G1 phase cell cycle arrest in NSCLC in response to LZ-106, an analog of enoxacin, is orchestrated through ROS overproduction in a P53-dependent manner.

LZ-106, a newly synthetized analog of quinolone, has been shown to be highly effective in non-small cell lung cancer (NSCLC) in both cultured cells and xenograft mouse model with low toxicity, yet the molecular mechanisms still require exploration. Here, we substantiated the involvement of P53 activation in intracellular reactive oxygen species (ROS) generation upon LZ-106 treatment and related P53 to the ROS-induced viability inhibition and apoptosis, which was exhibited in the previous research. P53 was shown to play an indispensable role in the elevated levels of intracellular ROS in LZ-106-treated NSCLC cells through ROS detection. We further identified the anti-proliferation effect of LZ-106 in NSCLC cells through G1 phase cell cycle arrest by cell cycle analysis, with the expression analysis of the key proteins, and discovered that the cell cycle arrest effect is also mediated by induction of ROS in a P53-dependent manner. In addition, the tumor suppression effect exhibited in vivo was demonstrated to be similar to that in vitro, which requires the participation of P53. Thus, LZ-106 is a potent antitumor drug possessing potent proliferation inhibition and apoptosis induction ability through the P53-dependent ROS modulation both in vitro and in vivo.

Silica nanoparticles as an enhancer in the IL-1ß-induced inflammation cycle of A549 cells.

Objective: The industrial production and combustion of coal can produce silica nanoparticles (nano-SiO2). It enters the human body mainly through the respiratory tract and exerts a toxic effect. However, whether nano-SiO2 can increase the IL-1ß-induced inflammatory expression in A549 cells has not been tested. Therefore, the synergistic toxicity of nano-SiO2 and IL-1ß to A549 was observed in our study. Materials and methods: We exposed A549 cells to nano-SiO2 (0, 100, 500, and 1000 µg/ml) for 12 and 24 h. The effect of nano-SiO2 on the viability of A549 cells was observed by the CCK-8 method. The A549 cells were exposed to nano-SiO2 (1 mg/mL) and cytokine IL-1ß (10 ng/mL) for 4 h, and we detected the expression of IL-1ß and IL-6 cytokines by real time quantitative polymerase chain (RT-qPCR) and enzyme linked immunosorbent assay (ELISA). The expression of ß-Actin, I-κB, phospho-ERK1/2 (P-ERK1/2), total-ERK1/2 (T-ERK1/2), phospho-JNK (P-JNK), total-JNK (T-JNK), phospho-P38 (P-P38), and total-P38 (T-P38) in A549 cells was detected by the Western Blot method. Results: The nano-SiO2 treatment resulted in a time-dependent decrease in the viability of A549 cells. The synergistic effect of nano-SiO2 and IL-1ß was observed on the new production of IL-1ß and IL-6 in A549 cells. The Western blot results showed that nano-SiO2 can increase the expression of IL-1ß and IL-6 by promoting the phosphorylation of ERK1/2 and elevating the phosphorylation of I-κB by IL-1ß. IL-1ß and IL-6 were induced by nano-SiO2, and the IL-1ß treatment with 20 µM of I-κBα phosphorylation inhibitor (PD98059) and 20 µM of ERK1/2 inhibitor (BAY11-7082) for 1 h was significantly lower than that of the control group in A549 cells. Discussion and conclusion: These results indicated that nano-SiO2 had a toxic effect on A549 cells, and this effect could increase IL-1ß on the A549 cell-induced inflammatory response. The results suggested that the release of IL-1ß and IL-6 in A549 was enhanced by the synergistic IL-1ß-induced phosphorylation of ERK1/2 and I-κB. This process is similar to a snowball, and it is possible that IL-1ß is continuously produced and repeatedly superimposed in A549 cells to produce an inflammatory effect; then, a vicious circle occurs, and an inflammatory storm is accelerated.

LZ-106, a potent lysosomotropic agent, causing TFEB-dependent cytoplasmic vacuolization.

