OBJECTIVE: To investigate interactions between SOX30 and DSC3 genes and their roles in the development of lung adenocarcinoma. METHODS: The GO enrichment analysis was used to screen possible downstream genes and pathways of SOX30 transcriptional regulation. The JASPAR database was used to predict the SOX30 transcription binding sites in the promoter region of DSC3. Expression data of SOX30 and DSC3 were downloaded from the TCGA database,and their relationships were evaluated using the Spearman rank correlation analysis. The reverse transcription-PCR was used to measure the mRNA levels of DSC3 in A549-SOX30+DSC3 miRNA stably transfected cells and the lung tissues of SOX30^+/+,SOX30^+/- and SOX30^-/- mice. The colony formation and wound healing assays were used to confirm whether the inhibition of the proliferation and migration of SOX30 in A549 cells depended on the expression of DSC3 or not. RESULTS: The GO enrichment analysis shows the function of SOX30 gene was significantly related to the cell junction proteins,such as focal adhesion,anchoring junction,and adhesion junction. And several SOX30 binding sites were found in the promoter region of DSC3. Furthermore,expression of DSC3 was in positive correlation with that of SOX30 in human lung adenocarcinoma(r=0.154,P=0.000). In addition,the ectopic expression of SOX30 in A549 cell lines dramatically increased DSC3 mRNA level (P < 0.05),and knockout of SOX30 significantly decreased DSC3 mRNA level in lung tissues of SOX30^-/- mice (P < 0.05). In addition,knockdown of DSC3 attenuated the inhibition effects of SOX30 on proliferation and migration in A549 cells (P < 0.05). CONCLUSION: SOX30 was a critical transcription factor in the expression regulation of DSC3,and DSC3 was an important downstream gene of SOX30. Therefore,SOX30-DSC3 may play an important role in the development of lung adenocarcinoma.

OBJECTIVE: To explore the influence of Chinese herbal compound QHF on human Huh7 hepatoma cell proliferation,apoptosis and invasion and expression of surface markers of cancer stem cells. METHODS: Human Huh7 hepatoma cells were cultured in vitro. Cell proliferation inhibition rate was detected by MTT assay;cell cycle,apoptosis and cell surface marker CD133 and CD44 expression were detected by Flow cytometry;transwell experiment was used to observe cell invasion. RESULTS: QHF inhibited the proliferation of Huh7 cells,and there was a dose-dependent inhibitory effect. In addition,QHF promoted apoptosis and significantly increased cell proportion in S phase of the cell cycle. QHF lowered expression of tumor stem cell surface markers:CD133 and CD44. CONCLUSION: QHF was able to inhibit human Huh7 hepatoma cell proliferation and migration in vitro,the inhibition was related to the reduced expression of tumor stem cell surface markers:CD133 and CD44.

OBJECTIVE: To explore the effect of TAK1 knockout in CD11c⁺ cells on concanavalin A (Con A)-induced immunological liver injury. METHODS: C57BL/6 mice with TAK1 knockout in CD11c⁺ cells were injected intravenously with normal saline or Con A (12 mg/kg) as NS+ and Con A (12 mg/kg)+ knockout groups,respectively. Normal C57BL/6 mice were injected intravenously with normal saline or Con A (12 mg/kg) as NS+ and Con A (12 mg/kg)+ control groups,respectively. After 6 hours,serum was collected to detect the levels of alanine aminotransferase (ALT),aspartate aminotransferase (AST) and lactic dehydrogenase (LDH). Organ coefficients of liver,kidney,thymus and other organs were detected. Histopathological observations and scores were performed. Serum was collected to detect the expression of tumor necrosis factor-α (TNF-α),interferon-γ (IFN-γ) and interleukins (IL-4/5/6 and IL-12). The levels of TNF-α and IFN-γ in liver were detected. RESULTS: Compared with NS+ control group,levels of ALT and LDH were significantly increased in Con A+ control group (P < 0.05). Con A+ control group showed severer pathological liver injury and higher pathological score (P < 0.05). The levels of TNF-α,IFN-γ,IL-4,IL-5,IL-6 in serum and TNF-α and IFN-γ in liver were significantly increased (P < 0.05). Compared with NS+ control group,the level of TNF-α in liver in NS+ knockout group was significantly decreased (P < 0.05). Compared with Con A+ control group,levels of ALT,AST and LDH were significantly increased in Con A+ knockout group (P < 0.05). Con A+ knockout group showed severer pathological liver injury and higher pathological score. The levels of TNF-α,IL-4 and IL-6 in serum and IFN-γ in liver were significantly increased (P < 0.05). CONCLUSION: Knockout of TAK1 in CD11c⁺ cells was not related to liver injury but aggravated concanavalin A-induced immunological liver injury.

