With the rapid development of industry and economy,the emergence of a large number of chemicals has made of risk management more difficult.Traditional risk assessment relies on animal experiments for toxicity testing.However,animal experiments are time-consuming,costly,and unable to meet the practical needs of risk assessment.The increasing maturity of toxicity testing alternative technologies signifies the possibility of rapid,sensitive,and accurate identification of chemical toxicity.This article focuses on the research and applications of alternative toxicity testing by reviewing the background,developments,and current research at home and abroad.It also discusses the progress in alternative testing methods in such areas as cosmetics and food safety risk assessment and explores the problems with the development of alternative testing technologies and risk assessment in China.This review aims to provide a reference for the system construction of cosmetics health risk assess-ment in China.

OBJECTIVE To explore the applicablity of 'BlueScreen HC'(BSHC),a high throughput genotoxicity screening system based on human growth arrest and DNA damage inducible 45α(GADD45α)gene,in detecting the genotoxicity of migrants mixtures from food contact materials(FCM).METHODS The 2000 bp sequence upstream of the open reading frame of human GADD45α gene was used as the promoter to construct the lentiviral plasmid pEZX-LvPG04,which was double labeled by purinamycin and Gausluciferase(Gluc),and the lentiviral plasmid was infected with human lymphoblastocyte TK6 to obtain a stable transmutation cell line TK6-Gluc.Methyl methylate(MMS)at concentrations of 0,1.56,3.13,6.25,12.5,25.0 and 50.0 mg·L-1 was selected as the genotoxin without liver S9,cyclophosphamide(CTX)0,0.78,1.56,3.13,6.25,12.5,25.0 mg·L-1 was selected as the pre-genotoxin with liver S9,and dimethyl sulfoxide(DMSO)0,0.35,0.69,1.38,2.75,5.5 and 11.0 g·L-1 was selected as the non-genotoxin.The constructed BSHC was verified with the above known genetic positive and negative substance respectively.Polybutyleneadipate-co-terephthalate(MS/PBAT)was tested using 4% (V/V)acetic acid,and 10%,20%,50% and 95% (V/V)ethanol as food simulants at 40℃for 24 hours to obtain 5 multi-component migrants of MS/PBAT that were obtained by using DMSO as a solvent.TK6-Gluc cells were treated with 5 multi-component migrants of MS/PBAT at concentrations of 0,0.38,0.76,1.53,3.05,6.10 and 12.20 g·L-1 with or without liver S9.Cells were treated without liver S9 for 48 h.Cells treated with liver S9-mix were incubated for 3 h at a final concen-tration of 1% (V/V)liver S9 before being washed and re-suspended in fresh recovery media for another 45 h.After exposure,the cell viability was detected using the CCK-8 cell activity kit,and the Gluc Lumi-nescence in the medium was detected with Secrete-PairTM Gaussia Luciferase Assay Kit.In addition,the mutagenicity on Salmonella typhimurium TA98 and TA100 was detected by micro-fluctuation Ames test with 5 multi-component migrants of MS/PBAT at concentrations of 3.05 and 12.20 g·L-1.The in vitro mammalian cell chromosome aberration test was performed on CHL cells with 5 multi-component migrants of MS/PBAT at concentrations of 3.05 and 12.20 g·L-1 to detect the chromosomal aberration.The results of genotoxicity were compared with those of BSHC.RESULTS The lowest effect centra-tion(LEC;<80% relative cell viability)and the coytotoxicity(<30% relative cell viability)was defined.A positive genotoxicity result threshold was determined at 1.8-fold relative induction.For the liver S9 protocol,the same process was followed,and the decision threshold derived was 1.5-fold relative Gluc induction.It is considered as genetic substance only when a positive genotoxicity result was reached and there was no cytotoxicity.Compared with the vehicle control group,no genotoxicity was observed at all concentration of DMSO by BSHC.MMS 12.5,25.0 and 50.0 mg·L-1 produced genotoxicity without liver S9 while CTX 6.25,12.5 and 25.0 mg·L-1 produced genotoxicity with liver S9.Significant cell growth inhibition was observed in 95% ethanol migrants of MS/PBAT at concentrations of 6.10 and 12.20 g·L-1,and in 50% ethanol migrants of MS/PBAT at a concentration of 12.20 g·L-1 without liver S9.No cytotoxicity with a relative cell viability below 30% was observed in any of the treatment groups,and no high expression of Gluc was observed.Therefore,none of the 5 multi-component migrants produced genotoxicity without liver S9.Significant cell growth inhibition was observed in 95% ethanol migrants of MS/PBAT at a concentration of 12.20 g·L-1,and in 4% acetic acid migrants of MS/PBAT at concentrations of 6.10 and 12.20 g·L-1 with liver S9.No cytotoxicity with a relative cell viability below 30% was observed in any of the treatment groups.No high expression of Gluc was observed.There-fore,none of the 5 multi-component migrants produced genotoxicity with liver S9.In the micro fluctua-tion Ames test,when 5 multi-component migrants of MS/PBAT were treated with concentrations of 3.05 and 12.20 g·L-1 on TA98 and TA100 strains,there was no significant difference in the number of muta-genic positive wells compared with DMSO control group with or without liver S9,indicating that no mutagenic effect was produced.When CHL cells were treated with 5 multi-component migrants of MS/PBAT at concentration of 3.05 and 12.20 g·L-1,compared with DMSO control group,there was no signifi-cant difference in chromosome aberration rate of CHL cells with or without liver S9.CONCLUSION BSHC based on GADD45α gene has been established,which can be used for in vitro genotoxicity eval-uation of migrants mixtures of FCM,but further exploration of its minimum effective concentrations is still needed,and more types of mixtures need to be applied for further validation.

