness. Results: The obtained results highlighted a significant correlation between age and adiponectin level (p = 0.0451, r = -0.412). Also, the IL-10 values are lower in cases where the intensity of the pain is more pronounced (p = 0.0405, r = -0.421). In addition, analyzing the data by gender, it was observed that in the case of males, stiffness is more related to age (p = 0.0079, r = 0.7993), compared to women (p = 0.0203, r = 0.6223). In the case of women, the progression of the disease tends to increase more intensively the WOMAC score's total values (p = 0.00031, r = 0.8342), compared with men (p = 0.0289, r = 7013). Regarding interleukins and BMI, significant correlations were observed only in the case of men. Conclusions: A significant correlation between age and adiponectin, and adiponectin and IL-6, suggests that advanced age may contribute to adiponectin reduction. Comparing men with women, it was observed that men's age is more related to rigidity, and IL-6 and IL-10 are directly correlated to BMI; in addition, women seem to be more sensitive to pain and stiffness.

Potential Diagnostic Biomarker Detection for Prostate Cancer Using Untargeted and Targeted Metabolomic Profiling.

Prostate cancer (PCa) remains one of the leading causes of cancer mortality in men worldwide, currently lacking specific, early detection and staging biomarkers. In this regard, modern research focuses efforts on the discovery of novel molecules that could represent potential future non-invasive biomarkers for the diagnosis of PCa, as well as therapeutic targets. Mounting evidence shows that cancer cells express an altered metabolism in their early stages, making metabolomics a promising tool for the discovery of altered pathways and potential biomarker molecules. In this study, we first performed untargeted metabolomic profiling on 48 PCa plasma samples and 23 healthy controls using ultra-high-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-[ESI+]-MS) for the discovery of metabolites with altered profiles. Secondly, we selected five molecules (L-proline, L-tryptophan, acetylcarnitine, lysophosphatidylcholine C18:2 and spermine) for the downstream targeted metabolomics and found out that all the molecules, regardless of the PCa stage, were decreased in the PCa plasma samples when compared to the controls, making them potential biomarkers for PCa detection. Moreover, spermine, acetylcarnitine and L-tryptophan had very high diagnostic accuracy, with AUC values of 0.992, 0.923 and 0.981, respectively. Consistent with other literature findings, these altered metabolites could represent future specific and non-invasive candidate biomarkers for PCa detection, which opens novel horizons in the field of metabolomics.

Efficiency of Platelet Transfusion in Patients with Moderate-to-Severe Chronic Kidney Disease and Thrombocytopenia.

There have been relatively few studies revealing a decreased platelet count in chronic kidney disease (CKD). Although this hematological abnormality is not as well documented as renal anemia, platelet functions are altered in the uremic environment and there is an increased risk of bleeding. The aim of this study was to assess the effectiveness of the administration of platelet concentrate in CKD based on how patient prognosis was influenced by platelet transfusion therapy. The study monitored 104 patients with CKD and thrombocytopenia who received platelet transfusion during their hospitalization in the period from 2015 to 2021. The complete blood cell count, serum urea and creatinine, and inflammatory status were tested upon admission. The number of transfused platelet units were considered for each patient. A Kruskal-Wallis H test showed that for one transfused platelet unit, the distribution of the number of platelets (×103/µL) was the same across the categories of associated diagnoses, which was seen as possible risk factors for thrombocytopenia, including liver cirrhosis and urosepsis. With a single exception, all patients exceeded the critical threshold of 20 × 103/µL and 14 patients remained under 50 × 103/µL. Even though our patients exceeded the critical threshold of platelet numbers, in patients with multiple comorbidities, severe, uncontrolled hemorrhages could not be prevented in 4.83% of cases.

COVID-19 Biomarkers Comparison: Children, Adults and Elders.

