A Novel lncRNA lncRNA-4045 Promotes the Progression of Hepatocellular Carcinoma by Affecting the Expression of AKR1B10.

BACKGROUND: Long noncoding RNAs (lncRNAs) have been shown to be related to the occurrence and development of a variety of cancers including hepatocellular carcinoma (HCC). However, a large number of potential HCC-related lncRNAs remain undiscovered and are yet to be fully understood. METHODS: Differentially expressed lncRNAs were first obtained from the tumor tissues and adjacent normal tissues of five HCC patients using high-throughput microarray chips. Then the expression levels of 10 differentially expressed lncRNAs were verified in 50 pairs of tissue samples from patients with HCC by quantitative real-time PCR (qRT-PCR). The oncogenic effects of lncRNA-4045 (ENST00000524045.6) in HCC cell lines were verified through a series of in vitro experiments including CCK-8 assay, plate clone formation assay, transwell assay, scratch assay, and flow cytometry. Subsequently, the potential target genes of lncRNA-4045 were predicted by bioinformatics analysis, fluorescence in situ hybridization assay, and RNA sequencing. The mechanism of lncRNA-4045 in HCC was explored by WB assay as well as rescue and enhancement experiments. RESULTS: The results from microarray chips showed 1,708 lncRNAs to have been significantly upregulated and 2725 lncRNAs to have been significantly downregulated in HCC tissues. Via validation in 50 HCC patients, a novel lncRNA lncRNA-4045 was found significantly upregulated in HCC tissues. Additionally, a series of in vitro experiments showed that lncRNA-4045 promoted the proliferation, invasion, and migration of HCC cell lines, and inhibited the apoptosis of HCC cell lines. The results of qRT-PCR in HCC tissues showed that the expression levels of AKR1B10 were significantly positively correlated with lncRNA-4045. LncRNA-4045 knockdown significantly down-regulated AKR1B10 protein expression, and overexpression of lncRNA-4045 led to significant up-regulation of AKR1B10 protein in HCC cell lines. Lastly, down-regulation of AKR1B10 could partially eliminate the enhancement of cell proliferation induced by lncRNA-4045 overexpression, while up-regulation of AKR1B10 was shown to enhance those effects. CONCLUSION: LncRNA-4045 may promote HCC via enhancement of the expression of AKR1B10 protein.

Liver-specific LINC01146, a promising prognostic indicator, inhibits the malignant phenotype of hepatocellular carcinoma cells both in vitro and in vivo.

BACKGROUND: Long non-coding RNAs (lncRNAs) are involved in the development of hepatocellular carcinoma (HCC). We aimed to investigate the function of LINC01146 in HCC. METHODS: The expression of LINC01146 in HCC tissues was explored via The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases and was verified using quantitative real-time polymerase chain reaction (qRT-PCR) in our HCC cohort. Kaplan-Meier analysis was used to assess the relationship between LINC01146 and the prognosis of HCC patients. Cell Counting Kit 8, colony formation assays, Transwell assays, flow cytometric assays, and tumour formation models in nude mice were conducted to reveal the effects of LINC01146 on HCC cells both in vitro and in vivo. Bioinformatic methods were used to explore the possible potential pathways of LINC01146 in HCC. RESULTS: LINC01146 was significantly decreased in HCC tissues compared with adjacent normal tissues and was found to be related to the clinical presentations of malignancy and the poor prognosis of HCC patients. Overexpression of LINC01146 inhibited the proliferation, migration, and invasion of HCC cells in vitro, while promoting their apoptosis. In contrast, downregulation of LINC01146 exerted the opposite effects on HCC cells in vitro. In addition, overexpression of LINC01146 significantly inhibited tumour growth, while downregulation of LINC01146 promoted tumour growth in vivo. Furthermore, the coexpressed genes of LINC01146 were mainly involved in the "metabolic pathway" and "complement and coagulation cascade pathway". CONCLUSION: LINC01146 expression was found to be decreased in HCC tissues and associated with the prognosis of HCC patients. It may serve as a cancer suppressor and prognostic biomarker in HCC.

Liver-specific lncRNA FAM99A may be a tumor suppressor and promising prognostic biomarker in hepatocellular carcinoma.

