Long non-coding RNA FKSG29 regulates oxidative stress and endothelial dysfunction in obstructive sleep apnea.

Altered expressions of pro-/anti-oxidant genes are known to regulate the pathophysiology of obstructive sleep apnea (OSA).We aim to explore the role of a novel long non-coding (lnc) RNA FKSG29 in the development of intermittent hypoxia with re-oxygenation (IHR)-induced endothelial dysfunction in OSA. Gene expression levels of key pro-/anti-oxidant genes, vasoactive genes, and the FKSG29 were measured in peripheral blood mononuclear cells from 12 subjects with primary snoring (PS) and 36 OSA patients. Human monocytic THP-1 cells and human umbilical vein endothelial cells (HUVEC) were used for gene knockout and double luciferase under IHR exposure. Gene expression levels of the FKSG29 lncRNA, NOX2, NOX5, and VEGFA genes were increased in OSA patients versus PS subjects, while SOD2 and VEGFB gene expressions were decreased. Subgroup analysis showed that gene expression of the miR-23a-3p, an endogenous competitive microRNA of the FKSG29, was decreased in sleep-disordered breathing patients with hypertension versus those without hypertension. In vitro IHR experiments showed that knock-down of the FKSG29 reversed IHR-induced ROS overt production, early apoptosis, up-regulations of the HIF1A/HIF2A/NOX2/NOX4/NOX5/VEGFA/VEGFB genes, and down-regulations of the VEGFB/SOD2 genes, while the protective effects of FKSG29 knock-down were abolished by miR-23a-3p knock-down. Dual-luciferase reporter assays confirmed that FKSG29 was a sponge of miR-23a-3p, which regulated IL6R directly. Immunofluorescence stain further demonstrated that FKSGH29 knock-down decreased IHR-induced uptake of oxidized low density lipoprotein and reversed IHR-induced IL6R/STAT3/GATA6/ICAM1/VCAM1 up-regulations. The findings indicate that the combined RNA interference may be a novel therapy for OSA-related endothelial dysfunction via regulating pro-/anti-oxidant imbalance or targeting miR-23a-IL6R-ICAM1/VCAM1 signaling.

MicroRNA-23a-3p Down-Regulation in Active Pulmonary Tuberculosis Patients with High Bacterial Burden Inhibits Mononuclear Cell Function and Phagocytosis through TLR4/TNF-α/TGF-ß1/IL-10 Signaling via Targeting IRF1/SP1.

The aim of this study is to explore the role of microRNAs (miR)-21/23a/146a/150/155 targeting the toll-like receptor pathway in active tuberculosis (TB) disease and latent TB infection (LTBI). Gene expression levels of the five miRs and predicted target genes were assessed in peripheral blood mononuclear cells from 46 patients with active pulmonary TB, 15 subjects with LTBI, and 17 non-infected healthy subjects (NIHS). THP-1 cell lines were transfected with miR-23a-3p mimics under stimuli with Mycobacterium TB-specific antigens. Both miR-155-5p and miR-150-5p gene expressions were decreased in the active TB group versus the NIHS group. Both miR-23a-3p and miR-146a-5p gene expressions were decreased in active TB patients with high bacterial burden versus those with low bacterial burden or control group (LTBI + NIHS). TLR2, TLR4, and interleukin (IL)10 gene expressions were all increased in active TB versus NIHS group. MiR-23a-3p mimic transfection reversed ESAT6-induced reduction of reactive oxygen species generation, and augmented ESAT6-induced late apoptosis and phagocytosis, in association with down-regulations of the predicted target genes, including tumor necrosis factor (TNF)-α, TLR4, TLR2, IL6, IL10, Notch1, IL6R, BCL2, TGF-ß1, SP1, and IRF1. In conclusion, the down-regulation of miR-23a-3p in active TB patients with high bacterial burden inhibited mononuclear cell function and phagocytosis through TLR4/TNF-α/TGF-ß1/IL-10 signaling via targeting IRF1/SP1.

Whole Genome DNA Methylation Analysis of Active Pulmonary Tuberculosis Disease Identifies Novel Epigenotypes: PARP9/miR-505/RASGRP4/GNG12 Gene Methylation and Clinical Phenotypes.