Background and Objectives: this study aimed to research links between C-reactive protein (CRP), lactate dehydrogenase (LDH), creatinekinase (CK), 25-OH vitamin D (25-OHD), ferritin (FER), high-density lipoprotein cholesterol (HDL)cholesterol and clinical severity in patients from the western part of Romania, and compare their potential use as biomarkers for intensive care units (ICU) admission and death in children, adults and elders. Materials and Methods: this study is a retrospective cohort study, performed on patients positively diagnosed with COVID-19. Available CRP, LDH, CK 25-OH vitamin D, ferritin, HDL cholesterol and clinical severity were recorded. The following were assessed: median group differences, association, correlation and receiver operating characteristic. Results: 381 children, 614 adults and 381 elders were studied between 1 March 2021 and 1 March 2022. Most children and adults presented mild symptomatology (53.28%, 35.02%, respectively), while most elders presented severe symptomatology (30.04%). ICU admission was 3.67% for children, 13.19% for adults and 46.09% for elders, while mortality was 0.79% for children, 8.63% for adults and 25.1% for elders. With the exception of CK, all other biomarkers showed some significant associations with clinical severity, ICU admission and death. Conclusions: CRP, LDH, 25-OH vitamin D, ferritin and HDL are important biomarkers for COVID-19 positive patients, especially in the pediatric population, while CK was mostly within normal ranges.

Linking Pregnancy and Long-Term Health: The Impact of Cardiovascular Risk on Telomere Shortening in Pregnant Women.

Background and Objectives: Telomeres are repetitive DNA sequences located at the end of chromosomes that play a crucial role in maintaining chromosomal stability. Shortening of telomeres has been associated with an increased risk of cardiovascular disease. The aim of this study was to investigate whether the length of telomeres in pregnant women with cardiovascular risk is shorter compared to those without cardiovascular risk. Materials and Methods: A total of 68 participants were enrolled, including 30 pregnant women with cardiovascular risk and 38 without cardiovascular risk, who were followed-up during their pregnancy between 2020 and 2022 at the Obstetrical and Gynecology Department of the "Pius Brînzeu" Emergency County Clinical Hospital in Timisoara, Romania. All included women underwent delivery via cesarean section at the same medical institution. The telomere length was measured in each participant using quantitative Polymerase chain reaction (PCR). Results: The results showed that the telomere length was negatively correlated with cardiovascular risk in pregnant women, with significantly shorter telomeres observed in the cardiovascular risk group (mean telomere length = 0.3537) compared to the group without cardiovascular risk (mean telomere length = 0.5728) (p = 0.0458). Conclusions: These findings suggest that cardiovascular risk during pregnancy may be associated with accelerated telomere shortening, which could have implications for the long-term health of both the mother and the child. Further research is needed to investigate the potential mechanisms underlying this association and to identify interventions that may mitigate the negative effects of cardiovascular risk on the telomere length during pregnancy.

Gene Network Analysis of the Transcriptome Impact of SARS-CoV-2 Interacting MicroRNAs in COVID-19 Disease.

According to the World Health Organization (WHO), as of June 2022, over 536 million confirmed COVID-19 disease cases and over 6.3 million deaths had been globally reported. COVID-19 is a multiorgan disease involving multiple intricated pathological mechanisms translated into clinical, biochemical, and molecular changes, including microRNAs. MicroRNAs are essential post-transcriptional regulators of gene expression, being involved in the modulation of most biological processes. In this study, we characterized the biological impact of SARS-CoV-2 interacting microRNAs differentially expressed in COVID-19 disease by analyzing their impact on five distinct tissue transcriptomes. To this end, we identified the microRNAs' predicted targets within the list of differentially expressed genes (DEGs) in tissues affected by high loads of SARS-CoV-2 virus. Next, we submitted the tissue-specific lists of the predicted microRNA-targeted DEGs to gene network functional enrichment analysis. Our data show that the upregulated microRNAs control processes such as mitochondrial respiration and cytokine and cell surface receptor signaling pathways in the heart, lymph node, and kidneys. In contrast, downregulated microRNAs are primarily involved in processes related to the mitotic cell cycle in the heart, lung, and kidneys. Our study provides the first exploratory, systematic look into the biological impact of the microRNAs associated with COVID-19, providing a new perspective for understanding its multiorgan physiopathology.

Diagnostic Value of microRNA-375 as Future Biomarker for Prostate Cancer Detection: A Meta-Analysis.