BACKGROUND: Increasing evidence shows that liver-specific long non-coding RNAs (lncRNAs) play important roles in the development of hepatocellular carcinoma (HCC). We identified a novel liver-specific lncRNA, FAM99A, and examined its clinical significance and biological functions in HCC. METHODS: The expression level and clinical value of FAM99A in HCC were examined using The Cancer Genome Atlas (TCGA), International Cancer Genome Consortium (ICGC), and Gene Expression Omnibus (GEO) databases, and were further verified using quantitative real-time polymerase chain reaction (qRT-PCR) in our HCC cohort. Univariate and multivariate Cox proportional hazards regression models were also applied to identify independent prognostic indicators for HCC patients. Cell counting kit-8, colony formation, and Transwell assays were performed to evaluate the effects of FAM99A on the proliferation, migration, and invasion abilities of HCC cells in vitro. A subcutaneous xenograft tumor model was implemented to determine the effect of FAM99A on the tumor growth of HCC cells in vivo. RNA pull-down and mass spectrometry assays were performed to reveal the potential molecular mechanisms of FAM99A in HCC. RESULTS: The three public online databases and qRT-PCR data showed that FAM99A was frequently downregulated in HCC tissues and inversely correlated with microvascular invasion and advanced histological grade of HCC patients. Kaplan-Meier survival analysis indicated that decreased FAM99A was significantly associated with poor overall survival of HCC patients based on TCGA database (P = 0.040), ICGC data portal (P < 0.001), and our HCC cohort (P = 0.010). A multivariate Cox proportional hazards regression model based on our HCC cohort suggested that FAM99A was an independent prognostic factor of overall survival for HCC patients (hazard ratio: 0.425, P = 0.039). Upregulation of FAM99A suppressed the proliferation, colony formation, migration, and invasion capacities of HCC cells in vitro, and knockdown of FAM99A had the opposite effects. A subcutaneous xenograft tumor model demonstrated that overexpression of FAM99A significantly inhibited the tumor growth of HCC cells in vivo. Seven tumor-related proteins (PCBP1, SRSF5, SRSF6, YBX1, IGF2BP2, HNRNPK, and HNRNPL) were recognized as possible FAM99A-binding proteins by the RNA pull-down and mass spectrometry assays. CONCLUSION: Our results suggest that FAM99A exerts cancer-inhibiting effects on HCC progression, and it may be a promising prognostic indicator for HCC patients.

Maternal overweight but not paternal overweight before pregnancy is associated with shorter newborn telomere length: evidence from Guangxi Zhuang birth cohort in China.

BACKGROUND: Telomere length (TL) is variable at birth and is inversely associated with body mass index (BMI) in adulthood. A growing number of evidences suggested that a higher maternal pre-pregnancy BMI results in adverse offspring health outcomes, especially shorter newborn TL. However, a newborn's genetic endowment is equally derived from both parents, the association between parental pre-pregnancy BMI and newborn TL has been rarely discussed. We aimed to determine the association between parental pre-pregnancy BMI and newborn TL. METHODS: A total of 1082 parent-newborn pairs were recruited from the Guangxi Zhuang Birth Cohort (GZBC). TL in cord blood was measured using quantitative real-time polymerase chain reaction (qPCR) and expressed as the ratio of telomere copy number to single-copy gene number (T/S). A series of linear regressions were performed to assess the associations between parental pre-pregnancy BMI and newborn TL. RESULTS: Mothers who were overweight before pregnancy had significantly shorter cord blood telomere length in their newborns than those who were normal weight before pregnancy [percentage change: - 7.96% (95% CI: - 14.49 to - 0.69%; P = 0.032)]. Further analysis of the combined effects of parental weight status on newborn TL showed that TL was significantly shortened among newborns whose mothers were overweight and fathers were of healthy weight when compared with those whose mothers and fathers were both of normal weight [percentage change: - 8.38% (95% CI: - 15.47 to - 0.92%; P = 0.028)]. Subgroup analysis indicated these effects were more pronounced among male newborns and those whose paternal age < 31 years or maternal age ≥ 28 years at delivery. CONCLUSIONS: Maternal pre-pregnancy overweight, but not paternal pre-pregnancy overweight is associated with shorter newborn TL. Weight control in reproductive women and effective healthy weight management before pregnancy may be of particular benefit for improving longevity and life quality of offspring.

Associations of bisphenol exposure with the risk of gestational diabetes mellitus: a nested case-control study in Guangxi, China.