We hypothesized that DNA methylation patterns may contribute to the development of active pulmonary tuberculosis (TB). Illumina's DNA methylation 450 K assay was used to identify differentially methylated loci (DML) in a discovery cohort of 12 active pulmonary TB patients and 6 healthy subjects (HS). DNA methylation levels were validated in an independent cohort of 64 TB patients and 24 HS. Microarray analysis identified 1028 DMLs in TB patients versus HS, and 3747 DMLs in TB patients after versus before anti-TB treatment, while autophagy was the most enriched signaling pathway. In the validation cohort, PARP9 and miR505 genes were hypomethylated in the TB patients versus HS, while RASGRP4 and GNG12 genes were hypermethylated, with the former two further hypomethylated in those with delayed sputum conversion, systemic symptoms, or far advanced lesions. MRPS18B and RPTOR genes were hypomethylated in TB patients with pleural involvement. RASGRP4 gene hypermethylation and RPTOR gene down-regulation were associated with high mycobacterial burden. TB patients with WIPI2/GNG12 hypermethylation or MRPS18B/FOXO3 hypomethylation had lower one-year survival. In vitro ESAT6 and CFP10 stimuli of THP-1 cells resulted in DNA de-methylation changes of the PARP9, RASGRP4, WIPI2, and FOXO3 genes. In conclusions, aberrant DNA methylation over the PARP9/miR505/RASGRP4/GNG12 genes may contribute to the development of active pulmonary TB disease and its clinical phenotypes, while aberrant DNA methylation over the WIPI2/GNG12/MARPS18B/FOXO3 genes may constitute a determinant of long-term outcomes.

miR-21-5p Under-Expression in Patients with Obstructive Sleep Apnea Modulates Intermittent Hypoxia with Re-Oxygenation-Induced-Cell Apoptosis and Cytotoxicity by Targeting Pro-Inflammatory TNF-α-TLR4 Signaling.

The purpose of this study is to explore the anti-inflammatory role of microRNAs (miR)-21 and miR-23 targeting the TLR/TNF-α pathway in response to chronic intermittent hypoxia with re-oxygenation (IHR) injury in patients with obstructive sleep apnea (OSA). Gene expression levels of the miR-21/23a, and their predicted target genes were assessed in peripheral blood mononuclear cells from 40 treatment-naive severe OSA patients, and 20 matched subjects with primary snoring (PS). Human monocytic THP-1 cell lines were induced to undergo apoptosis under IHR exposures, and transfected with miR-21-5p mimic. Both miR-21-5p and miR-23-3p gene expressions were decreased in OSA patients as compared with that in PS subjects, while TNF-α gene expression was increased. Both miR-21-5p and miR-23-3p gene expressions were negatively correlated with apnea hypopnea index and oxygen desaturation index, while TNF-α gene expression positively correlated with apnea hypopnea index. In vitro IHR treatment resulted in decreased miR-21-5p and miR-23-3p expressions. Apoptosis, cytotoxicity, and gene expressions of their predicted target genes-including TNF-α, ELF2, NFAT5, HIF-2α, IL6, IL6R, EDNRB, and TLR4-were all increased in response to IHR, while all were reversed with miR-21-5p mimic transfection under IHR condition. The findings provide biological insight into mechanisms by which IHR-suppressed miRs protect cell apoptosis via inhibit inflammation, and indicate that over-expression of the miR-21-5p may be a new therapy for OSA.

Co-upregulation of Toll-like receptors 2 and 6 on peripheral blood cells in patients with obstructive sleep apnea.