Background and Objectives: Responding to the need for additional biomarkers for the diagnosis of prostate cancer (PCa), mounting studies show that microRNAs (miRNAs/miRs) possess great potential as future promising diagnostic tools. However, the usefulness of these miRNAs is still highly debated, as the degree of inconsistency between study designs and results is still elevated. Herein, we present a meta-analysis evaluating the diagnostic value and accuracy of circulating miR-375, as it is one of the most studied types of miRs in PCa. Materials and Methods: The diagnostic accuracy of miR-375 was evaluated using the QUADAS-2 tool, analyzing different statistical parameters. The seven studies (from six articles) that matched our selection included 422 PCa patients and 212 controls (70 healthy volunteers + 142 with benign prostate diseases). Results and Conclusion: We obtained a p-value of 0.76 for sensitivity, 0.83 for specificity, 16 for DOR, 4.6 for LR+, 0.29 for LR-, and 0.87 for AUC (95% CI 0.83-0.89). Our results confirm that miRNA-375 has high diagnostic potential for PCa, suggesting its usefulness as a powerful biomarker. More comprehensive studies are warranted to better assess its true value as a diagnostic biomarker for this urologic disease.

Identification of Genomic Variants of SARS-CoV-2 Using Nanopore Sequencing.

Background and Objectives: SARS-CoV-2 is the first global threat and life-changing event of the twenty-first century. Although efficient treatments and vaccines have been developed, due to the virus's ability to mutate in key regions of the genome, whole viral genome sequencing is needed for efficient monitoring, evaluation of the spread, and even the adjustment of the molecular diagnostic assays. Materials and Methods: In this study, Nanopore and Ion Torrent sequencing technologies were used to detect the main SARS-CoV-2 circulating strains in Timis County, Romania, between February 2021 and May 2022. Results: We identified 22 virus lineages belonging to seven clades: 20A, 20I (Alpha, V1), 21B (Kappa), 21I (Delta), 21J (Delta), 21K (Omicron), and 21L (Omicron). Conclusions: Results obtained with both methods are comparable, and we confirm the utility of Nanopore sequencing in large-scale epidemiological surveillance due to the lower cost and reduced time for library preparation.

Plasma hsa-mir-19b is a potential LevoDopa therapy marker.

Parkinson's disease (PD) is the second most common neurodegenerative disorder among the elderly, the diagnostic and prognostic of which is based mostly on clinical signs. LevoDopa replacement is the gold standard therapy for PD, as it ameliorates the motor symptoms. However, it does not affect the progression of the disease and its long-term use triggers severe complications. There are no bona fide biomarkers for monitoring the patients' response to LevoDopa and predicting the efficacy of levodopa treatment. Here, we have combined qPCR microRNA array screening with analysis of validated miRs in naïve versus Levodopa-treated PD patients. We have identified plasma miR-19b as a possible biomarker for LevoDopa therapy and validated this result in human differentiated dopaminergic neurons exposed to LevoDopa. In silico analysis suggests that the LevoDopa-induced miR-19b regulates ubiquitin-mediated proteolysis.

Cognitive dysfunction in young subjects with periodontal disease.

BACKGROUND: Periodontal disease is an inflammatory, dysbiotic condition. Studies have shown that in the elderly, periodontal disease was associated with cognitive dysfunction and Alzheimer's disease. OBJECTIVE: To investigate whether young healthy subjects with periodontal disease have lower cognition compared to those without periodontal disease. The salivary cytokines (IL-1ß, TNF-α) levels in relation to cognition were also tested. METHODS: In a monocenter, cross-sectional study, forty subjects [mean age (SD) = 34 (5) and 48% female] from western Romania were classified into periodontal disease conditions using radiographic assessment: 10 subjects had aggressive periodontitis (AGG_P), 20 chronic mild-moderate periodontitis (CR_P), and 10 no periodontitis (NL_P). Neuropsychological assessment performed by standardized neurologists and psychologist included Rey Auditory Verbal Learning Test (RAVLT), Montreal Cognitive Assessment test (MOCA), Mini-Mental State Examination (MMSE), and Prague tests. Salivary cytokines levels were determined by ELISA. RESULTS: RAVLT and MOCA delayed recall scores were lower in AGG_P group compared to NL_P and CR_P. The learning curve was also different with subjects with AGG_P showing reduced learning performance. Contrary to our hypothesis, salivary IL-1ß associated with immediate but not delayed cognitive scores. CONCLUSIONS: These results showed for the first time that subjects with AGG_P had cognitive dysfunction and IL-1ß may play a role in this process.

