SIRT4 in ageing.

Ageing is a phenomenon in which cells, tissues and organs undergo systemic pathological changes as individuals age, leading to the occurrence of ageing-related diseases and the end of life. It is associated with many phenotypes known as ageing characteristics, such as genomic instability, nutritional imbalance, mitochondrial dysfunction, cell senescence, stem cell depletion, and an altered microenvironment. The sirtuin family (SIRT), known as longevity proteins, is thought to delay ageing and prolong life, and mammals, including humans, have seven family members (SIRT1-7). SIRT4 has been studied less among the sirtuin family thus far, but it has been reported that it has important physiological functions in organisms, such as promoting DNA damage repair, participating in the energy metabolism of three substances, inhibiting inflammatory reactions and apoptosis, and regulating mitochondrial function. Recently, some studies have demonstrated the involvement of SIRT4 in age-related processes, but knowledge in this field is still scarce. Therefore, this review aims to analyse the relationship between SIRT4 and ageing characteristics as well as some age-related diseases (e.g., cardiovascular diseases, metabolic diseases, neurodegenerative diseases and cancer).

A novel fungal negative-stranded RNA virus related to mymonaviruses in Auricularia heimuer.

Here, we report the characterization of a novel (-)ssRNA mycovirus isolated from Auricularia heimuer CCMJ1222, using a combination of RNA-seq, reverse transcription polymerase chain reaction, 5' and 3' rapid amplification of cDNA ends, and Sanger sequencing. Based on database searches, sequence alignment, and phylogenetic analysis, we designated the virus as "Auricularia heimuer negative-stranded RNA virus 1" (AhNsRV1). This virus has a monopartite RNA genome related to mymonaviruses (order Mononegavirales). The AhNsRV1 genome consists of 11,441 nucleotides and contains six open reading frames (ORFs). The largest ORF encodes a putative RNA-dependent RNA polymerase; the other ORFs encode hypothetical proteins with no conserved domains or known function. AhNsRV1 is the first (-)ssRNA virus and the third virus known to infect A. heimuer.

LncRNA NEAT1 promoted MPP+­induced ferroptosis via regulating miR­150­5p/BAP1 pathway in SK­N­SH cells.

As widely reported, dysregulated ferroptosis is closely associated with Parkinson's disease (PD) progression. The goal of the present study was to probe the roles of long non­coding RNA (lncRNA) nuclear enriched assembly transcript 1 (NEAT1) in regulating ferroptosis in PD. PD cell model was constructed by subjecting SK­N­SH cells to 1­methyl­4­phenylpyridinium (MPP+) for 24 h. The RNA levels of NEAT1, miRNA (miR)­150­5p, and BRCA1­associated protein 1 (BAP1) were evaluated using qRT­PCR. The protein levels of glutathione peroxidase 4 (GPX4), BAP1, and solute carrier family 7 member 11 (SLC7A11) were determined using western blot. Cell viability was assessed using 3­(4,5­dimethylthiazolyl2)­2, 5­diphenyltetrazolium bromide (MTT) assay. In addition, fluorescent probe 2,7­dichlorodihydrofluorescein diacetate (DCFH­DA) was employed to determine the ROS level. Moreover, the levels of GSH, MDA, and Fe2+ were also measured. Finally, the interactions among NEAT1, miR­150­5p, and BAP1 were identified by dual luciferase reporter gene assay, and/or RIP assay. Upregulated NEAT1 was observed in PD cell model. Knockdown of NEAT1 elevated viability and GSH level in PD cell model and reduced ROS, MDA, and Fe2+ levels. Moreover, NEAT1 functioned as a sponge to suppress miR­150­5p expression. Moreover, miR­150­5p overexpression suppressed ferroptosis in PD cell model. We subsequently found that miR­150­5p regulated SLC7A11 expression by directly binding to BAP1. miR­150­5p inhibition or BAP1 overexpression mitigated the anti­ferroptosis effect meditated by sh­NEAT1. Taken together, knockdown of NEAT1 mitigated MPP+­induced ferroptosis through regulating BAP1/SLC7A11 axis by sponging miR­150­5p, indicating the potential of NEAT1 as a promising therapeutic target for PD.