PURPOSE: Toll-like receptor (TLR) 2 can heterodimerise with TLR6 to detect diacylated lipoproteins. Hypoxia inducible factor-1 α co-ordinates selective induction of TLR2 and TLR6 during persistent hypoxia. We hypothesized that TLR 2/6 co-expression may be upregulated by chronic intermittent hypoxia with re-oxygenation (IHR) in obstructive sleep apnea (OSA). METHODS: TLR2/6 expressions on blood immune cells were measured in 144 patients with sleep-disordered breathing (SDB), including primary snoring (PS, n = 24), moderate to severe OSA (MSO, n = 60), very severe OSA (VSO, n = 36), and very severe OSA on continuous positive airway pressure (CPAP) treatment (VSOC, n = 24). An in vitro IHR experiment was also undertaken. RESULTS: Patients in both the MSO and VSO groups had increased TLR2/6 co-expression on CD16(+) neutrophil than those in the PS group. Patients in the VSOC group had reduced TLR2/6 co-expression on neutrophil than those in either the MSO or VSO group. Blood absolute neutrophil count was positively but weakly correlated with TLR2/6 co-expression on neutrophil. TLR2/6 co-expressions on both CD14(+) monocyte and CD3(+)CD4(+)T helper cell, and TLR2 expressions on both monocyte and T helper cell in SDB patients with low Minimum SaO2 (≦70%) were all higher than those with high Minimum SaO2. In vitro IHR for 1-4 days resulted in TLR2/6 co-upregulation on both neutrophil and monocyte. CONCLUSIONS: OSA patients had increased TLR2/6 co-expressions on blood immune cells, which were related to their immune cell counts and could be reversed with CPAP treatment. In vitro IHR could induce TLR2/6 co-upregulation.

Application and outlook of electrochemical technology in single-cell analysis.

Cellular heterogeneity, especially in some important diseased cells like tumor cells, acts as an invisible driver for disease development like cancer progression in the tumor ecosystem, contributing to differences in the macroscopic and microscopic detection of disease lesions like tumors. Traditional analysis techniques choose group information masked by the mean as the analysis sample, making it difficult to achieve precise diagnosis and target treatment, on which could be shed light via the single-cell level determination/bioanalysis. Hence, in this article we have reviewed the special characteristic differences among various kinds of typical single-cell bioanalysis strategies and electrochemical techniques, and then focused on the recent advance and special bio-applications of electrochemiluminescence and micro-nano electrochemical sensing mediated in single-cell bioimaging & bioanalysis. Especially, we have summarized the relevant research exploration of the possibility to establish the in-situ single-cell electrochemical methods to detect cell heterogeneity through determination of specific biomolecules and bioimaging of some important biological processes. Eventually, this review has explored some important advances of electrochemical single-cell detection techniques for the real-time cellular bioimaging and diagnostics of some disease lesions like tumors. It raises the possibility to provide the specific in-situ platform to exploit the versatile, sensitive, and high-resolution electrochemical single-cell analysis for the promising biomedical applications like rapid tracing of some disease lesions or in vivo bioimaging for precise cancer theranostics.

Autophagy impairment in patients with obstructive sleep apnea modulates intermittent hypoxia-induced oxidative stress and cell apoptosis via hypermethylation of the ATG5 gene promoter region.

BACKGROUND: Autophagy is a catabolic process that recycles damaged organelles and acts as a pro-survival mechanism, but little is known about autophagy dysfunction and epigenetic regulation in patients with obstructive sleep apnea (OSA). METHODS: Protein/gene expressions and DNA methylation levels of the autophagy-related genes (ATG) were examined in blood leukocytes from 64 patients with treatment-naïve OSA and 24 subjects with primary snoring (PS). RESULTS: LC3B protein expression of blood monocytes, and ATG5 protein expression of blood neutrophils were decreased in OSA patients versus PS subjects, while p62 protein expression of cytotoxic T cell was increased, particularly in those with nocturia. ATG5, ULK1, and BECN1 gene expressions of peripheral blood mononuclear cells were decreased in OSA patients versus PS subjects. LC3B gene promoter regions were hypermethylated in OSA patients, particularly in those with excessive daytime sleepiness, while ATG5 gene promoter regions were hypermethylated in those with morning headache or memory impairment. LC3B protein expression of blood monocytes and DNA methylation levels of the LC3B gene promoter region were negatively and positively correlated with apnea hyponea index, respectively. In vitro intermittent hypoxia with re-oxygenation exposure to human THP-1/HUVEC cell lines resulted in LC3B/ATG5/ULK1/BECN1 down-regulations and p62 up-regulation along with increased apoptosis and oxidative stress, while rapamycin and umbilical cord-mesenchymal stem cell treatment reversed these abnormalities through de-methylation of the ATG5 gene promoter. CONCLUSIONS: Impaired autophagy activity in OSA patients was regulated by aberrant DNA methylation, correlated with clinical phenotypes, and contributed to increased cell apoptosis and oxidative stress. Autophagy enhancers may be novel therapeutics for OSA-related neurocognitive dysfunction.