Long Non-Coding RNA Expression in Laser Micro-Dissected Luminal A and Triple Negative Breast Cancer Tissue Samples-A Pilot Study.

Background and Objectives: Breast cancer (BC) remains one of the major causes of cancer death in women worldwide. The difficulties in assessing the deep molecular mechanisms involved in this pathology arise from its high complexity and diverse tissue subtypes. Long non-coding RNAs (lncRNAs) were shown to have great tissue specificity, being differentially expressed within the BC tissue subtypes. Materials and Methods: Herein, we performed lncRNA profiling by PCR array in triple negative breast cancer (TNBC) and luminal A tissue samples from 18 BC patients (nine TNBC and nine luminal A), followed by individual validation in BC tissue and cell lines. Tissue samples were previously archived in formalin-fixed paraffin-embedded (FFPE) samples, and the areas of interest were dissected using laser capture microdissection (LCM) technology. Results: Two lncRNAs (OTX2-AS1 and SOX2OT) were differentially expressed in the profiling analysis (fold change of 205.22 and 0.02, respectively, p < 0.05 in both cases); however, they did not reach statistical significance in the individual validation measurement (p > 0.05) when analyzed with specific individual assays. In addition, GAS5 and NEAT1 lncRNAs were individually assessed as they were previously described in the literature as being associated with BC. GAS5 was significantly downregulated in both TNBC tissues and cell lines compared to luminal A samples, while NEAT1 was significantly downregulated only in TNBC cells vs. luminal A. Conclusions: Therefore, we identified GAS5 lncRNA as having a differential expression in TNBC tissues and cells compared to luminal A, with possible implications in the molecular mechanisms of the TNBC subtype. This proof of principle study also suggests that LCM could be a useful technique for limiting the sample heterogeneity for lncRNA gene expression analysis in BC FFPE tissues. Future studies of larger cohort sizes are needed in order to assess the biomarker potential of lncRNA GAS5 in BC.

Clinical Relevance of Plasma Concentrations of MBL in Accordance with IgE Levels in Children Diagnosed with Bronchial Asthma.

Background and objectives: Bronchial asthma is a heterogeneous, multifactorial pulmonary disease characterized by variable airway obstruction caused by chronic inflammation. Our study investigates the clinical relevance of MBL plasma levels in accordance with IgE values in children who attended a pediatric consult for respiratory symptoms with bronchial asthma. Materials and Methods: The study population consists of patients <18-years-old and included 43 patients with bronchial asthma and 64 age-matched healthy subjects as a control group. We used the ELISA Human MBL Immunoassay kit and the electrochemiluminescence immunoassay (ECLIA) kit for IgE determination. Results: Our results show significantly different distributions of patients in the bronchial asthma group and control group. The measured values were within the normal range for most controls, while the bronchial asthma patients displayed higher values of plasma MBL and IgE levels. We observed a wider heterogeneity in MBL concentrations in bronchial asthma patients when compared to the healthy age-matched controls. Our results also suggest a potential clinical usefulness of plasma MBL concentrations in accordance with IgE and eosinophil cells levels in the diagnosis of bronchial asthma, and our results may suggest a prognostic role of MBL in the evolution of asthmatic disease; however, further studies are necessary to confirm these findings. Conclusions:We can say that plasma MBL concentrations present a relative diagnostic role for bronchial asthma in pediatric patients and may suggest a more severe disease progression; however, further studies are needed to elucidate the role played by MBL in the determination and evolution of this disease.

MicroRNAs mediate liver transcriptome changes upon soy diet intervention in mice.