Cytoplasmic vacuolization usually occurs in cells treated with different agents and substances. We found that LZ-106, an analog of enoxacin, is a potent lysosomotropic agent, contributing to the formation of cytoplasmic vacuoles in cells. Studies of LZ-106-induced vacuolization in H460 cells showed acid environment inside these vacuoles. Further study demonstrated that markers in the late endosomes and lysosomes, like LAMP1 and RAB7, on the surface of the vacuoles, implying that these vacuoles might derive from endosomes and/or lysosomes. By studying the fluorescence intensity of LZ-106, we discovered that LZ-106 tended to locate in acid organelles, and Bafilomycin A1, a V-ATPase inhibitor, was able to suppress its acid organelles localization. Also, we noticed that LZ-106 could induce lysosome stress, involving pH increment and lysosomal membrane damage. Moreover, the expression levels of some lysosome-related proteins, like LAMP1, EEA1, and Cathepsin B, were also altered upon LZ-106 treatment. At last, we confirmed LZ-106 can activate TFEB, a key regulator of lysosomes. Knockdown of TFEB could also reverse LZ-106's effect on vacuolization in H460 cells. Taken together, due to LZ-106's lysosomotropic properties, it is able to accumulate in the acid organelles and induce lysosomal dysfunction in H460 cells, leading to TFEB activation and the following cytoplasmic vacuolization.

Serum miR-638 Combined with Squamous Cell Carcinoma-Related Antigen as Potential Screening Biomarkers for Cervical Squamous Cell Carcinoma.

Aims: Cervical cancer is the second most common cause of cancer-related deaths in developing nations. Human papillomavirus prophylactic vaccines are not widely available, and there are shortages of gynecologists and cytologists in the already overburdened health care systems. The aim of this study was to identify circulating microRNAs (miRNAs) that could be used as feasible screening tests for cervical cancer in low-resource regions. Materials and Methods: Serum expression levels of five miRNAs were measured and validated by quantitative real-time polymerase chain reaction in cervical squamous cell carcinoma (CSCC) patients, cervical intraepithelial neoplasia patients, and healthy individuals. Squamous cell carcinoma-related antigen (SCC-Ag) was also measured in the serum. Results: Serum miR-638, miR-203a-3p, miR-1914-5p, and miR-521 levels were downregulated in the CSCC group (p < 0.05). Receiver operating characteristic (ROC) curve analysis indicated that the area under the ROC curve (AUC) values for miR-638 and miR-521 were 0.734 and 0.742, respectively, for discriminating CSCC patients from healthy controls. Furthermore, the combined use of miR-638 and SCC-Ag yielded the best screening performance and increased the AUC value, sensitivity, and specificity to 0.956, 94.87%, and 80.00%, respectively. Conclusion: This study suggested that miR-638 and miR-521 have independent screening value and that the combined measurement of miR-638 and SCC-Ag resulted in a better ability to discriminate patients with CSCC from healthy individuals.

PM2.5 promotes replication of VSV by ubiquitination degradation of phospho-IRF3 in A549 cells.

Both PM2.5 and respiratory viruses are part of the atmospheric constituents. Respiratory viruses are often associated with PM2.5 exposure, but the mechanism of toxicity remains to be explored. The vitro models that adequately reproduce healthy cells or diseased cells exposing to PM2.5 and infecting VSV can provide a useful tool for studying innate immune mechanisms and investigating new therapeutic focus. In the environment of PM2.5, an infection model in which VSV infected A549 cells was established, that mimics the state in which the antiviral innate immune pathways are activated after the respiratory system is infected with RNA viruses. Subsequently, the model was exposed to PM2.5 for 24 h. PM2.5 could be ingested by A549 cells and synergize with VSV to inhibit cell viability and promote apoptosis. The expression of VSV-G were more abundant after VSV-infected A549 cells were exposed to PM2.5. Furthermore, PM2.5 inhibits VSV-induced IFN-ß expression in A549 cells. ISG15, CCL-5, and CXCL-10 had the same expression tendency with IFN-ß mRNA, consistently. Interestingly, when MG132 was applied, the expression of p-IRF-3 and IFN-ß proteins reduced by PM2.5 were refreshed. Conversely, the expression of VSV-G proteins were decreased. PM2.5 could degrade p-IRF-3 proteins by ubiquitination pathway to inhibit VSV-induced IFN-ß expression in A549 cells. Therefore, replication of the VSV viruses was promoted.

Association of Polymorphisms in miRNA Processing Genes With Type 2 Diabetes Mellitus and Its Vascular Complications in a Southern Chinese Population.