Next generation sequencing reveals miR-431-3p/miR-1303 as immune-regulating microRNAs for active tuberculosis.

OBJECTIVES: RNA therapeutics is an emerging field that widens the range of treatable targets and would improve disease outcome through bypassing the antibiotic bactericidal targets to kill Mycobacterium tuberculosis (M.tb). METHODS: We screened for microRNA with immune-regulatory functions against M.tb by next generation sequencing of peripheral blood mononuclear cells, followed by validation in an independent cohort. RESULTS: Twenty three differentially expressed microRNAs were identified between 12 active pulmonary TB patients and 4 healthy subjects, and 35 microRNAs before and after 6-month anti-TB therapy. Enriched predicted target pathways included proteoglycan, HIF-1 signaling, longevity-regulating, central carbon metabolism, and autophagy. We validated miR-431-3p down-regulation and miR-1303 up-regulation accompanied with corresponding changes in their predicted target genes in an independent validation cohort of 46 active TB patients, 30 latent TB infection subjects, and 24 non-infected healthy subjects. In vitro experiments of transfections with miR-431-3p mimic/miR-1303 short interfering RNA in THP-1 cells under ESAT-6 stimuli showed that miR-431-3p and miR-1303 were capable to augment and suppress autophagy/apoptosis/phagocytosis of macrophage via targeting MDR1/MMP16/RIPOR2 and ATG5, respectively. CONCLUSIONS: This study provides a proof of concept for microRNA-based host-directed immunotherapy for active TB disease. The combined miR-431-3p over-expression and miR-1303 knock-down revealed new vulnerabilities of treatment-refractory TB disease.

RNA interference inhibits high mobility group box 1 by lipopolysaccharide-activated murine macrophage RAW 264.7 secretion.

BACKGROUND: This study aims to evaluate the influence of RNA interference (RNAi) on the high mobility group box 1 (HMGB-1) in the lipopolysaccharide (LPS)-induced murine macrophage cell line RAW 264.7. MATERIALS AND METHODS: In order to observe the effect of RNAi on HMGB-1, tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), and transforming growth factor ß (TGF-ß) levels, the RAW 264.7 cell line was divided into five treatment groups to measure separately as a function of time of negative control, LPS stimulation only, LPS + HMGB-1 short interfering RNAs (siRNAs), LPS + negative control siRNAs (siNC), and LPS + nafamostat mesilate (NM). RESULTS: Measurement does show HMGB-1 expression in the LPS-activated macrophages in an explicit time-dependent manner. The HMGB-1 cellular level is consistently knocked down 80%∼85% by the siRNA; TNF-α, IL-6, and TGF-ß levels in turn significantly decrease following siRNA delivery to the inflammatory response. HMGB-1 expression is lower in the LPS + NM group than the LPS + HMGB-1 siRNA group at the initial stage, however, a significantly lower level of HMGB-1 in the siRNA group is observed 48 h later. The decrease of TNF-α, IL-6, and TGF-ß levels in the LPS-induced inflammatory response is also observed in both groups. CONCLUSIONS: Our results demonstrate that HMGB-1 RNAi treatment of LPS-stimulated macrophages inhibit HMGB-1 and remarkably reduce the LPS-induced inflammatory responses. Hence, RNAi is highly recommended as a potential candidate for a new therapeutic strategy to minimize or, to a lesser extent, prevent the LPS-induced inflammatory injury.

Thermally assisted mobility of nanodroplets on surfaces with weak defects.