Soy-based diets have triggered the interest of the research community due to their beneficial effects on a wide variety of pathologies like breast and prostate cancer, diabetes and atherosclerosis. However, the molecular details underlying these effects are far from being completely understood; several recent attempts have been made to elucidate the soy-induced liver transcriptome changes in different animal models. Here we used Next Generation Sequencing to identify a set of microRNAs specifically modulated by short-term soy-enriched diet in young male mice and estimated their impact on the liver transcriptome assessed by microarray. Clustering and topological community detection (CTCD) network analysis of STRING generated interactions of transcriptome data led to the identification of four topological communities of genes characteristically altered and putatively targeted by microRNAs upon soy diet intervention.

Active and secondhand smoke exposure throughout life and DNA methylation in breast tumors.

PURPOSE: Tobacco smoke exposure has been associated with altered DNA methylation. However, there is a paucity of information regarding tobacco smoke exposure and DNA methylation of breast tumors. METHODS: We conducted a case-only analysis using breast tumor tissue from 493 postmenopausal and 225 premenopausal cases in the Western New York Exposures and Breast Cancer (WEB) study. Methylation of nine genes (SFN, SCGB3A1, RARB, GSTP1, CDKN2A, CCND2, BRCA1, FHIT, and SYK) was measured with pyrosequencing. Participants reported their secondhand smoke (SHS) and active smoking exposure for seven time periods. Unconditional logistic regression was used to estimate odds ratios (OR) of having methylation higher than the median. RESULTS: SHS exposure was associated with tumor DNA methylation among postmenopausal but not premenopausal women. Active smoking at certain ages was associated with increased methylation of GSTP1, FHIT, and CDKN2A and decreased methylation of SCGB3A1 and BRCA1 among both pre- and postmenopausal women. CONCLUSION: Exposure to tobacco smoke may contribute to breast carcinogenesis via alterations in DNA methylation. Further studies in a larger panel of genes are warranted.

Mature miR-99a Upregulation in the Amniotic Fluid Samples from Female Fetus Down Syndrome Pregnancies: A Pilot Study.

Background and Objective: Although Down syndrome is the most frequent aneuploidy, its pathogenic molecular mechanisms are not yet fully understood. The aim of our study is to quantify-by qRT-PCR-the expression levels of both the mature forms and the pri-miRNAs of the microRNAs resident on chromosome 21 (miR(21)) in the amniotic fluid samples from Down syndrome singleton pregnancies and to estimate the impact of the differentially expressed microRNAs on Down syndrome fetal heart and amniocytes transcriptomes. Materials and methods: We collected amniotic fluid samples harvested by trained obstetricians as part of the second trimester screening/diagnostic procedure for aneuploidies to assess the trisomy 21 status by QF-PCR and karyotyping. Next, we evaluated-by Taqman qRT-PCR-the expression levels of both the mature forms and the pri-miRNA precursors of the microRNAs resident on chromosome 21 in amniotic fluid samples from singleton Down syndrome and euploid pregnancies. Further, we combined miRWalk 3.0 microRNA target prediction with GEO DataSets analysis to estimate the impact of hsa-miR-99a abnormal expression on Down syndrome heart and amniocytes transcriptome. Results: We found a statistically significant up-regulation of the mature form of miR-99a, but not pri-miR-99a, in the amniotic fluid samples from Down syndrome pregnancies with female fetuses. GATHER functional enrichment analysis of miRWalk3.0-predicted targets from Down syndrome amniocytes and fetal hearts transcriptome GEODataSets outlined both focal adhesion and cytokine-cytokine receptor interaction signaling as novel signaling pathways impacted by miR-99a and associated with cardiac defects in female Down syndrome patients. Conclusions: The significant overexpression of miR-99a, but not pri-miR-99a, points towards an alteration of the post-transcriptional mechanisms of hsa-miR-99a maturation and/or stability in the female trisomic milieu, with a potential impact on signaling pathways important for proper development of the heart.

Biomarker Potential of Plasma MicroRNA-150-5p in Prostate Cancer.