A double-blinded, placebo-controlled randomized trial evaluating the efficacy and safety of Zhigancao Tang granules for treating HFpEF: study protocol for a randomized controlled trial.

BACKGROUND: Heart failure with preserved ejection fraction (HFpEF) is a clinical syndrome characterized by diastolic dysfunction. Despite the increasing incidence of HFpEF, there is no available therapy that reduces the mortality rate of HFpEF. Zhigancao Tang has been used traditionally for the treatment of cardiovascular diseases in China. The use of traditional Chinese medicine (TCM) is associated with improvements in clinical syndromes and quality of life of patients. A randomized clinical trial should be conducted to provide clear evidence regarding the efficacy and safety of Zhigancao Tang granules for the treatment of HFpEF. METHODS: A randomized, double-blinded, placebo-controlled clinical trial was proposed. A total of 122 patients with HFpEF will be randomly assigned to receive Zhigancao Tang granules or placebo for 12 weeks. The primary outcome measure is cardiac function. The secondary outcomes include measurement of the integral TCM syndrome score, echocardiography, 6-min walk test, N-terminal-pro hormone B-type natriuretic peptide level, atrial natriuretic peptide level, Minnesota Living with Heart Failure scale, and Lee's scale. The outcome measures will be evaluated at baseline, 4 weeks, and 12 weeks. Adverse events will be evaluated from baseline till the 12-week follow-up period. DISCUSSION: The results of this trial will demonstrate whether Zhigancao Tang granules are effective and safe for treating HFpEF. TRIAL REGISTRATION: ClinicalTrials.gov NCT04317339 . Registered on 23 March 2020.

A Biomimetic Zinc Alloy Scaffold Coated with Brushite for Enhanced Cranial Bone Regeneration.

Bone tissue engineering is considered as a promising pathway for bone regeneration and defect reconstruction, in which scaffolds play an important role. Zn alloy, which is a biodegradable metal material that has advantages of metallic and biodegradable characteristics, has its special features, especially the ideal degradation rate and acceptable biocompatibility, which make it worthy to be further investigated for medical applications. In this study, new biodegradable porous Zn alloy scaffolds with Ca-P coating were attempted to repair cranial bone defect, and in vitro and in vivo assays were conducted to evaluate its biocompatibility, osteo-inductivity, and osteo-conductivity. The results indicated that coated Zn alloy possessed good biocompatibility, with no cytotoxicity. It could also promote osteogenic differentiation and calcium deposition of rabbit BMSCs in vitro, and new bone formation around the scaffold in vivo. The biodegradable porous Zn alloy scaffold with Ca-P coating is considered to be promising in cranial bone defect repair.

Challenges and strategies for in situ endothelialization and long-term lumen patency of vascular grafts.

Vascular diseases are the most prevalent cause of ischemic necrosis of tissue and organ, which even result in dysfunction and death. Vascular regeneration or artificial vascular graft, as the conventional treatment modality, has received keen attentions. However, small-diameter (diameter < 4 mm) vascular grafts have a high risk of thrombosis and intimal hyperplasia (IH), which makes long-term lumen patency challengeable. Endothelial cells (ECs) form the inner endothelium layer, and are crucial for anti-coagulation and thrombogenesis. Thus, promoting in situ endothelialization in vascular graft remodeling takes top priority, which requires recruitment of endothelia progenitor cells (EPCs), migration, adhesion, proliferation and activation of EPCs and ECs. Chemotaxis aimed at ligands on EPC surface can be utilized for EPC homing, while nanofibrous structure, biocompatible surface and cell-capturing molecules on graft surface can be applied for cell adhesion. Moreover, cell orientation can be regulated by topography of scaffold, and cell bioactivity can be modulated by growth factors and therapeutic genes. Additionally, surface modification can also reduce thrombogenesis, and some drug release can inhibit IH. Considering the influence of macrophages on ECs and smooth muscle cells (SMCs), scaffolds loaded with drugs that can promote M2 polarization are alternative strategies. In conclusion, the advanced strategies for enhanced long-term lumen patency of vascular grafts are summarized in this review. Strategies for recruitment of EPCs, adhesion, proliferation and activation of EPCs and ECs, anti-thrombogenesis, anti-IH, and immunomodulation are discussed. Ideal vascular grafts with appropriate surface modification, loading and fabrication strategies are required in further studies.