A growing number of epidemiologic studies have estimated the associations between endocrine-disrupting chemicals and gestational diabetes mellitus (GDM). However, reports on the association between bisphenol A (BPA) substitutes and GDM are limited. This investigation aimed to explore the associations of maternal serum BPA, bisphenol B (BPB), bisphenol F (BPF), bisphenol S (BPS), and tetrabromobisphenol A (TBBPA) with the risk of GDM. A nested case-control study was performed among 500 pregnant women. In conditional logistic regression models, the OR for BPS was significantly increased in the medium exposure groups (OR = 1.77; 95% CI: 1.01, 3.13) compared with the reference group, while BPA (OR: 0.38, 95%CI: 0.29, 0.50) and TBBPA (OR: 0.67, 95%CI: 0.54, 0.85) were negatively associated with the risk of GDM. In the Bayesian kernel machine regression (BKMR) analysis, the joint effect of bisphenols was positively associated with the risk of GDM. BPS showed positively relationship, while BPA and TBBPA showed negatively relationship, respectively. The quantile g-computation revealed a statistically significant and negative joint effect of the five bisphenols on the risk of GDM (OR: 0.57; 95% CI: 0.46, 0.72) with BPA (70.2%), TBBPA (21.3%), and BPB (8.5%) had positive contribution to the overall effect. These findings suggested that BPS had a positive effect on the risk of GDM, while BPA and TBBPA had negative effect on the risk of GDM. Moreover, exposure to the mixture of the five bisphenols was negatively associated with the risk of GDM.

Construction and Comprehensive Analysis of a circRNA-miRNA-mRNA Regulatory Network to Reveal the Pathogenesis of Hepatocellular Carcinoma.

Background: Circular RNAs (circRNAs) have been demonstrated to be closely related to the carcinogenesis of human cancer in recent years. However, the molecular mechanism of circRNAs in the pathogenesis of hepatocellular carcinoma (HCC) has not been fully elucidated. We aimed to identify critical circRNAs and explore their potential regulatory network in HCC. Methods: The robust rank aggregation (RRA) algorithm and weighted gene co-expression network analysis (WGCNA) were conducted to unearth the differentially expressed circRNAs (DEcircRNAs) in HCC. The expression levels of DEcircRNAs were validated by quantitative real-time polymerase chain reaction (qRT-PCR). A circRNA-miRNA-mRNA regulatory network was constructed by computational biology, and protein-protein interaction (PPI) network, functional enrichment analysis, survival analysis, and infiltrating immune cells analysis were performed to uncover the potential regulatory mechanisms of the network. Results: A total of 22 DEcircRNAs were screened out from four microarray datasets (GSE94508, GSE97332, GSE155949, and GSE164803) utilizing the RRA algorithm. Meanwhile, an HCC-related module containing 404 circRNAs was identified by WGCNA analysis. After intersection, only four circRNAs were recognized in both algorithms. Following qRT-PCR validation, three circRNAs (hsa_circRNA_091581, hsa_circRNA_066568, and hsa_circRNA_105031) were chosen for further analysis. As a result, a circRNA-miRNA-mRNA network containing three circRNAs, 17 miRNAs, and 222 mRNAs was established. Seven core genes (ESR1, BUB1, PRC1, LOX, CCT5, YWHAZ, and DDX39B) were determined from the PPI network of 222 mRNAs, and a circRNA-miRNA-hubgene network was also constructed. Functional enrichment analysis suggested that these seven hub genes were closely correlated with several cancer related pathways. Survival analysis revealed that the expression levels of the seven core genes were significantly associated with the prognosis of HCC patients. In addition, we also found that these seven hub genes were remarkably related to the infiltrating levels of immune cells. Conclusion: Our research identified three pivotal HCC-related circRNAs and provided novel insights into the underlying mechanisms of the circRNA-miRNA-mRNA regulatory network in HCC.

Exposure to phthalates in early pregnancy and the risk of fetal growth restriction: a nested case-control study in a Zhuang Chinese population.