Background and Objectives: Over decades, prostate cancer (PCa) has become one of the leading causes of cancer mortality in men. Extensive evidence exists that microRNAs (miRNAs or miRs) are key players in PCa and a new class of non-invasive cancer biomarkers. Materials and Methods: We performed miRNA profiling in plasma and tissues of PCa patients and attempted the validation of candidate individual miRs as biomarkers. Results: The comparison of tissue and plasma profiling results revealed five commonly dysregulated miRs, namely, miR-130a-3p, miR-145-5p, miR-148a-3p, miR-150-5p, and miR-365a-3p, of which only three show concordant changes-miR-130a-3p and miR-150-5p were downregulated and miR-148a-3p was upregulated in both tissue and plasma samples, respectively. MiR-150-5p was validated as significantly downregulated in both plasma and tissue cancer samples, with a fold change of -2.697 (p < 0.001), and -1.693 (p = 0.035), respectively. ROC analysis showed an area under the curve (AUC) of 0.817 (95% CI: 0.680-0.995) for plasma samples and 0.809 (95% CI: 0.616-1.001) for tissue samples. Conclusions: We provide data indicating that miR-150-5p plasma variations in PCa patients are associated with concordant changes in prostate cancer tissues; however, given the heterogeneous nature of previous findings of miR-150-5p expression in PCa cells, additional future studies of a larger sample size are warranted in order to confirm the biomarker potential and role of miRNA-150-5p in PCa biology.

Lifetime exposure to ambient air pollution and methylation of tumor suppressor genes in breast tumors.

BACKGROUND: We previously reported increased risk of breast cancer associated with early life exposure to two measures of air pollution exposure, total suspended particulates (TSP) and traffic emissions (TE), possible proxies for exposure to polycyclic aromatic hydrocarbons (PAHs). Exposure to PAHs has been shown to be associated with aberrant patterns of DNA methylation in peripheral blood of healthy individuals. Exposure to PAHs and methylation in breast tumor tissue has received little attention. We examined the association of early life exposure to TSP and TE with patterns of DNA methylation in breast tumors. METHODS: We conducted a study of women enrolled in the Western New York Exposures and Breast Cancer (WEB) Study. Methylation of nine genes (SFN, SCGB3A1, RARB, GSTP1, CDKN2A CCND2, BRCA1, FHIT, and SYK) was assessed using bisulfite-based pyrosequencing. TSP exposure at each woman's home address at birth, menarche, and when she had her first child was estimated. TE exposure was modeled for each woman's residence at menarche, her first birth, and twenty and ten years prior to diagnosis. Unconditional logistic regression was employed to estimate odds ratios (OR) of having methylation greater than the median value, adjusting for age, secondhand smoke exposure before age 20, current smoking status, and estrogen receptor status. RESULTS: Exposure to higher TSP at a woman's first birth was associated with lower methylation of SCGB3A1 (OR = 0.48, 95% CI: 0.23-0.99) and higher methylation of SYK (OR = 1.86, 95% CI: 1.03-3.35). TE at menarche was associated with increased methylation of SYK (OR = 2.37, 95% CI: 1.05-5.33). TE at first birth and ten years prior to diagnosis was associated with decreased methylation of CCND2 (OR ten years prior to diagnosis=0.48, 95% CI: 0.26-0.89). Although these associations were nominally significant, none were significant after adjustment for multiple comparisons (p < 0.01). CONCLUSIONS: We observed suggestive evidence that exposure to ambient air pollution throughout life, measured as TSP and TE, may be associated with DNA methylation of some tumor suppressor genes in breast tumor tissue. Future studies with a larger sample size that assess methylation of more sites are warranted.

Metabolomic profiles of current cigarette smokers.