Preliminary study on mechanical characteristics of maxillofacial soft and hard tissues for virtual surgery.

PURPOSE: Virtual surgery system can provide us a realistic and immersive training environment, in which haptic force-feedback gives operators 'touching feeling.' Appropriate deformation models of soft and hard tissues are required for the achievement of real-time haptic feedback. To improve accuracy of modeling and haptic feedback simulation for maxillofacial virtual surgery, mechanical characteristics of soft and hard tissues should be explored. METHODS: Craniofacial soft tissues from one male and female cadavers were divided into two layers: skin and muscle. Maxillofacial tissues were divided into frontal, chin, temporalis, masseter regions. Insertion and cutting process were conducted using VMX42 5-axis linkage system and recorded by piezoelectric dynamometer. Maximum stiffness values were analyzed, and insertion curves before puncture were fitted using a polynomial model. Elasticity modulus and hardness of maxillofacial hard tissues were measured and analyzed using Berkovich nanoindentation. RESULTS: Tissues in different maxillofacial regions, as well as from different layers (skin and muscle), displayed various mechanical performance. Maximum stiffness values and cutting force of soft tissues in male and female had significant difference. The third-order polynomial was demonstrated to fit the insertion curves well before puncture. Furthermore, elasticity modulus and hardness of enamel were significantly greater than that of zygoma, maxilla and mandible. CONCLUSION: Mechanical properties of hard tissues are relatively stable, which can be applied in virtual surgery system for physical model construction. Insertion model and cutting force for soft tissues are meaningful and applicable and can be utilized to promote the accuracy of response for haptic feedback sensations.

Cytochalasin D Promotes Osteogenic Differentiation of MC3T3-E1 Cells via p38-MAPK Signaling Pathway.

BACKGROUND: Bone defect caused by trauma, tumor resection, infection or congenital malformation is a common clinical disease. Bone tissue engineering is regarded as a promising way of bone defect reconstruction. Thus, agents that can promote osteogenesis have received great attention. Cytochalasin D (Cyto D), a metabolite derived from molds, proves to be able to modify actin, reorganize cytoskeleton, and then promote the osteogenic differentiation. OBJECTIVE: The purpose of this study was to explore the effect and mechanism of Cyto D on osteogenic differentiation of mouse pre-osteoblast MC3T3-E1 cells. METHODS: The optimum concentration of Cyto D was explored. The osteogenic differentiation of MC3T3-E1 cells induced by Cyto D was assessed by alkaline phosphatase (ALP) staining, Alizarin Red S (ARS) staining, western blotting and quantitative real-time polymerase chain reaction (RT-qPCR). In addition, a specific pathway inhibitor was utilized to explore whether MAPK pathways were involved in this process. RESULTS: The results showed that the optimized concentration of action was 10-2µg/ml. The expression of Runx2, OCN and OSX was up-regulated by the supplement of Cyto D. ALP activity, calcium deposition, and phosphorylation level of p38 protein were also improved. Inhibition of the pathway significantly reduced the activation of p38, and the expression of osteogenic-related genes. CONCLUSION: Cyto D can promote the osteogenic differentiation of MC3T3 cells via the p38-MAPK signaling pathway, but not the ERK1/2 or JNK, and it is a potential agent to improve the osteogenesis of MC3T3 cells.

Adsorption of Cd(II) from aqueous solutions by rape straw biochar derived from different modification processes.