HYPOTHESIS: Thermal activation plays an essential role in contact line dynamics on nanorough surfaces. However, the relation between the aforementioned concept and the sliding motion of nanodroplets remains unclear. As a result, thermally assisted motion of nanodroplets on nanorough surfaces is investigated in this work. EXPERIMENTS: Steady slide and random motion of nanodroplets on surfaces with weak defects are investigated by Many-body Dissipative Particle Dynamics. The surface roughness is characterized by the slip length acquired from the velocity profile associated with the flowing film. FINDINGS: The slip length is found to decline with increasing the defect density. The linear relationship between the sliding velocity and driving force gives the mobility and reveals the absence of contact line pinning. On the basis of the Navier condition, a simple relation is derived and states that the mobility is proportional to the slip length and the reciprocal of the product of viscosity and contact area. Our simulation results agree excellently with the theoretical prediction. In the absence of external forces, a two-dimensional Brownian motion of nanodroplets is observed and its mean square displacement decreases with increasing the defect density. The diffusivity is proportional to the mobility, consistent with the Einstein relation. This consequence suggests that thermal fluctuations are able to overcome contact line pinning caused by weak defects.

H3K23/H3K36 hypoacetylation and HDAC1 up-regulation are associated with adverse consequences in obstructive sleep apnea patients.

The aim of this study is to determine the roles of global histone acetylation (Ac)/methylation (me), their modifying enzymes, and gene-specific histone enrichment in obstructive sleep apnea (OSA). Global histone modifications, and their modifying enzyme expressions were assessed in peripheral blood mononuclear cells from 56 patients with OSA and 16 matched subjects with primary snoring (PS). HIF-1α gene promoter-specific H3K36Ac enrichment was assessed in another cohort (28 OSA, 8 PS). Both global histone H3K23Ac and H3K36Ac expressions were decreased in OSA patients versus PS subjects. H3K23Ac expressions were further decreased in OSA patients with prevalent hypertension. HDAC1 expressions were higher in OSA patients, especially in those with excessive daytime sleepiness, and reduced after more than 6 months of continuous positive airway pressure treatment. H3K79me3 expression was increased in those with high C-reactive protein levels. Decreased KDM6B protein expressions were noted in those with a high hypoxic load, and associated with a higher risk for incident cardiovascular events or hypertension. HIF-1α gene promoter-specific H3K36Ac enrichment was decreased in OSA patients versus PS subjects. In vitro intermittent hypoxia with re-oxygenation stimuli resulted in HDAC1 over-expression and HIF-1α gene promoter-specific H3K36Ac under-expression, while HDAC1 inhibitor, SAHA, reversed oxidative stress through inhibiting NOX1. In conclusions, H3K23/H3K36 hypoacetylation is associated with the development of hypertension and disease severity in sleep-disordered breathing patients, probably through up-regulation of HDAC1, while H3K79 hypermethylation is associated with higher risk of cardiovascular diseases, probably through down-regulation of KDM6B.

MicroRNA Sequencing Analysis in Obstructive Sleep Apnea and Depression: Anti-Oxidant and MAOA-Inhibiting Effects of miR-15b-5p and miR-92b-3p through Targeting PTGS1-NF-κB-SP1 Signaling.

The aim of this study was to identify novel microRNAs related to obstructive sleep apnea (OSA) characterized by intermittent hypoxia with re-oxygenation (IHR) injury. Illumina MiSeq was used to identify OSA-associated microRNAs, which were validated in an independent cohort. The interaction between candidate microRNA and target genes was detected in the human THP-1, HUVEC, and SH-SY5Y cell lines. Next-generation sequencing analysis identified 22 differentially expressed miRs (12 up-regulated and 10 down-regulated) in OSA patients. Enriched predicted target pathways included senescence, adherens junction, and AGE-RAGE/TNF-α/HIF-1α signaling. In the validation cohort, miR-92b-3p and miR-15b-5p gene expressions were decreased in OSA patients, and negatively correlated with an apnea hypopnea index. PTGS1 (COX1) gene expression was increased in OSA patients, especially in those with depression. Transfection with miR-15b-5p/miR-92b-3p mimic in vitro reversed IHR-induced early apoptosis, reactive oxygen species production, MAOA hyperactivity, and up-regulations of their predicted target genes, including PTGS1, ADRB1, GABRB2, GARG1, LEP, TNFSF13B, VEGFA, and CXCL5. The luciferase assay revealed the suppressed PTGS1 expression by miR-92b-3p. Down-regulated miR-15b-5p/miR-92b-3p in OSA patients could contribute to IHR-induced oxidative stress and MAOA hyperactivity through the eicosanoid inflammatory pathway via directly targeting PTGS1-NF-κB-SP1 signaling. Over-expression of the miR-15b-5p/miR-92b-3p may be a new therapeutic strategy for OSA-related depression.