Smoking-related biomarkers for lung cancer and other diseases are needed to enhance early detection strategies and to provide a science base for tobacco product regulation. An untargeted metabolomics approach by ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry (UHPLC-Q-TOF MS) totaling 957 assays was used in a novel experimental design where 105 current smokers smoked two cigarettes 1 h apart. Blood was collected immediately before and after each cigarette allowing for within-subject replication. Dynamic changes of the metabolomic profiles from smokers' four blood samples were observed and biomarkers affected by cigarette smoking were identified. Thirty-one metabolites were definitively shown to be affected by acute effect of cigarette smoking, uniquely including menthol-glucuronide, the reduction of glutamate, oleamide, and 13 glycerophospholipids. This first time identification of a menthol metabolite in smokers' blood serves as proof-of-principle for using metabolomics to identify new tobacco-exposure biomarkers, and also provides new opportunities in studying menthol-containing tobacco products in humans. Gender and race differences also were observed. Network analysis revealed 12 molecules involved in cancer, notably inhibition of cAMP. These novel tobacco-related biomarkers provide new insights to the effects of smoking which may be important in carcinogenesis but not previously linked with tobacco-related diseases. © 2016 Wiley Periodicals, Inc.

Genetic variation in one-carbon metabolism in relation to genome-wide DNA methylation in breast tissue from heathy women.

Single nucleotide polymorphisms (SNPs) in one-carbon metabolism genes and lifestyle factors (alcohol drinking and breast folate) may be determinants of whole-genome methylation in the breast. DNA methylation profiling was performed using the Illumina Infinium HumanMethylation450 BeadChip in 81 normal breast tissues from women undergoing reduction mammoplasty and no history of cancer. ANCOVA, adjusting for age, race and BMI, was used to identify differentially-methylated (DM) CpGs. Gene expression, by the Affymetrix GeneChip Human Transcriptome Array 2.0, was correlated with DM. Biological networks of DM genes were assigned using Ingenuity Pathway Analysis. Fifty-seven CpG sites were DM in association with eight SNPs in FTHFD, MTHFD1, MTHFR, MTR, MTRR, and TYMS (P <5.0 x 10-5); 56% of the DM CpGs were associated with FTHFD SNPs, including DM within FTHFD. Gene expression was negatively correlated with FTHFD methylation (r=-0.25, P=0.017). Four DM CpGs identified by SNPs in MTRR, MTHFR, and FTHFD were significantly associated with alcohol consumption and/or breast folate. The top biological network of DM CpGs was associated with Energy Production, Molecular Transportation, and Nucleic Acid Metabolism. This is the first comprehensive study of the association between SNPs in one-carbon metabolism genes and genome-wide DNA methylation in normal breast tissues. These SNPs, especially FTHFD, as well as alcohol intake and folate exposure, appear to affect DM in breast tissues of healthy women. The finding that SNPs in FTHFD and MTR are associated with their own methylation is novel and highlights a role for these SNPs as cis-methylation quantitative trait loci.

Persistent alterations of gene expression profiling of human peripheral blood mononuclear cells from smokers.

The number of validated biomarkers of tobacco smoke exposure is limited, and none exist for tobacco-related cancer. Additional biomarkers for smoke, effects on cellular systems in vivo are needed to improve early detection of lung cancer, and to assist the Food and Drug Administration in regulating exposures to tobacco products. We assessed the effects of smoking on the gene expression using human cell cultures and blood from a cross-sectional study. We profiled global transcriptional changes in cultured smokers' peripheral blood mononuclear cells (PBMCs) treated with cigarette smoke condensate (CSC) in vitro (n = 7) and from well-characterized smokers' blood (n = 36). ANOVA with adjustment for covariates and Pearson correlation were used for statistical analysis in this study. CSC in vitro altered the expression of 1 178 genes (177 genes with > 1.5-fold-change) at P < 0.05. In vivo, PBMCs of heavy and light smokers differed for 614 genes (29 with > 1.5-fold-change) at P < 0.05 (309 remaining significant after adjustment for age, race, and gender). Forty-one genes were persistently altered both in vitro and in vivo, 22 having the same expression pattern reported for non-small cell lung cancer. Our data provides evidence that persistent alterations of gene expression in vitro and in vivo may relate to carcinogenic effects of cigarette smoke, and the identified genes may serve as potential biomarkers for cancer. The use of an in vitro model to corroborate results from human studies provides a novel way to understand human exposure and effect. © 2015 Wiley Periodicals, Inc.