In order to deal with cadmium (Cd(II)) pollution, three modified biochar materials: alkaline treatment of biochar (BC-NaOH), KMnO4 impregnation of biochar (BC-MnOx) and FeCl3 magnetic treatment of biochar (BC-FeOx), were investigated. Nitrogen adsorption-desorption isotherms, Fourier transform infrared spectroscopy (FTIR), Boehm titration, and scanning electron microscopy (SEM) were used to determine the characteristics of adsorbents and explore the main adsorption mechanism. The results show that manganese oxide particles are carried successfully within the biochar, contributing to micropore creation, boosting specific surface area and forming innersphere complexes with oxygen-containing groups, while also increasing the number of oxygen-containing groups. The adsorption sites created by the loaded manganese oxide, rather than specific surface areas, play the most important roles in cadmium adsorption. Batch adsorption experiments demonstrate a Langmuir model fit for Cd(II), and BC-MnOx provided the highest sorption capacity (81.10 mg g-1). The sorption kinetics of Cd(II) on adsorbents follows pseudo-second-order kinetics and the adsorption rate of the BC-MnOx material was the highest (14.46 g (mg·h)-1). Therefore, biochar modification methods involving KMnO4 impregnation may provide effective ways of enhancing Cd(II) removal from aqueous solutions.

[Effect of Modified Biochars on Soil Cadmium Stabilization in Paddy Soil Suffered from Original or Exogenous Contamination].

To investigate the passivation of different modified biochars on the speciation and availability of cadmium contaminated soil, the modified biochars were treated by different approaches (acid/base treatment, impregnation with manganese oxides, magnetic modification) and biochars(BC) were used as soil passivating agents for soil culture experiments. The result indicated that the content of available cadmium decreased significantly by BC and modified biochars in originally contaminated soil. Compared with CK, the percentage of available cadmium in originally contaminated soil was reduced by more than 50% using impregnable biochars by KMnO4(BC-KMnO4) and basic biochars by NaOH(BC-NaOH). And the content of available cadmium decreased significantly by three modified biochars which were BC-KMnO4, BC-NaOH and FeCl3 magnetization biochars(BC-FeCl3) in exogenously contaminated soil. Particularly, the best performance was observed with BC-KMnO4 that reduced 30% available cadmium in exogenously contaminated soil. However, the passivation of BC was not significant, and the content of available cadmium slightly increased (3.8%-24.5%) by BC-HNO3 in exogenously contaminated soil. Furthermore, the content of exchangeable cadmium was increased by 20.2% with 2.5% BC-HNO3 in exogenously contaminated soil, while significantly decreased by other modified biochars and BC, and the 10% BC-KMnO4 reduced 65.1% exchangeable cadmium in originally contaminated soil. Meanwhile, soil pH was increased significantly by BC, BC-KMnO4 and BC-NaOH, while was reduced by BC-HNO3. The contents of organic carbon and exchangeable base cations in soil were improved by all the treatments. The results of regression analysis showed that the content of available cadmium in originally contaminated soil was significantly negatively correlated with soil pH, soil exchangeable Na+, while the content of available cadmium in exogenously contaminated soil was significantly negatively correlated with soil pH, soil organic carbon, soil exchangeable Mg2+, Na+, K+. Accordingly, the lower available cadmium in contaminated soil may correlate with the increasing content of organic carbon, exchangeable base cations and pH. In summary, the materials of BC-KMnO4 could be used as a superior passivating agent for in situ remediation of cadmium pollution, while the materials of BC-HNO3 could slightly activate cadmium in the soil, leading to some risk in in situ remediation.

[Study on sperm damage caused by trichloroethylene in male rats].