Objective:To investigate the role of autophagy mediated by mTOR signaling pathway in the inhibition of osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) induced by cadmium.Methods:HBMSCs were divided into 0, 2.5 or 5.0 μmol/L groups according to the exposure dose of cadmium chloride (CdCl 2), and each group was treated for 1 day, 4 days and (or) 7 days. The ALP activity and mRNA and protein expression levels of osteogenesis markers (ALP, RUNX2 and OSTERIX), autophagy-related proteins (LC3 and Beclin-1) and mTOR signaling pathway related proteins (mTOR, p-mTOR and p-p70S6K) expression, alkaline phosphatase staining and alizarin red staining were detected. MHY 1485 was selected as the signaling pathway activator. The control group, CdCl 2 group (5.0 μmol/L), MHY 1485 group and CdCl 2+MHY 1485 combined treatment group were set. After 7 days of treatment, the expression levels of autophagy related proteins and mTOR signaling pathway related proteins of hBMSCs in each group were detected. Results:There was no significant difference in ALP activity between 0, 2.5 and 5.0 μmol/L groups on day 1 and 4 ( P>0.05); On day 7, compared with the 0 μmol/L group, the ALP activity, expression of osteogenic markers (ALP, RUNX2, OSTERIX) and mTOR signaling pathway related proteins (mTOR, p-mTOR, p-p70S6K) expression decreased in the 2.5 and 5.0 μmol/L group ( P<0.05). Compared with the 0 μmol/L group, the staining of the 2.5 and 5.0 μmol/L groups became lighter, and the formation of ALP and mineralized nodules was reduced. Compared with the CdCl 2 group, the autophagy related protein expression in the CdCl 2+MHY 1485 combined treatment group decreased, and the mTOR signaling pathway related protein expression increased. The difference was statistically significant ( P<0.05). Conclusion:The inhibition of osteogenic differentiation of hBMSCs by cadmium may be related to autophagy mediated by mTOR signaling pathway.