OBJECTIVE: To detect expression of mutations in tumor-related genes,such as Kras,Trp53 and Smad4 in pancreatic tissues of diet-induced obese mice and to explore whether obesity is associated with genome instability in the pancreas. METHODS: Male C57BL/6 mice were fed normal chow diet and high-fat diet for 24 weeks. The body weight,food intake,body composition and plasma lipid were measured. Pancreatic genomic DNA was extracted and the exon 2 of Kras,exon 8 of Trp53 and exon 9 of Smad4 were amplified by using high-fidelity PCR. The PCR products were then subjected to the Sanger sequencing protocol. Cell proliferation was assessed by qPCR and immunohistochemistry for Ki67. RESULTS: Intake of 24 weeks of high-fat diet led to severe obesity in mice. High-fat diet feeding increased the body weight,fat mass,lean mass,hepatosomatic index and body-fat ratio of epididymal (P < 0.05). Body weight gain in high-fat diet group was significantly higher than those of chow diet group from eating the same weight of food(P < 0.05). Plasma lipid such as triglyceride,total cholesterol and low density lipoprotein cholesterol were significantly higher in high-fat diet group(P < 0.05). Moreover,the level of Ki67 mRNA was increased in the high-fat diet group(P < 0.05). There were several Ki67 positive cells in pancreatic duct,indicating that proliferation of pancreatic cells in obese mice were elevated. We detected 1 locus mutation in the Kras gene in 1 out of 50 samples (rate:2%). There was no mutation in the other two genes. CONCLUSION: Obesity resulted in instability of Kras gene in pancreatic tissue,which could play a role in the initiation and progression of pancreatic cancer.

OBJECTIVE: To investigate the effects of chlorocholine chloride (CCC) on expression of skeletal development-related genes and functional proteins in mouse pre-osteoblast cell line MC3T3-E1. METHODS: MC3T3-E1 cells in logarithmic growth phase were treated with 0,8,40,200,1 000 μg/mL CCC. At different time points (24,48,72 h) after treatment,cytotoxicity was determined by using the MTT assay. At 48 h after treatment,mRNA expression of skeletal and osteogenic development-related genes was detected using qPCR:alkaline phosphatase (ALP),osteocalcin(OCN),growth factor 1 (IGF-1),growth hormone receptor (GHR),RANKL,osteoprotegerin (OPG),macrophage colony-stimulating factor (M-CSF). In addition,expression of functional proteins related to bone development and MAPK signal pathways was determined by Western blot:ALP,GHR,bone morphogenetic protein 2 (BMP2),transcription factor runt-related protein 2 (Runx2). RESULTS: CCC had no significant effect on cytotoxicity of MC3T3-E1 at different time points (24,48,72 h). mRNA expression of ALP,OCN and RANKL in the 1 000 μg/mL groups increased significantly (P < 0.05);the ratio of RANKL and OPG in the 1 000 μg/mL group increased significantly (P < 0.05);mRNA expression of GHR,IGF-1,OPG,M-CSF did not change significantly (P > 0.05). Protein expression of GHR and BMP2 in the 1 000 μg/mL group was significantly decreased,while ALP and RUNX2 did not change significantly. Phosphorylated ERK protein expression decreased and phosphorylated JNK protein expression increased. CONCLUSION: CCC has almost no cytotoxicity on pre-cultured mouse MC3T3-E1 cells,but inhibited the differentiation of MC3T3-E1 cells into mature osteoblasts and promoted osteoclast differentiation. Moreover,involvement of mitochondria on differentiation of pre-mouse osteoblasts might be related to the MAPK pathway. The decrease of phosphorylated ERK and the increase of phosphorylated JNK might have inhibited migration of MC3T3-E1 to mature osteoblasts differentiation of cells,hence their differentiation to become mature osteoblasts.