OBJECTIVE: To study in vitro sperm damage caused by trichloroethylene in male rats. METHODS: Sperms of Sprague-Dawley (SD) rats were collected 4 hours after being contaminated by trichloroethylene of 0, 2, 4, 6, 8, and 10 mmol/L in vitro. Giemsa staining was performed to observe the morphological changes of sperms, and flow cytometer was used to detect the changes in mitochondrial membrane potential. RESULTS: The sperm motilities in 6, 8, and 10 mmol/L trichloroethylene groups decreased significantly compared with that in control group (P <0.01); the sperm aberration rates in 8 and 10 mmol/L trichloroethylene groups were significantly higher than that in control group (P<0.01). With the increase in exposure dose, the proportion of sperms with reduced mitochondrial membrane potential increased, and there were significant differences in sperm apoptosis rate between the 4, 6, 8, and 10 mmol/L trichloroethylene groups and control group (P<0.01). CONCLUSION: In vitro exposure to trichloroethylene can reduce sperm motility and increase the aberration rate and apoptosis rate of sperms in male SD rats.

[Effects of bisphenol A on OCT4 and SOX2 genes expression in mouse embryonic stem cells].

OBJECTIVE: To explore the effects of bisphenol A (BPA) exposure on toxicity characteristic and OCT4 and SOX2 gene expression of mouse embryonic stem cells (mESC). METHODS: mESC were cultured, and treated with the doses of 10(-8), 10(-7), 10(-6), 10(-5), 10(-4) mol/L respectively of BPA and DMSO (the solvent control group)for 24 hours, and three groups of cells were treated with the same method. The morphological changes of mESC in the control and exposure groups were observed through an inverted microscope. Cell counting kit 8 (CCK8) was used to detect the effects of BPA on proliferation of mESC, and based on the results, the half inhibitory concentration (IC50) was calculated. Real-time fluorescent quantitative polymerase chain reaction (RT-QPCR) and western blotting were used to detect the expression of OCT4 and SOX2. RESULTS: BPA had certain toxicity on mESC, the treatment of BPA significantly increased cell toxicity in a concentration-dependent manner, and the IC50 was 4.3×10(-4) mol/L, combined with the BPA exposure concentration of the environment and the related literature, eventually taking the five concentrations of 10(-8), 10(-7), 10(-6), 10(-5), 10(-4) mol/L as the experimental groups. The mESC morphology were effected after the treatment of BPA for 24 h, compared with the control group, the number of cells decreased, appearing some floating cells, and the cell cloning became irregular and differentiation in the higher concentration groups. The OCT4 mRNA expression level in the 10(-7) mol/L (1.146 ± 0.087), 10(-6) mol/L (1.156 ± 0.030), 10(-5) mol/L (1.158 ± 0.103) and the 10(-4) mol/L (1.374 ± 0.053) dose group were all significantly higher than the control group (1.000 ± 0.000) (t values were -2.384, -2.953, -3.203, -4.021 respectively, P value all < 0.05). Meanwhile, the SOX2 mRNA expression level in the 10(-4) mol/L (1.113 ± 0.052) were higher than the control group (1.000 ± 0.000) (t value was -2.765, P value < 0.05). Moreover, the OCT4 protein expression level in the 10(-5) mol/L (1.360 ± 0.168) and 10(-4) mol/L (1.602 ± 0.151) were all significantly higher than the control group (1.000 ± 0.000) (t values were -3.538, -4.002 respectively, P value all < 0.05), while no obvious change of the SOX2 protein expression level was detected in all treated groups. CONCLUSION: BPA in a certain dose range could upregulate the expression of OCT4 gene in mouse embryonic stem cells while had no significant effect on the expression of SOX2 gene.

[Genome DNA hypomethylation in HaCaT cells after short exposure to SiO2 nanoparticles].