Objective: To evaluate the potential association between the genetic variants in miRNA processing genes (RAN, XPO5, DICER1, and TARBP2) and susceptibility to type 2 diabetes mellitus (T2DM) and its vascular complications, as well as to further investigate their interaction with environmental factors in type 2 diabetes. Methods: We conducted a case-control study in genotyping of five polymorphic loci, including RAN rs14035, XPO5 rs11077, DICER1 rs13078, DICER1 rs3742330, and TARBP2 rs784567, in miRNA processing genes to explore the risk factors for T2DM and diabetic vascular complications. Haplotype analyses, interactions of gene-gene and interactions of gene-environment were performed too. Results: We identified a 36% decreased risk of developing T2DM in individuals with the minor A allele in DICER1 rs13078 (OR: 0.64; 95%CI: 0.42-0.95; P: 0.026). The AA haplotype in DICER1 was also associated with a protective effect on T2DM compared with the AT haplotype (OR: 0.63; 95%CI: 0.42-0.94; P-value: 0.023). T2DM patients with the TT+TC genotype at RAN rs14035 had a 1.89-fold higher risk of developing macrovascular complications than patients with the CC genotype (OR: 1.89; 95%CI: 1.04-3.45; P-value: 0.037). We also identified two three-factor interaction models. One is a three-factor [DICER1 rs13078, body mass index (BMI), and triglyceride (TG)] interaction model for T2DM (OR: 5.93; 95%CI: 1.25-28.26; P = 0.025). Another three-factor [RAN rs14035, hypertension (HP), and duration of T2DM (DOD)] interaction model was found for macrovascular complications of T2DM (OR = 41.60, 95%CI = 11.75-147.35, P < 0.001). Conclusion: Our study provides new evidence that two single nucleotide polymorphisms (SNPs) of the miRNA processing genes, DICER1 and RAN, and their interactions with certain environmental factors might contribute to the risk of T2DM and its vascular complications in the southern Chinese population.

Association Study Between Methylation in the Promoter Regions of cGAS, MAVS, and TRAF3 Genes and the Risk of Cervical Precancerous Lesions and Cervical Cancer in a Southern Chinese Population.

A case-control study was used to explore the association between the methylation status in the promoter regions of the cGAS, MAVS, and TRAF3 genes and the diseases of cervical precancerous lesions (CPL) and cervical cancer (CC) in a Southern Chinese population, and to further explore their interaction effects with high-risk human papillomavirus (hrHPV) infection and environmental factors in these diseases. The study protocol was approved by the ethics committee of The First Affiliated Hospital of Jinan University, and this study was performed in 97 healthy controls, 75 patients with CPL and 33 patients with CC, while each participant has read and signed the informed consent forms before enrolment. The promoter methylation status genes were detected from the bisulfite-treated DNA by the bisulfite sequencing PCR (BSP) technique, which was carried out using MethPrimer. The cGAS, MAVS, and TRAF3 promoter methylation levels in CPL (CPL cGAS = 35.40%, CPL MAVS = 24.26%, and CPL TRAF3 = 96.76%) were significantly higher than those in the control (Control cGAS = 31.87%, Control MAVS = 21.16%, and Control TRAF3 = 96.26%, PcGAS < 0.001, PMAVS < 0.001, and PTRAF3 = 0.001); however, there was no significant differences between the CC and control. In the logistic regression model with adjusted covariates, compared with the individuals whose cGAS methylation levels were less than or equal to 31.87%, the women with the levels more than 31.87% increased the risk of CPL by 2.49 times (ORa = 2.49, 95% CI = 1.31-4.75, P a = 0.006). The women with MAVS methylation levels above 21.16% were 1.97 times more likely to have CPL than the those with the levels less than 21.16% (ORa = 1.97, 95% CI = 1.06-3.69, P a = 0.033). A synergistic interaction was found between hrHPV and gene promoter methylation levels of cGAS and MAVS in CPL; however, no potential interaction was observed in CC. The promoter methylation levels in cGAS, MAVS, and TRAF3 genes are higher in CPL than in control, indicating that hypermethylation might be an early event in the progression of cervical intraepithelial neoplasia (CIN). The interaction between the promoter methylation levels in cGAS and MAVS genes and hrHPV infection might play a role in the development of CPL.

Toxoplasma gondii Cathepsin C1 inhibits NF-κB signalling through the positive regulation of the HIF-1α/EPO axis.

Toxoplasma gondii has evolved many successful strategies for immune evasion. However, the parasite-derived effectors involved in modulating NF-κB signalling pathway are largely unknown. T. gondii Cathepsin C1 (CPC1) is widely conserved among T. gondii strains and is important for T. gondii intracellular growth and proliferation. Our study showed that CPC1 protein could abrogate NF-κB activation after screening dense granule proteins. CPC1 suppressed NF-κB activation at or downstream of p65 and decreased the production of IL-1, IL-8, IL-6, IL-12, and TNF-α. Western blot analysis revealed that CPC1 inhibited phospho-p65 and CPC1 proteins primarily settled in cytoplasm. RNA sequencing analysis revealed that overexpression of CPC1 significantly upregulated erythropoietin (EPO), which can be induced by the hypoxia-inducible factor -1α (HIF-1α) during hypoxia. Furthermore, dual-luciferase reporter assays confirmed that CPC1 upregulated HIF-1α. Finally, both the knockdown of EPO and restriction of HIF-1α partially eliminated the suppression impact of CPC1 on the NF-κB signalling pathway. Our study identified a previously unrecognized role of CPC1 in the negative regulation of NF-κB activation through positive regulation of the HIF-1α/EPO axis. For the first time, CPC1 was shown to play an important role in immune evasion during T. gondii infection.