Objective:To investigate the role of autophagy mediated by mTOR signaling pathway in the inhibition of osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) induced by cadmium.Methods:HBMSCs were divided into 0, 2.5 or 5.0 μmol/L groups according to the exposure dose of cadmium chloride (CdCl 2), and each group was treated for 1 day, 4 days and (or) 7 days. The ALP activity and mRNA and protein expression levels of osteogenesis markers (ALP, RUNX2 and OSTERIX), autophagy-related proteins (LC3 and Beclin-1) and mTOR signaling pathway related proteins (mTOR, p-mTOR and p-p70S6K) expression, alkaline phosphatase staining and alizarin red staining were detected. MHY 1485 was selected as the signaling pathway activator. The control group, CdCl 2 group (5.0 μmol/L), MHY 1485 group and CdCl 2+MHY 1485 combined treatment group were set. After 7 days of treatment, the expression levels of autophagy related proteins and mTOR signaling pathway related proteins of hBMSCs in each group were detected. Results:There was no significant difference in ALP activity between 0, 2.5 and 5.0 μmol/L groups on day 1 and 4 ( P>0.05); On day 7, compared with the 0 μmol/L group, the ALP activity, expression of osteogenic markers (ALP, RUNX2, OSTERIX) and mTOR signaling pathway related proteins (mTOR, p-mTOR, p-p70S6K) expression decreased in the 2.5 and 5.0 μmol/L group ( P<0.05). Compared with the 0 μmol/L group, the staining of the 2.5 and 5.0 μmol/L groups became lighter, and the formation of ALP and mineralized nodules was reduced. Compared with the CdCl 2 group, the autophagy related protein expression in the CdCl 2+MHY 1485 combined treatment group decreased, and the mTOR signaling pathway related protein expression increased. The difference was statistically significant ( P<0.05). Conclusion:The inhibition of osteogenic differentiation of hBMSCs by cadmium may be related to autophagy mediated by mTOR signaling pathway.

OBJECTIVE: To survey the residents for their understanding of Coronavirus Disease 2019 (COVID-19)-related knowledge, attitude and practices (KAP) in two hard hit provinces of China to facilitate the governmental decisions on strategies against the disease. METHODS: We invited the participants from Hubei and Henan Provinces of China for an internetbased survey starting from 12:00 on February 21, 2020 to 12:00 on February 23. The survey included the general conditions, KAP of COVID-19, psychological status and living conditions of the residents. RESULTS: The effective response rate of the questionnaire was 98.9%. The mean (P25, P75) age of the participants was 19 (16, 40) years, and 54.3% of them were students. Social media were the most important source of information concerning the pandemic of the respondents. The respondents had a high awareness of person-to-person transmission of the virus through the respiratory tract or droplets but showed a relatively low level of awareness of the population susceptible to COVID-19 and its specific symptoms. The results of multivariate analysis showed that women, undergraduate students (including college students) and higher degree holders had better knowledge of COVID-19 ( < 0.05); the proportion of respondents who expressed to have different levels of psychological stressed such as worry, anxiety and panic reached 77.2%; 16.7% of the responders considered psychological interventions necessary for their psychological conditions; 63.6% of the respondents confessed a bias against the people returning from Hubei and Henan provinces, while 22.4% worried that they might be biased because of their residence in Hubei and Henan. The rate of personal protective equipment shortage was as high as 69.4%; the rates of the responders who would "covering the mouth and nose when coughing or sneezing", "properly use masks in accordance with regulations", "maintain proper hand hygiene ", "avoid gatherings with relatives and friends" and "refrain from going to public places" were 92.4%, 95.9%, 93.5%, 88.8% and 93.1%, respectively. Women and groups with good knowledge of the disease reported better protective behaviors against the diseases ( < 0.05). CONCLUSIONS The residents in Hubei and Henan Provinces have generally good KAP related to COVID-19, and the online platforms plays a positive role to in circulating epidemic-related information. It is essential to further increase the supply of the protective materials and pay more attention to the mental health of the residents during the pandemic, and psychological counseling and psychological protection should be provided if necessary.
Attitude ; Betacoronavirus ; Coronavirus Infections ; Female ; Humans ; Pandemics ; Pneumonia, Viral