OBJECTIVE: To observe the effect of SiO2 nanoparticles on genome DNA methylation profile in cultured cells. METHODS: HaCaT cells were treated with nm-SiO2 at 2.5, 5 and 10 microg/ml and micro-SiO2 at 10 microg/ml for 24h and DAC treatment was given at 10 microg/ml group for 48h. The mC/(mC + C) percent was quantified by high performance capillary electrophoresis (HPCE) assay, and the expression level of mRNA and protein was detected by Real-time Q-PCR and westernblot assay. The activity of DNMTs was determined by DNA Methyltransferase Activity/Inhibition Assay Kit. RESULTS: HPCE assay showed that nm-SiO2-treated cells were decreased in some degree. An average proportion of methylated mC/ (mC + C) was 4.82% in control, 2.7% in 2.5 microg/ml and 2.17% in 10 microg/ml groups, while 3.1% in micro-SiO2 groups, which got the consistent downtrend of genome methylation level during increasing nm-SiO2 dose nanoparticles. The mRNA expression level for DNMT1 decreased gradually with increased dose of nm-SiO2 nanoparticles. The alterations at protein level were similar to those at the mRNA level. CONCLUSION: Genomic DNA methylation levels were decreased in HaCaT cells after short-term exposure to SiO2 nanoparticles.

Effect of low dose bisphenol A on the early differentiation of human embryonic stem cells into mammary epithelial cells.

It has been previously reported that bisphenol A (BPA) can disturb the development of mammary structure and increase the risk of breast cancer in experimental animals. In this study, an in vitro model of human embryonic stem cell (hESC) differentiation into mammary epithelial cells was applied to investigate the effect of low dose BPA on the early stages of mammogenesis. A newly established hESC line was directionally differentiated into mammary epithelial cells by a well-established three-dimensional (3D) culture system. The differentiated mammary epithelial cells were characterized by immunofluorescence and western blotting assay, and were called induced differentiated mammary epithelial cells (iDMECs) based on these data. The hESCs were treated with low doses of BPA range 10(-9)-10(-6)M during the differentiation process, with DMSO as the solvent control and 17-ß-estrodiol (E2) as the estrogen-positive control. Our results showed that low dose BPA and E2 could influence the mammosphere area of iDMECs and upregulate the expression level of Oct4 and Nanog proteins, while only BPA could downregulate the expression of E-cadherin protein. Taken together, this study provides some insights into the effects of low dose BPA on the early differentiation stage of mammary epithelial cells and suggests an easier canceration status of iDMECs under the effect of low dose BPA during its early differentiation stage.

[Detection of trace uranium in air with field spectrometry].

As a natural radioactive element, uranium and its compounds exist as aerosol and transfer in air. In gas phase, uranium can cause various kinds of radioactive damage to human body. The change in its concentration in a local area is related to the exploration and utilization of nuclear energy. Therefore, the development of field method for rapid uranium detection in air sample is very important. In this contribution, the air samples over uranium ores collected by a general pump was absorbed with 2.0 mol x L(-1) nitrate and then reacted with solid reagent kit. When the reaction between trace uranium and chromogenic reagent was finished, the homemade portable photometer was used to measure the absorbance. The results showed that the concentration of uranium in air samples over low grade uranium ores can be successfully determined by the present method and the values agree with that obtained by ICP-MS. The RSD measured by the new method was 1.72%. The application of the new field spectrometry in discriminating uranium ores from other ores has the potential advantages of easy operation, cost-saving and high accuracy.

Methylation of PARP-1 promoter involved in the regulation of nano-SiO2-induced decrease of PARP-1 mRNA expression.

Nano silicon dioxide (nano-SiO2) is becoming more and more widely applied in the fields of industry. The potential toxic effects of nano-SiO2 and its hazard to human health are drawing more attention. The mRNA expression of poly(ADP-ribose) polymerases-1(PARP-1), a pivotal repair gene, has been decreased by nano-SiO2 exposure. However, the effect of epigenetic modification on nano-SiO2-induced low PARP-1 expression has not been reported. In this study, HaCaT cells with or without DNA methyltransferase 1(DNMT1) knock down were incubated with nano-SiO2 and then further treated with DNMT inhibitor, 5-aza-2-deoxycytidine (DAC), which is a kind of key epigenetic modification reagents. Real-time Q-PCR and western blotting were used to examine the mRNA and protein expression of PARP-1. For promoter methylation status of PARP-1, methylation-specific PCR (MSP) and Bisulfite sequencing assay were performed. Results showed a dramatic decrease of PARP-1 expression on mRNA and protein level and a simultaneously obvious increase in the level of PARP-1 methylation in nano-SiO2-treated cells compared to the control group. Further, the expression and promoter methylation of PARP-1 in HaCaT cells were restored following DNMT1 knock down, suggesting that the effects of PARP-1 promoter hypermethylation are mediated at least in part by DNMT1. Taken together, methylation of PARP-1 promoter might be involved in the regulation of nano-SiO2-induced decrease of PARP-1 expression.