Association Between MALAT1 and THRIL Polymorphisms and Precancerous Cervical Lesions.

BACKGROUND: The occurrence of cervical cancer is a complex process, for which human papillomavirus (HPV) infection is a risk factor, although not all women infected with HPV will develop the disease. Knockout of mammalian lung metastasis associated transcript 1 (MALAT1) is associated with increased risk for several cancer types, whereas the long non-coding RNA (lncRNA) THRIL is essential for induction of tumor necrosis factor-α expression, which plays important roles in HPV infection. MATERIALS AND METHODS: To investigate the effects of polymorphisms in the lncRNAs MALAT1 and THRIL on the susceptibility to precancerous cervical lesions, 12 single nucleotide polymorphisms (SNPs) were analyzed from 164 cervical precancerous lesion cases and 428 controls. Gene-gene and gene-environment interactions and haplotype associations were also evaluated. RESULTS: We found a significantly decreased risk of precancerous cervical lesions for the THRIL rs7133268 AG genotype (odds ratio adjusted = 0.63, 95% confidence interval: 0.42-0.94, p = 0.025). Multifactor dimensionality reduction analysis identified a significant two-locus interaction model involved in HPV infection and THRIL rs7133268 (training balanced accuracy = 0.6957, testing balanced accuracy = 0.6948, cross-validation consistency = 10/10, p = 0.0046). Other SNPs, including the two identified for MALAT1, were not significantly related to the risk of precancerous cervical lesions. CONCLUSION: Our results suggest that the rs7133268 polymorphism of the lncRNA THRIL gene can reduce the genetic susceptibility of precancerous cervical lesions and in turn reduce the risk of HPV infection.

Association Analysis of NLRP3 Inflammation-Related Gene Promotor Methylation as Well as Mediating Effects on T2DM and Vascular Complications in a Southern Han Chinese Population.

Objective: To explore the association between the methylation levels in the promoter regions of the NLRP3, AIM2, and ASC genes and T2DM and its vascular complications in a Southern Han Chinese population and further analyze their interaction and mediating effects with environmental factors in T2DM. Methods: A case-control study was used to determine the association between population characteristics, the methylation level in the promoter region of the NLRP3, AIM2, and ASC genes and T2DM and vascular complications. A mediating effect among genes-environment-T2DM and the interaction of gene-gene or gene-environment factors was explored. Results: In the logistic regression model with adjusted covariants, healthy people with lower total methylation levels in the AIM2 promoter region exhibited a 2.29-fold [OR: 2.29 (1.28~6.66), P = 0.011] increased risk of developing T2DM compared with higher-methylation individuals. T2DM patients without any vascular complications who had lower methylation levels (

Association of variants of miRNA processing genes with cervical precancerous lesion risk in a southern Chinese population.

The miRNA processing genes play essential roles in the biosynthesis of mammalian miRNAs, and their genetic variants are involved in the development of various cancers. Our study aimed to determine the potential association between miRNA processing gene polymorphisms and cervical precancerous lesions. Five single nucleotide polymorphisms (SNPs), including Ran-GTP (RAN) rs14035, exportin-5 (XPO5) rs11077, DICER1 rs3742330, DICER1 rs13078, and TARBP2 rs784567, were genotyped in a case-control study to estimate risk factors of cervical precancerous lesions. The gene-environment interactions and haplotype association were estimated. We identified a 27% decreased risk of cervical precancerous lesions for individuals with minor G allele in DICER1 rs3742330 (odds ratio (OR) = 0.73, 95% confidence interval (95% CI) = 0.58-0.92, P = 0.009). The AG and AG/GG genotypes in DICER1 rs3742330 were also found to decrease the risk of cervical precancerous lesions (AG compared with AA: OR = 0.51, 95% CI = 0.35-0.73, P <0.001; AG/GG compared with AA: OR = 0.54, 95% CI = 0.39-0.77, P = 0.001). The GT haplotype in DICER1 had a risk effect on cervical precancerous lesions compared with the AT haplotype (OR = 1.36, 95% CI = 1.08-1.73, P = 0.010). A two-factor (DICER1 rs3742330 and human papillomavirus (HPV) infection) and two three-factor (model 1: rs3742330, passive smoking, and HPV infection; model 2: rs3742330, abortion history, and HPV infection) interaction models for cervical precancerous lesions were identified. In conclusion, the genetic variants in the miRNA processing genes and interactions with certain environmental factors might contribute to the risk of cervical precancerous lesions in southern Chinese women.