Phthalates (PAEs) are common endocrine disrupting chemicals (EDCs) that disrupt fetal development. The present study aimed to evaluate the effects of single and coexposure to phthalates in early pregnancy on fetal growth restriction (FGR) by a nested case-control study based on the Guangxi Zhuang Birth Cohort (GZBC). Maternal serum concentrations of seven phthalates in 97 neonates with FGR and 291 matched controls were detected through gas chromatography-mass spectrometry (GC-MS). The associations between phthalates and FGR were analyzed using multiple logistic regression, weight quantile sum (WQS) regression, and Bayesian kernel machine regression (BKMR) models. We found that exposures to butyl-benzyl phthalate (BBP, ORadj = 1.849, 95% CI: 1.080-3.177, Padj = 0.025, Ptrend = 0.046), di (2-ethyl-hexyl) phthalate (DEHP, ORadj = 3.893, 95% CI: 1.305-11.910, Padj = 0.015, Ptrend = 0.098) and dimethyl phthalate (DMP, ORadj = 1.722, 95% CI: 1.089-2.725, Padj = 0.020, Ptrend = 0.002) were significantly positively associated with the risk of FGR, while mono-butyl phthalate (MBP) showed a significant negative association with FGR (ORhigh = 0.192, 95% CI: 0.036-0.795, Padj = 0.033, Ptrend = 0.035) only among girls. The WQS model identified that BBP, di(2-ethyl)phthalate (DEP), DMP, DEHP, di-n-butyl phthalate (DBP), and MBP were highly weighted in the association with FGR. The BKMR model supported the positive association between joint exposure to phthalates and the risk of FGR and identified no significant interaction between the seven phthalates. Overall, maternal exposure to BBP, DEHP, and DMP may cause adverse effects on FGR, especially with combined effects.

Exposure to perfluoroalkyl substances in early pregnancy and the risk of hypertensive disorders of pregnancy: A nested case-control study in Guangxi, China.

BACKGROUND: Previous epidemiological studies have examined the associations between exposure to perfluoroalkyl substances (PFASs) and the risk of hypertensive disorders of pregnancy (HDP). However, these studies have drawn discrepant conclusions and have some limitations. METHODS: A nested case-control study was conducted with the Guangxi Zhuang Birth Cohort (GZBC), a prospective, ongoing birth cohort that was implemented in Guangxi, China, in June 2015. Maternal serum concentrations of nine PFASs were measured using ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS). The associations between PFAS exposure and the risk of HDP were assessed using logistic regression (single-exposure), weighted quantile sum (WQS) regression, and Bayesian kernel machine regression (BKMR) models. RESULTS: A total of 136 HDP cases and 408 controls were enrolled in this study. In logistic regression models, perfluoroundecanoic acid (PFUnA), perfluorooctane sulfonate (PFOS), perfluorononanoic acid (PFNA) and perfluorobutanesulfonic acid (PFBS) were positively associated with HDP, while perfluorohexane sulfonate (PFHxS) was inversely associated with HDP. In the BKMR analysis, the joint effect of PFASs was positively associated with HDP. PFOS and PFBS showed positive trends, while PFHxS and PFHpA showed inverse trends. In WQS regression analysis, we calculated two WQS indices that were estimated using constraints in both the positive and negative directions of effects. Both WQS indices were significantly associated with HDP (OR: 2.663, 95% CI: 1.795-3.951; OR: 0.338, 95% CI: 0.229-0.499, respectively). PFBS, PFOS and PFUnA had significant weights in the positive effect direction; PFHxS, perfluoroheptanoic acid (PFHpA) and perfluorododecanoic acid (PFDoA) had significant weights in the negative effect direction. CONCLUSION: Considering all model results, we found that combined exposure to nine PFASs had a positive effect on the development of HDP. Moreover, PFOS and PFBS were positively associated with the HDP risk, while PFHxS and PFHpA were negatively associated with the HDP risk in women in Guangxi, China.

Association of single and multiple prefluoroalkyl substances exposure with preterm birth: Results from a Chinese birth cohort study.

BACKGROUND: Perfluoroalkyl substances (PFASs) are persistent organic pollutants that may lead the adverse birth outcomes, including preterm birth (PTB). However, previous studies have reported inconsistent results on the association between PFASs and PTB, and lack of the epidemiological evidence regarding the effect of PFASs mixture on PTB. This study aimed to explore association of individual and multiple exposure to PFASs with PTB. METHODS: The study subjects were consisted of 1341 pregnant women from Guangxi Zhuang Birth Cohort in Guangxi, China, from June 2015 to April 2019. Nine PFASs concentrations in the maternal serum were examined by ultrahigh liquid performance chromatography-tandem mass spectrometry, and the gestational weeks were obtained from medical records. We applied binary logistics regression model to explore correlation between individual PFAS and PTB and inspected the combined effect of PFASs mixture on PTB by applying Bayesian kernel machine regression (BKMR) and weighted quantile sum (WQS) regression models. RESULTS: In adjusted logistics regression model, perfluorooctane sulfonate (PFOS), perfluoroheptanoic acid (PFHpA), perfluorobutanesulfonic acid (PFBS), ∑perfluorinated sulfonic acids (PFSA), and ∑PFASs were positively associated with the risk of PTB. In contrast, perfluoroundecanoic acid (PFUnA), perfluorohexane sulfonate (PFHxS), and perfluorooctanoic acid (PFOA) were negatively associated with the risk of PTB. These associations of n PFOS and PFHpA with PTB were found to be more pronounced in male infants. Restricted cubic splines (RCSs) showed an inverse U-shaped relationship between PFBS and PTB. Analysis from BKMR model showed a positive association between PFASs mixture and PTB, and no evidence of interactions among the nine PFASs were detected. Additionally, PFHpA, PFOS, and PFBS were identified as the main contributors for the effect of PFASs mixture on increasing the risk of PTB by BKMR and WQS models. CONCLUSION: Prenatal exposure to higher levels of PFASs mixture was associated with higher risk of PTB.