Role of poly(ADP-ribose) glycohydrolase in the regulation of cell fate in response to benzo(a)pyrene.

Poly(ADP-ribosyl)ation is a crucial regulator of cell fate in response to genotoxic stress. Poly(ADP-ribosyl)ation plays important roles in multiple cellular processes, including DNA repair, chromosomal stability, chromatin function, apoptosis, and transcriptional regulation. Poly(ADP-ribose) (PAR) degradation is carried out mainly by poly(ADP-ribose) glycohydrolase (PARG) enzymes. Benzo(a)pyrene (BaP) is a known human carcinogen. Previous studies in our laboratory demonstrated that exposure to BaP caused a concentration-dependent DNA damage in human bronchial epithelial (16HBE) cells. The role of PARG in the regulation of DNA damage induced by BaP is still unclear. To gain insight into the function of PARG and PAR in response to BaP, we used lentiviral gene silencing to generate 16HBE cell lines with stably suppressed PARG, and determined parameters of cell death and cell cycle following BaP exposure. We found that PARG was partially dependent on PAR synthesis, PARG depletion led to PAR accumulation. BaP-induced cell death was regulated by PARG, the absence of which was beneficial for undamaged cells. Our results further suggested that PARG probably has influence on ATM/p53 pathway and metabolic activation of BaP. Experimental evidences provided from this study suggest significant preventive properties of PAR accumulation in the toxicity caused by BaP.

Effects of long-term low-dose formaldehyde exposure on global genomic hypomethylation in 16HBE cells.

Formaldehyde (FA), a volatile organic compound, is a ubiquitous air pollutant that is classified as 'Carcinogenic to humans (Group 1)' by IARC (2006). As a well-recognized human carcinogen, its carcinogenic mechanisms are still poorly understood. Previous studies have emphasized on genetic changes. However, little is known about the epigenetic mechanisms of FA exposure. In this study, We not only characterized the epigenomic response to long-term low-dose FA exposure in 16HBE cells, but also examined the expression of DNA methyltransferases (DNMTs) and the methyl-CpG-binding protein DNA-binding domain protein 2 (MBD2). Each week the 16HBE cells were treated with 10 µM FA for 24 h (h). After 24 weeks (W) of exposure to FA, the level of genomic DNA methylation gradually decreased in a time-related manner. Moreover, our results showed that FA exposure down-regulated the expression of DNMT3a and DNMT3b at both mRNA and protein level, and up-regulated the levels of DNMT1 and MBD2 at both mRNA and protein level. Our study indicated that long-term FA exposure could disrupt genomic DNA methylation, which may be one of the possible underlying carcinogenic mechanisms of FA.

[Role of poly (ADP-ribose) polymerase 1 on DNA methylation variation induced by B(a)P in human bronchial epithelial cell].