Using betaxolol for the prevention of paronychia induced by epidermal growth factor receptor inhibitors: a case-control cohort study.

BACKGROUND: High rates of posttreatment discomfort, infection, recurrence, and increased time to return to work have been noted after nail plate avulsion resulting from epidermal growth factor receptor tyrosine kinase inhibitor(EGFR-TKI)-induced paronychia, which may even interrupt the course of treatment for EGFR-TKI therapy. Thus, we conducted this study to determine how effectively a topical ß-blocker, betaxolol, prevents EGFR-TKI-induced paronychia. METHODS: This case-control cohort study included a total of 131 non-small-cell lung cancer patients. The prevention group comprised 40 patients treated with topical betaxolol 0.25% solution to prevent paronychia while they received EGFR-TKI therapy. The control group comprised 91 patients who did not preventively use topical betaxolol 0.25% solution while receiving EGFR-TKI therapy. The patients' age, gender, antineoplastic regimen, duration of antineoplastic treatment before the appearance of lesions, number of involved digits (fingernails or toenails) with lesions, grading of paronychia, and pain score were recorded. RESULTS: In terms of the cumulative incidence of paronychia, significant differences (P < 0.01) were noted at both the 2nd and 3rd months after starting EGFR-TKIs. Furthermore, the average visual analogue scale scores were 3.125 and 6.29 in the prevention group and control group, respectively (P < 0.01). The average grades of paronychia were 1.5 and 2.12 in the prevention group and control group, respectively (P < 0.01). The average numbers of involved digits were 2.25 (range: 1-5 digits) in the prevention group and 3.03 (range: 1-7) in the control group (P = 0.07). CONCLUSIONS: Preventively using topical betaxolol can significantly decrease the incidence, VAS score, and grading of EGFR-TKI-induced paronychia.

Aberrant DNA methylation levels of the formyl peptide receptor 1/2/3 genes are associated with obstructive sleep apnea and its clinical phenotypes.

BACKGROUND: FPR1 over-expression and insufficiency of FPR2 and FPR3 are associated with disease severity of obstructive sleep apnea (OSA). We hypothesized that epigenetic modification of the FPR1/2/3 genes may underlie intermittent hypoxia with re-oxygenation (IHR) injury in OSA. METHODS: DNA methylation levels over 17 CpG sites of the FPR1/2/3 genes and their gene expression levels in the peripheral blood mononuclear cells were determined in 40 treatment-naïve OSA patients, 12 severe OSA patients under long-term continuous positive airway pressure treatment, 16 primary snoring (PS) subjects, and 10 healthy non-snorers (HS). RESULTS: Both -524 and -264 CpG sites of the FPR1 gene were hypomethylated in treatment-naïve OSA versus HS, while -264 CpG site methylation level was negatively correlated with FPR1/FPR3 gene expression ratio and associated with prevalent diabetes mellitus. Both +8802 and +8845 CpG sites of the FPR2 gene were hypermethylated in treatment-naive OSA versus HS, while hypermethylated +9132 and +9150 CpG sites were both associated with prevalent hypertension. FPR3 gene expression and DNA methylation levels over -842/-516 CpG sites of the FPR3 gene were both decreased in treatment-naive OSA versus HS, while hypermethylated -429 CpG site was associated with elevated serum C-reactive protein level. In vitro IHR stimuli in human monocytic THP-1 cells resulted in gene promoter hypomethylation-mediated FPR1 over-expression, increased production of reactive oxygen species, and increased cell apoptosis, which could be reversed with re-methylation agent, folic acid, treatment. CONCLUSIONS: Aberrant DNA methylation patterns of the FPR1/2/3 gene promoters contribute to disease severity and diabetes mellitus or cardiovascular disease in OSA patients, probably through regulating FPR1/2/3 gene expressions.

Blood M2a monocyte polarization and increased formyl peptide receptor 1 expression are associated with progression from latent tuberculosis infection to active pulmonary tuberculosis disease.