OBJECTIVE: To study the anti-liver cancer effect of phenylethanol glycosides from cistanche (CPhGs) in mice. METHODS: H22 tumor-bearing mice were established by using axillary subcutaneous injections. Six groups of 10 mice were used in our study:the experiment established blank control group,H22 tumor-bearing model group,Ganfule positive treatment group (1 351.5 mg/kg),and CPhGs high,medium and low dose groups (500,250,125 mg/kg). In addition to the blank control and the model groups,the rest of them were given Ganfule or CPhGs by gavage for 10 consecutive days. During the period,conditions of these mice were monitored,and the tumor growth of H22 tumor-bearing mice was observed. At the end of the experiment,blood was collected from the eyeball,and serum interleukin-2 (IL-2),tumor necrosis factor-α (TNF-α) and α-fetoprotein (AFP) content were detected by enzyme linked immunosorbent assay (ELISA). Biological samples were collected to calculate the spleen,liver coefficient and inhibition rate. HE staining was conducted to observe pathological histologic changes. RESULTS: The axillary tumor in the model group grew early and grew the fastest,followed by the CPhGs low and medium dose groups. The tumor growth of the high dose group and the positive treatment group were slower. Compared with the model group,the quality of tumor in the CPhGs medium and high dose groups was significantly reduced,and the inhibition rate of each dose group was increased with increasing doses of CPhGs (P < 0.05). The spleen coefficient of the positive treatment group and each CPhGs dose group were significantly increased,and liver coefficient were obviously decreased (P < 0.05). Among the high dose and positive treatment groups,IL-2 contents were significantly elevated (P < 0.05). Each CPhGs dose group and positive treatment group,TNF-α contents were significantly reduced (P < 0.05). Among CPhGs medium,high dose and positive treatment groups,AFP content were significantly reduced (P < 0.05). Pathological results show that the tumor cells in each CPhGs dose group were suppressed with reduced numbers,the heterogeneity was reduced,and there were large numbers of necrotic cells. CONCLUSION: CPhGs exposure reduced liver injury of H22 tumor-bearing mice,adversely affected tumor growth which was probably related to reduce AFP levels in serum of tumor-bearing mice,and improved immune function of tumor-bearing mice.

OBJECTIVE: To study mechanisms in sensitivity of paclitaxel-resistant human lung adenocarcinoma cells from treatment with resveratrol. METHODS: A549/Taxol-R,a human lung adenocarcinoma cell line that resist paclitaxel,was established and treated with different concentrations of resveratrol and paclitaxel alone or in combination. MTT assay was performed and SPSS was used to calculate IC₅₀ of A549/Taxol-R to paclitaxel. Change of IC₅₀ was determined after treatment with resveratrol at different concentrations. Apoptosis rates were detected with flow cytometry. Expressions of Bax,Bcl-2 and Caspase-3 were detected by Western blot. RESULTS: The inhibitory rate of resveratrol to A549/Taxol-R cells was dependent on concentration and time. After treated with resveratrol,the IC₅₀ of paclitaxel on A549/Taxol-R cells decreased from (17.50±1.24) μg/mL to (4.18±0.62) μg/mL. The apoptosis and necrosis rates of A549/Taxol-R cells that were treated with combined resveratrol and paclitaxel were higher than those that treated with single drugs (P < 0.05). Expression of Bax and Caspase3 in the combined treatment group was higher than that in the single treatment group,while expression of Bcl-2 was relatively down-regulated (P < 0.05). CONCLUSION: Resveratrol may increase sensitivity of paclitaxel-resistant human lung adenocarcinoma cells to paclitaxel by promoting cell apoptosis. The mechanism may be related to promotion of expression of Bax and Caspase3 and down-regulation of Bcl-2 expression.

OBJECTIVE: To investigate the protective effect of Rhodiola extract (RE) on cisplatin(DDP)-induced oxidative damage in human embryonic kidney cells (HEK293). METHODS: HEK293 cells were cultured in vitro. The cells were divided into control group,DDP group (0,0.5,1,2,4,8,16,32,64 and 128 mg/L),RE group (0,0.25,0.5,1,2,4,8,16,32 and 64 mg/L),and RE (2-16 mg/L)+DDP (20 mg/L) group. The effect of RE and DDP on the growth of HEK293 cells and the protective effect of RE on DDP-induced cytotoxicity were determined using the CCK-8 assay. Glutathione (GSH) content was measured by dithiobisnitrobenzoate method. Malondialdehyde (MDA) content was measured by thiobarbituric acid method. Superoxide dismutase (SOD) activity was examined by xanthine oxidase method. RESULTS: Compared with the control group,cell survival rates decreased significantly (P < 0.01),and in a dose-dependent manner (r=0.85,P=0.002). The survival rate of RE increased gradually in the range of 0.5-16 mg/L (P < 0.01). When RE concentration was 64 mg/L,the growth of HEK293 cells was inhibited significantly (P < 0.01). The inhibitory effect of 6-16 mg/L RE preconditioning on cell survival with 20 mg/L DDP was significantly higher than that of DDP group (P < 0.01). Compared with DDP group,RE could significantly inhibit the increase of MDA content induced by DDP and the decrease of SOD activity and GSH content (P < 0.01). CONCLUSION: RE treatment enhances the viability of HEK293 cells. In addition,DDP-induced damages in HEK293 cells were remarkedly reversed by RE,and provided protection against DDP-induced nephrotoxicity.