Association Between SLC30A8 rs13266634 Polymorphism and Risk of T2DM and IGR in Chinese Population: A Systematic Review and Meta-Analysis.

Introduction: Published data regarding the association between solute carrier family 30, member 8 (SLC30A8) rs13266634 polymorphism and type 2 diabetes mellitus (T2DM) and impaired glucose regulation (IGR) risks in Chinese population are in-consistent. The purpose of this meta-analysis was to evaluate the association between SLC30A8 rs13266634 and T2DM/IGR in a Chinese population. Material and Methods: Three English (PubMed, Embase, and Web of Science) and three Chinese databases (Wanfang, CNKI, and CBMD database) were used for searching articles from January 2005 to January 2018. Odds ratio (OR) and 95% confidence interval (95%CI) were calculated with the random-effect model. Trial sequential analysis was also utilized. Results: Twenty-eight case-control studies with 25,912 cases and 26,975 controls were included for SLC30A8 and T2DM. Pooled risk allele C frequency for rs13266634 was 60.6% (95%CI: 59.2-62.0%) in the T2DM group and 54.8% (95%CI: 53.2-56.4%) in the control group which had estimated OR of 1.23 (95%CI: 1.17-1.28). Individuals who carried major homozygous CC and heterozygous CT genotype were at 1.51 and 1.23 times higher risk of T2DM, respectively, than those carrying minor homozygous TT. The most appropriate genetic analysis model was the co-dominant model based on comparison of OR1, OR2 and OR3. Five articles that involved 4,627 cases and 6,166 controls were included for SLC30A8 and IGR. However, no association was found between SLC30A8 rs13266634 and IGR (C vs. T, OR = 1.13, 95%CI: 0.98-1.30, p = 0.082). TSA revealed that the pooled sample sizes of T2DM exceeded the estimated required information size but not the IGR. Conclusion: The present meta-analysis demonstrated that SLC30A8 rs13266634 was a potential risk factor for T2DM, and more studies should be performed to confirm the association between rs13266634 polymorphism and IGR.

Global Epidemiology of Dengue Outbreaks in 1990-2015: A Systematic Review and Meta-Analysis.

Dengue is an arthropod-borne infectious disease caused by dengue virus (DENV) infection and transmitted by Aedes mosquitoes. Approximately 50-100 million people are infected with DENV each year, resulting in a high economic burden on both governments and individuals. Here, we conducted a systematic review and meta-analysis to summarize information regarding the epidemiology, clinical characteristics, and serotype distribution and risk factors for global dengue outbreaks occurring from 1990 to 2015. We searched the PubMed, Embase and Web of Science databases through December 2016 using the term "dengue outbreak." In total, 3,853 studies were identified, of which 243 studies describing 262 dengue outbreaks met our inclusion criteria. The majority of outbreak-associated dengue cases were reported in the Western Pacific Region, particularly after the year 2010; these cases were primarily identified in China, Singapore and Malaysia. The pooled mean age of dengue-infected individuals was 30.1 years; of the included patients, 54.5% were male, 23.2% had DHF, 62.0% had secondary infections, and 1.3% died. The mean age of dengue patients reported after 2010 was older than that of patients reported before 2010 (34.0 vs. 27.2 years); however, the proportions of patients who had DHF, had secondary infections and died significantly decreased after 2010. Fever, malaise, headache, and asthenia were the most frequently reported clinical symptoms and signs among dengue patients. In addition, among the identified clinical symptoms and signs, positive tourniquet test (OR = 4.86), ascites (OR = 13.91) and shock (OR = 308.09) were identified as the best predictors of dengue infection, DHF and mortality, respectively (both P < 0.05). The main risk factors for dengue infection, DHF and mortality were living with uncovered water container (OR = 1.65), suffering from hypotension (OR = 6.18) and suffering from diabetes mellitus (OR = 2.53), respectively (all P < 0.05). The serotype distribution varied with time and across WHO regions. Overall, co-infections were reported in 47.7% of the evaluated outbreaks, and the highest pooled mortality rate (2.0%) was identified in DENV-2 dominated outbreaks. Our study emphasizes the necessity of implementing programs focused on targeted prevention, early identification, and effective treatment.