Objective: To study the alterations of mitochondrial biological characteristics during both cellular replicative and premature senescence induced by hydrogen peroxide in human embryonic lung fibroblasts (HEFs). Methods: The premature senescence was induced by 400 μmol/L H₂O₂ once a day at the same time and with 2 hours each time, after four consecutive days the premature senescence models were classified into premature senescence initiation group (PSi) and premature senescence persistence group (PSp). Based on the life span of HEFs, the cell replicative senescence was divided into five groups included young-age (22 PDL), middle-age (35 PDL), replicative senescence (49 PDL), PSi and PSp. The mitochondrial distribution, relative content, adenosine triphosphate (ATP) contents, 8-hydroxydeoxyguanosine (8-OHdG) levels, the relative mitochondrial transcription factor A (TFAM) as well as mitochondrial DNA methyltransferase 1 (mtDNMT1) mRNA levels, mtDNA copy number, the relative TFAM protein level and the total enzyme activity of mitochondrial DNA methyltransferases (mtDNMTs) were detected in five senescence groups. Results: The mtDNA copy number, 8-OHdG contents, level of mtDNMT1 mRNA and mtDNMTs activity in 49 PDL group were higher than those in 22 PDL group (all P values <0.05); The level of 8-OHdG in PSi was higher than that in 22 PDL group (P<0.05); The ATP contents, mtDNA copy number, the mRNA and protein expression levels of TFAM and mtDNMTs activity of PSp were higher than those in 22 PDL group (all P values<0.05). Conclusion During the cellular senescence of HEFs, the higher mtDNA copy number and mtDNMTs activity were common features regardless of replicative or premature senescence, with possibility that oxidative stress was involved in modifying the occurrence of premature senescence.

Objective To study the alterations of mitochondrial biological characteristics during both cellular replicative and premature senescence induced by hydrogen peroxide in human embryonic lung fibroblasts (HEFs). Methods The premature senescence was induced by 400 μmol/L H2O2 once a day at the same time and with 2 hours each time, after four consecutive days the premature senescence models were classified into premature senescence initiation group (PSi) and premature senescence persistence group (PSp). Based on the life span of HEFs, the cell replicative senescence was divided into five groups included young-age (22 PDL), middle?age (35 PDL), replicative senescence (49 PDL), PSi and PSp. The mitochondrial distribution, relative content, adenosine triphosphate (ATP) contents, 8?hydroxydeoxyguanosine (8?OHdG) levels, the relative mitochondrial transcription factor A (TFAM) as well as mitochondrial DNA methyltransferase 1 (mtDNMT1) mRNA levels, mtDNA copy number, the relative TFAM protein level and the total enzyme activity of mitochondrial DNA methyltransferases (mtDNMTs) were detected in five senescence groups. Results The mtDNA copy number, 8?OHdG contents, level of mtDNMT1 mRNA and mtDNMTs activity in 49 PDL group were higher than those in 22 PDL group (all P values<0.05); The level of 8?OHdG in PSi was higher than that in 22 PDL group (P<0.05); The ATP contents, mtDNA copy number, the mRNA and protein expression levels of TFAM and mtDNMTs activity of PSp were higher than those in 22 PDL group (all P values<0.05). Conclusion During the cellular senescence of HEFs, the higher mtDNA copy number and mtDNMTs activity were common features regardless of replicative or premature senescence, with possibility that oxidative stress was involved in modifying the occurrence of premature senescence.

Objective To study the alterations of mitochondrial biological characteristics during both cellular replicative and premature senescence induced by hydrogen peroxide in human embryonic lung fibroblasts (HEFs). Methods The premature senescence was induced by 400 μmol/L H2O2 once a day at the same time and with 2 hours each time, after four consecutive days the premature senescence models were classified into premature senescence initiation group (PSi) and premature senescence persistence group (PSp). Based on the life span of HEFs, the cell replicative senescence was divided into five groups included young-age (22 PDL), middle?age (35 PDL), replicative senescence (49 PDL), PSi and PSp. The mitochondrial distribution, relative content, adenosine triphosphate (ATP) contents, 8?hydroxydeoxyguanosine (8?OHdG) levels, the relative mitochondrial transcription factor A (TFAM) as well as mitochondrial DNA methyltransferase 1 (mtDNMT1) mRNA levels, mtDNA copy number, the relative TFAM protein level and the total enzyme activity of mitochondrial DNA methyltransferases (mtDNMTs) were detected in five senescence groups. Results The mtDNA copy number, 8?OHdG contents, level of mtDNMT1 mRNA and mtDNMTs activity in 49 PDL group were higher than those in 22 PDL group (all P values<0.05); The level of 8?OHdG in PSi was higher than that in 22 PDL group (P<0.05); The ATP contents, mtDNA copy number, the mRNA and protein expression levels of TFAM and mtDNMTs activity of PSp were higher than those in 22 PDL group (all P values<0.05). Conclusion During the cellular senescence of HEFs, the higher mtDNA copy number and mtDNMTs activity were common features regardless of replicative or premature senescence, with possibility that oxidative stress was involved in modifying the occurrence of premature senescence.