OBJECTIVE: To investigate associations between genotypic polymorphisms of the mitotic checkpoint gene,MAD1L1,and risk of breast cancer among women in the Chaoshan area. METHODS: This was a case-control study consisted of 191 new cases of breast cancer selected from the Tumor Hospital of SUMC between December,2015 and December,2017 and 225 healthy resident controls at the same period. TaqMan allelic discrimination was used to genotype the MAD1L1 locus rs1801368,and related information such as demographic characteristics,reproductive factors,smoking and drinking status,medical history and clinicopathological indexes for the cases were collected. RESULTS: The CC,CT and TT genotypes of rs1801368 accounted for 20.63%,42.86%,36.51% in the cases,and accounted for 30.14%,41.15%,28.71% in controls,genotype distribution showed no statistical significance between cases and controls,while logistic regression analysis showed that after adjusting the confounding factors such as age,age of menarche,menopause and so on,women carrying mutant homozygous TT genotype were more susceptible to breast cancer than those with wild homozygous CC genotype (OR=3.399,95% CI:1.288-8.973,P=0.013). CONCLUSION: Genetic variation of the SNP locus rs1801368 in mitotic checkpoint gene MAD1L1 was associated with breast cancer risk. Therefore,women who had the variant genotype would be susceptible to breast cancer.

OBJECTIVE: The aim of this study was to investigate reproductive toxicity of tebuconazoleine tebuconazoleb (TEB) in Caenorhabditis elegans(C. elegans). METHODS: Wild-type(N2) and him-5(e1490) strains from L2-larvae to L4-larvae stages were exposed to TEB at concentrations of 0.1,1.0 and 10.0 μg/L. Brood size and generation time measurements were used to evaluate fertility defects in nematodes. Furthermore,the number of outcross progeny was used to detect reproductive toxicity on male worms. Multiple endpoints were employed to evaluate defects on spermatogenesis,including the number of germ cells in mitosis and meiosis zoon,the number of germs in the germ line and the expression levels of related genes. Sperm size,morphology and activation ability were used to evaluate the adverse effect on spermiogenesis. The level of related genes was detected by real-time fluorescence quantitative PCR. RESULTS: The brood sizes at a concentration of 10.0 μg/L and the number of outcross progeny at concentrations more than 1.0 μg/L were reduced. However,generation times were not changed compared with that from the control group. The number of total germ cells,mitotic cells,meiotic cells and spermatids were less than that from the control group at concentrations of more than 1.0 μg/L. mRNA levels of daf-2,akt-1,and daf-16 were significantly increased at concentrations of 1 and 10 μg/L. Additionally,the mRNA level of age-1 was decreased at the concentrations of 0.1 to 10.0 μg/L compared to the control group. The sperm diameter and cross-sectional sperm area were reduced at concentrations of 0.1-10.0 μg/L,the activation ability of the sperm was declined in the 1.0 μg/L and 10.0 μg/L treatment groups. The expression level of try-5,spe-4,spe-6,fer-1 mRNA were increased significantly and swm-1 mRNA was decreased compared with the control group. CONCLUSION: TEB exposure causeds reproductive toxicity in Caenorhabditis elegans by affecting spermatogenesis and spermiogenesis.

OBJECTIVE: To explore the biological characteristics of sperm-tumor cell fusion model. METHODS: A co-cultivation in vitro methods was used to study the time-effectiveness,cell inoculation density dependence and cell type specificity of the fusion model. The chimeric sperm-HeLa monoclonal cells were obtained by using micromanipulation and monoclonal culture method,which was then identified by using PCR test. The chimeric tumor cells in the cervical cancer paraffin were detected by amplification Y chromosome specific gene TSPY. RESULTS: Fusion between the sperm and tumor cells generally occurred in 1.5 hours after co-cultivation,the fusion rates was inversely proportional to the cell inoculation density,and the tumor cells who derived from an adenocarcinoma had a higher rate of cell fusion than other tumor cells and normal controls. One chimeric tumor tissue was observed in 20 cases of cervical cancer specimens,and one righteous mutation was found in TSPY gene. CONCLUSION: Cell fusion between the sperm and tumor cells displayed time-effectiveness,cell inoculation density dependence and cell type specificity. The chimeric tumor cells could be found in cervical cancer specimens.