Association of serum per- and polyfluoroalkyl substances and gestational anemia during different trimesters in Zhuang ethnic pregnancy women of Guangxi, China.

BACKGROUND: Gestational anemia is a complication of pregnancy, and a low level of hemoglobin (Hb) has been linked to adverse pregnancy outcomes. Previous studies reported that PFASs were more strongly associated with Hb than red blood cells, indicating that Hb is more susceptible to the effect of PFASs. However, the evidences regarding the effects of per- and polyfluoroalkyl substances (PFASs) on gestational anemia are currently limited. Therefore, it is important to explore the effects of PFASs on anemia in Chinese pregnant women. METHODS: A total of 821 pregnant women were recruited between June 2015 and April 2019 in the Guangxi Zhuang Birth Cohort. The concentrations of PFASs were assessed in maternal serum before 12 gestational weeks. To determine both individual and combined associations of PFASs exposure with anemia in the three stages of pregnancy, binary logistic regression, Bayesian kernel machine regression (BKMR), and weighted quantile sum (WQS) regression models were employed. RESULTS: In single-pollutant analysis, maternal exposure to perfluorododecanoic acid (PFDoA) and perfluoroheptanoic acid (PFHpA) were associated with anemia in the first trimester, exposure to PFHpA and perfluorobutanesulfonic acid (PFBS) were associated with anemia in the second trimester, and exposure to perfluorodecanoic acid (PFDA) and perfluorononanoic acid (PFNA) were associated with anemia in the third trimester. Notably, perfluoroundecanoic acid (PFUnA) had a nonlinear association with anemia in the third trimester. In multiple-pollutant analysis, a positive association of PFDoA with anemia in the first trimester and a negative association of PFBS with anemia in the second trimester were confirmed by BKMR. Exposure to PFASs mixture was not associated with anemia in all three trimesters. In WQS, there was a significantly negative association between the PFAS mixture and anemia in the second trimester. CONCLUSION: Maternal exposure to PFASs is associated with gestational anemia in different trimesters.

Associations between prenatal multiple plasma metal exposure and newborn telomere length: Effect modification by maternal age and infant sex.

Exposure to metals during pregnancy may affect maternal and infant health. However, studies on the combined effects of metals on the telomere length (TL) of newborns are limited. A prospective cohort study was conducted among 1313 mother-newborn pairs in the Guangxi Zhuang Birth Cohort. The concentrations of metals in maternal plasma during the first trimester were measured using inductively coupled plasma-mass spectrometry. We explored the associations between nine plasma metals and newborn TL using generalized linear models (GLMs), principal component analysis (PCA), quantile g-computation (qgcomp), and Bayesian kernel machine regression (BKMR). The GLMs revealed the inverse association between plasma arsenic (percent change, -5.56%; 95% CI: -7.69%, -3.38%) and barium concentrations (-9.84%; 95% CI: -13.81%, -5.68%) and newborn TL. Lead levels were related to significant decreases in newborn TL only in females. The PCA revealed a negative association between the PC3 and newborn TL (-4.52%; 95% CI: -6.34%, -2.68%). In the BKMR, the joint effect of metals was negatively associated with newborn TL. Qgcomp indicated that each one-tertile increase in metal mixture levels was associated with shorter newborn TL (-9.39%; 95% CI: -14.32%, -4.18%). The single and joint effects of multiple metals were more pronounced among pregnant women carrying female fetuses and among pregnant women <28 years of age. The finding suggests that prenatal exposure to arsenic, barium, antimony, and lead and mixed metals may shorten newborn TLs. The relationship between metal exposures and newborn TL may exhibit heterogeneities according to infant sex and maternal age.