Objective: A new ion exchange column technology was used to establish an efficient and sensitive method for the detection of inorganic arsenic. Methods: Based on the new As Specia Fast Column, the pretreatment methods, liquid phase separation and mass spectrometry determination conditions of inorganic arsenic in rice were optimized. Finally, arsenic compounds were separated by As Specia Fast Column and detected by liquid chromatography inductively coupled plasma mass spectrometry. The external standard method was used for quantitative analysis. The detection limit, precision and accuracy of the method were determined by measuring the content of arsenic compounds in rice samples and rice standard samples. At the same time, three Guangdong rice samples were selected as the experimental samples of this study, and 1 g of each sample was weighed and measured in parallel three times. The method was compared with the method of liquid chromatography-atomic fluorescence spectrometry (LC-AFS) and liquid chromatography-inductively coupled plasma mass spectrometry (LC-ICP-MS) in the national standard. Results: The inorganic arsenic in rice was extracted with 0.5% nitric acid solution at 65 ℃ for 15 h, and the pH was adjusted to alkaline. The mobile phase A (8 mmol/L HNO₃, 50 mmol/L NH₃·H₂O) and mobile phase B (40 mmol/L HNO₃, 80 mmol/L NH₃·H₂O) were used as the mobile phase gradient elution (93%) . Five arsenic compounds can reach baseline separation under the conditions of RF power of 1 500 W and atomization gas flow of 0.97 L/min. The detection limits ranged from 0.114 to 0.331 μg/L, and the inorganic arsenic content in rice samples ranged from 0.063 to 0.232 mg/kg. The results of determination of arsenic compounds in rice flour reference materials were all within the uncertainty range indicated by the standard. The recoveries were 86.7%~106.7%, and the precision was 1.9%-12.5%. Compared with national standards, the results of determination of arsenate in rice were relatively close (using this method, LC-AFS, LC-ICP-MS to detect the content of arsenate in rice samples 1 was 0.231, 0.226, 0.236 mg/kg, respectively). However, due to insufficient sensitivity, the national standard method is difficult to detect low levels of arsenic compounds (Arsenobetaine was not detected in rice sample 1). The method can detect the content of arsenobetaine in rice sample 1 was 0.023 mg/kg. Conclusion The established method can meet the requirements of inorganic arsenic determination in rice, and it is more rapid and accurate than the current national standard. It can better monitor and evaluate the content of i-As in rice, and provide accurate data for comprehensively grasping and evaluating the safety of rice consumption of residents.

Objective To establish an HPLC method for simultaneous determination of Isoquercitrin and astragalin in Moringa oleifera leaves prescription. Methods By using HPLC method with Cosmosil-C18 column (4.6 mm × 250 mm, 5 μm) and the column temperature was 40 degrees Celsius, the mobile phase contains a 0.1％phosphoric acid in water(A) and acetonitrile(B), the detection wavelength was set at 350nm by UV detector and the flow rate was 1.3 ml/min. Results The Isoquercitrin and astragalin was better separated in 30 minutes, and Isoquercitrin had a good linear correlation at the range of 0.24-4.73 μg, while astragalin was 0.11-2.19 μg, the average spiked recoveries of isoquercetin and astragalin were 99.09％ (RSD=0.60％) and 99.08％ (RSD=1.37％), respectively. Conclusions The proposed method is simple,accurate,and repeatable and can be used for quality control as well as a powerful tool to evaluate the processing of Moringa oleifera leaves. It can provide the reference basis and data support for the determination of content of Moringa oleifera health products.