OBJECTIVE: To investigate DNA methylation variation in human cells induces by B(a)P, and to explore the role of PARP1 during this process. METHODS: The changes of DNA methylation of 16HBE and its PARP1-deficient cells exposed to B(a)P (1.0, 2.0, 5.0, 10.0, 15.0, 30.0 µmol/L) were investigated by immunofluorescence and high performance capillary electrophoresis, and simultaneously, the expression level of PARP 1 and DNMT 1 were monitored dynamically. RESULTS: The percentage of methylated DNA of overall genome (mCpG%) in 16HBE and 16HBE-shPARP1 cells were separately (4.04 ± 0.08)% and (9.69 ± 0.50)%. After being treated by 5-DAC for 72 hours, mCpG% decreased to (3.15 ± 0.14)% and (6.07 ± 0.54)%. After both being exposed to B(a)P for 72 hours, the mCpG% in 16HBE group (ascending rank) were separately (5.10 ± 0.13), (4.25 ± 0.10), (3.91 ± 0.10), (4.23 ± 0.27), (3.70 ± 0.15), (3.08 ± 0.07); while the figures in 16HBE-shPARP1 group (ascending rank) were respectively (10.63 ± 0.60), (13.08 ± 0.68), (9.75 ± 0.55), (7.32 ± 0.67), (6.90 ± 0.49) and (6.27 ± 0.21). The difference of the results was statistically significant (F values were 61.67 and 60.91, P < 0.01). For 16HBE group, expression of PARP 1 and DNMT 1 were 141.0%, 158.0%, 167.0%, 239.0%, 149.0%, 82.9% and 108.0%, 117.0%, 125.0%, 162.0%, 275.0%, 233.0% comparing with the control group, whose difference also has statistical significance (t values were 11.45, 17.32, 32.24, 33.44, 20.21 and 9.87, P < 0.01). For 16HBE-shPARP1 group, expression of PARP 1 and DNMT 1 were 169.0%, 217.0%, 259.0%, 323.0%, 321.0%, 256.0% and 86.0%, 135.0%, 151.0%, 180.0%, 229.0%, 186.0% comparing with the control group, with statistical significance (t values were 9.06, 15.92, 22.68, 26.23, 37.19 and 21.15, P < 0.01). When the dose of B(a)P reached 5.0 µmol/L, the mRNA expression of DNMT 1 in 16HBE group (ascending rank) were 125.0%, 162.0%, 275.0%, 233.0% times of it in control group, with statistical significance (t values were 12.74, 24.92, 55.11, 59.07, P < 0.01); while the dose of B(a)P reached 2.0 µmol/L, the mRNA expression of DNMT 1 in 16HBE-shPARP1 group were 135.0%, 151.0%, 180.0%, 229.0%, 186.0% of the results in control group, and the differences were statistically significant (t values were 23.82, 40.17, 32.69, 74.85, 46.76, P < 0.01). CONCLUSION: The hypomethylation of 16HBE cells induced by B(a)P might be one important molecular phenomenon in its malignant transformation process. It suggests that PARP1 could regulate DNA methylation by inhibiting the enzyme activity of DNMT1, and this effect could be alleviated by PARP1-deficiency.

Chromium(VI) causes down regulation of biotinidase in human bronchial epithelial cells by modifications of histone acetylation.

Hexavalent chromium (Cr(VI)), a commonly used industrial metal, is a well-known mutagen and carcinogen, and occupational exposure can induce a broad spectrum of adverse health effects, including cancers. Although Cr(VI)-induced DNA damage is thought to be the primary mechanism of chromate genotoxicity and mutagenicity, there is an increasing number of reports showing that epigenetic mechanisms of gene regulation might be a central target of Cr(VI) toxicity. Epigenetic changes, such as changes in phosphorylation, altered DNA methylation status, histone acetylation and signaling pathways, have been observed after chromium exposure. Nevertheless, to better demonstrate the roles of epigenetic modifications in Cr(VI)-induced carcinogenesis, more work needs to be carried out. This study is aimed to investigate changes in biotinidase (BTD) and holocarboxylase synthetase (HCS), two major proteins which maintain homeostasis of the newfound epigenetic modification: histone biotinylation, in cells exposed to Cr(VI). The data showed that Cr(VI) decreased BTD expression at the transcriptional level in human bronchial epithelial cells (16HBE). In addition, using the epigenetic modifiers, 5-Aza-2'-deoxycytidine (Aza) and Trichostatin A (TSA), we found that modifications of histone acetylation reversed the inhibition of BTD, suggesting that Cr(VI) may cause down regulation of BTD by modifications of histone acetylation.