OBJECTIVES: This study aims to explore the role of M2a polarization and formyl peptide receptor (FPR) regulation in the reactivation of Mycobacterium tuberculosis (Mtb) infection. METHODS: M1/M2a monocyte percentage and FPR1/2/3 protein expression of blood immune cells were measured in 38 patients with sputum culture (+) active pulmonary TB disease, 18 subjects with latent TB infection (LTBI), and 28 noninfected healthy subjects (NIHS) using flow cytometry method. RESULTS: M1 percentage was decreased in active TB versus either NIHS or LTBI group, while M2a percentage and M2a/M1 percentage ratio were increased. FPR1 expression on M1/M2a, FPR2 expression on M1, and FPR3 expression of M1 were all decreased in active TB versus LTBI group, while FPR1 over FPR2 expression ratio on NK T cell was increased in active TB versus either NIHS or LTBI group. In 11 patients with active TB disease, M1 percentage became normal again after anti-TB treatment. In vitro Mtb-specific antigen stimulation of monocytic THP-1 cells resulted in M2a polarization in association with increased FPR2 expression on M2a. CONCLUSIONS: Increased M2a and decreased M1 phenotypes of blood monocyte may serve as a marker for active TB disease, while decreased FPR1 on blood monocyte may indicate LTBI status.

Aberrant DNA methylation of the toll-like receptors 2 and 6 genes in patients with obstructive sleep apnea.

Obstructive sleep apnea (OSA) is a syndrome leading to chronic intermittent hypoxia, and the up-regulation of toll-like receptors (TLR) 2 and 6 on peripheral blood cells has been reported. We hypothesized that DNA methylation in TLR2 and TLR6 genes may play a role in the development of OSA and its excessive daytime sleepiness (EDS) phenotype. DNA methylation over 28 cytosine-phosphate-guanine (CpG) sites of the TLR2 promoter region and 3 CpG sites of the TLR6 gene body, and their protein expressions were measured by using pyrosequencing and ELISA methods in 18 heathy subjects (HS) and 58 patients with severe OSA (divided into 18 non-EDS and 40 EDS group). Patients with severe OSA had higher DNA methylation levels over five CpG sites (#1, #2, #3, #25 and #28) and lower DNA methylation levels over CpG site #18 of the TLR2 promoter region, higher DNA methylation levels over two CpG sites (#1 and #3) of the TLR6 gene body, and higher protein expressions of TLR6 than HS. The CpG site #2 of the TLR6 gene body was hypermethylated in severe OSA patients with EDS. Both DNA methylation levels over CpG site #1 of the TLR6 gene body and protein expressions of TLR6 were reduced after more than 6 months of nasal CPAP treatment in seven selected patients. Aberrant DNA methylation of the TLR2 promoter region and TLR6 gene body are associated with the consequence of severe OSA and its EDS phenotype.

Formyl peptide receptor 1 up-regulation and formyl peptide receptor 2/3 down-regulation of blood immune cells along with defective lipoxin A4/resolvin D1 production in obstructive sleep apnea patients.

BACKGROUND: This study aims to investigate the role of FPR 1/2/3 expressions in patients with obstructive sleep apnea (OSA). METHOD: We made cross-sectional comparisons of FPR1/2/3 expressions of blood neutrophil, M1/M2a monocyte, and natural killer (NK) cell between 16 healthy subjects (HS), 16 primary snoring (PS) subjects, 46 treatment-naive OSA patients, and 18 severe OSA patients under long-term continuous positive airway pressure treatment (severe OSA on CPAP). RESULTS: FPR1 expressions on neutrophil were increased in treatment-naive OSA and severe OSA on CPAP groups versus either HS or PS. FPR2 expressions on neutrophil were decreased in treatment-naive OSA versus HS, and returned to normal in severe OSA on CPAP group. FPR1/FPR2 expression ratio on neutrophil was increased in treatment-naive OSA versus either HS or PS. Serum lipoxin A4, resolvin D1 levels, and FPR3 expressions of M1, M2a and NK cells were all decreased in treatment-naive OSA versus HS. OSA patients with hypertension had decreased FPR2 expressions on neutrophil and FPR3 expressions of NK cell. FPR1 expression, FPR1/FPR2 expression ratio on neutrophil, and FPR3 expression of M1 cell were all reversed after > 6-month CPAP treatment in 9 selected patients. In vitro intermittent hypoxia with re-oxygenation treatment in THP-1 cells resulted in increased FPR1/FPR2 expression ratio of M1 cells, and increased FPR1/FPR3 expression ratio of M2a cells. CONCLUSIONS: FPR1 over-expression and insufficiency of FPR2 and FPR3 in association with defective lipoxin A4 and resolving D1 production were associated with disease severity of OSA and its adverse consequences.