OBJECTIVE: To investigate the effects of long-term and repeatedly exposure to radon from hot spring on oxidative damage and antioxidation in the population. METHODS: A simple random sampling method was used to select 38 residents around radon hot spring in Jiangzha township as high radon group,and 39 residents of Axirong township were selected as control group. The levels of 8-hydroxydeoxyguanosine (8-OHdG) and thioredoxin reductase (TrxR) in plasma of the two groups were measured by enzyme-linked immune sorbent assay (ELISA). The Mann-Whitney U test was used to compare the differences in 8-OHdG and TrxR between the two groups. Questionnaire surveys were conducted on residents' cognition of radon. RESULTS: Levels of 8-OHdG in peripheral plasma of residents with high radon group were significantly decreased and levels of TrxR were significantly increased (Z=-3.316,-3.773,P < 0.05),which were 0.571 and 2.325 times that of the control group respectively. Radon exposure was associated with differential expression of 8-OHdG and TrxR (P < 0.05). In addition,the two groups of residents lacked knowledge about radon and its impact on human health and about measures to reduce it. CONCLUSION: Long-term and repeatedly exposure to radon from hot springs enhanced body's antioxidant capacity to a certain extent,and reduced the level of oxidative damage. Local residents had a serious lack of awareness of radon,and they should be educated about such knowledge.

OBJECTIVE: To investigate the effects of different doses of di (2-ethylhexyl) phthalate (DEHP) on blood cell parameters in rats after 1 mg/kg selenium's intervention. METHODS: Seventy-seven healthy male SD rats were randomly divided into 7 groups:the control group,three DEHP groups with different concentrations (300,600,900 mg/kg),and Se +different doses of DEHP (1 mg/kg Se + 300,600,and 900 mg/kg DEHP). The rats were continuously fed for 90 days. During the experiment,the rats were observed for signs of poisoning (spiritual excitement,loose hair,partial hair loss). Body weight and whole blood were used to measure changes in various blood cell parameters at the end of the treatment time. RESULTS: Compared with the control group,the body weights of the rats exposed to 900 mg/kg DEHP were significantly decreased (P < 0.05). The proportion of toxic signs in the 600 and 900 mg/kg DEHP groups were 63.63% (7/11) and 100% (11/11),which were significantly higher than 0 (0/11) in the control group (P < 0.05). Among DEHP treated rats,the body weights were significantly increased after 1 mg/kg Se intervention (P < 0.05). The signs of poisoning in the 1 mg/kg Se+300 and 600 mg/kg DEHP groups were significantly lower. Compared with the control group,the WBC of the rats exposed to 600 and 900 mg/kg DEHP was significantly lower and the MONO was significantly higher (P < 0.05). Compared with the same dose of DEHP-treated group,the WBC of the 1 mg/kg Se+600 mg/kg DEHP group was significantly increased and the MONO was significantly decreased (P < 0.05). Among the erythrocyte parameters and compared with the control group,the MCH and HDW of the rats were significantly increased after exposure to DEHP;the RBC and HCT of the 600 and 900 mg/kg DEHP treated groups were significantly lower (P < 0.05). Compared with the same dose of DEHP-treated group,there was no significant difference in the red blood cell parameters of the rats after intervention by 1 mg/kg Se. Among the platelet parameters,no DEHP exposed group and Se intervention group had significant effects. CONCLUSION: Exposure of rats to 300-900 mg/kg DEHP affected certain red and white blood cell parameters. Intervention by selenium protected certain blood cell functions under the doses of 300-600 mg/kg DEHP.

OBJECTIVE: To construct a plasmid which interfered with expression of the human SRF gene,and to study the role of SRF in oral squamous cell carcinoma. METHODS: A SRF gene-specific knockdown fragment was designed by using the Thermo Fisher's RNAi design tool,and then by double digestion of the pLKO.1 vector,gel extraction,and connection of specific fragment and the linearized vector by T4 ligase,then the pLKO.1-hSRF plasmid was obtained. The recombinant plasmids were identified by sequencing and restriction enzyme digestion. 293T cells were used to generate lentivirus by co-transfecting pLKO.1-hSRF with helper plasmids pVSV-G and pHR. The virus were collected and used to infect the oral squamous carcinoma cells,SAS. Stable cells were selected by puromycin and verified by Western blot and real-time quantitative PCR. RESULTS: The lentiviral knockdown plasmid pLKO.1-hSRF was identified by sequencing and restriction enzyme digestion. The expression of SRF protein and mRNA level of SAS cells infected with the lentiviral plasmid were significantly decreased compared with the control group. CONCLUSION: The lentiviral knockdown plasmid pLKO.1-hSRF was successfully constructed and the SAS cell line of low expression of SRF was obtained.