Increased S100A15 expression and decreased DNA methylation of its gene promoter are involved in high metastasis potential and poor outcome of lung adenocarcinoma.

PURPOSE: This study aims to determine the functional role of S100A15 and its promoter DNA methylation patterns in lung cancer progression. EXPERIMENTAL DESIGN: We analyzed 178 formalin-fixed paraffin embedded specimens from lung cancer patients, including 24 early stage and 91 advanced stage adenocarcinoma. S100A15 protein expression was evaluated by immunohistochemistry stain, and its DNA methylation levels were measured by pyrosequencing. RESULTS: S100A15 nuclear staining was increased in lung adenocarcinoma patients with distant metastasis versus those without distant metastasis. There was reduced one/three-year overall survival in adenocarcinoma patients receiving first line target therapy and harboring high nuclear expressions of S100A15. Both DNA methylation levels over -423 and -248 CpG sites of the S100A15 gene promoter were decreased in adenocarcinoma patients with distant metastasis, and the former was associated with lower one-year overall survival. The highly invasive CL1-5 cell lines display decreased DNA methylation over -412/-248/-56 CpG sites of the S100A15 gene promoter and increased S100A15 gene/protein expressions as compared with the less invasive CL1-0 cell lines. Knockdown of S100A15 in CL1-5 cell line inhibited cell proliferation, migration, and invasion, while over-expression of S100A15 in CL1-0 cell line promoted cell proliferation, migration, and invasion. RNA sequencing analysis revealed potential biological effects of S100A15 over-expression and knock-down with CTNNB1, ZEB1, CDC42, HSP90AA1, BST2, and PCNA being the pivotal down-stream mediators. CONCLUSIONS: Increased S100A15 expression and decreased DNA methylation of its gene promoter region were associated with high metastasis potential and poor outcome in lung adenocarcinoma, probably through triggering CTNNB1 -centered pathways.

Histone H3K14 hypoacetylation and H3K27 hypermethylation along with HDAC1 up-regulation and KDM6B down-regulation are associated with active pulmonary tuberculosis disease.

The aim of this study is to determine the roles of global histone acetylation (Ac)/methylation (me), their modifying enzymes, and gene-specific histone enrichment in active pulmonary tuberculosis (TB) disease. Global histone H3K27me3, H3K27me2, H3K9me3, H3K9Ac, and H3K14Ac expressions, and their modifying enzyme expressions, including KDM1A, KDM6B, EZH2, HDAC1, and HDAC2, were assessed in blood leukocytes from 81 patients with active pulmonary TB disease and 44 matched healthy subjects (HS). TLR2, TNF-α, IFN-γ, and IL12B-specific histone enrichment of peripheral blood mononuclear cells was measured by chromatin immunoprecipitation method. We found that Global H3K14Ac was decreased and H3K27me2 was increased in TB patients as compared with that in HS. TB patients with low H3K14Ac had lower one-year survival. Global H3K27me3 was increased in TB patients with high bacterial burden, or systemic symptoms as compared with that in those without the attribute or HS. HDAC1 gene/protein expressions were increased in TB patients as compared with that in HS, whereas KDM6B gene/protein expressions were decreased. Global H3K27me2, HDAC1 and KDM6B protein expressions were all reversed to normal after 6-month anti-TB treatment. TNF-α/IL12B promoter-specific H3K14Ac and TNF-α/IL12B/IFN-γ promoter-specific H3K27me2 enrichment were all decreased in 10 TB patients as compared with that in 10 HS. Among them, IL12B-specific H3K27me2 enrichment was reversed to normal after treatment, while the other 4 remained depressed. In conclusions, H3K14 hypoacetylation and H3K27 hypermethylation play a role in the development of active pulmonary TB disease or its clinical phenotypes, probably through up-regulation of HDAC1 and down-regulation of KDM6B, respectively.