OBJECTIVE： This study aimed to investigate effects of nuezhenide (NED) on LPS-induced proinflammatory response in macrophages. METHODS： RAW264.7 cells were treated with 50 μmol/L NED for 12 hours. Differential gene expressions were detected via transcriptome，cell viability using the CCK8 kit，and apoptosis rates measured using Annexin V-FITC/PI staining，and expression and localization of Nrf2 protein were detected by immunofluorescence. The cells were treated with LPS (100 ng/mL) and NED (12.5，25 and 50 μmol/L) for 12 hours. Then，TNF-α and IL-6 contents in the supernatant of cultured cells were determined by ELISA，and expressions of TNF-α and IL-6 by RT-PCR. Moreover，ROS，mitochondrial mass，and membrane potential were measured by flow cytometry after staining with DCFH-DA，MitoTracker-Green and JC-1，respectively. Enzyme activities of SOD and CAT，activities of mitochondrial complexes I and Ⅲ，as well as ATP contents were determined using commercial kits. RESULTS： Our results show that 50 μmol/L NED had no significant effects on survival and apoptosis of the RAW264.7 macrophages (P>0.05). However，NED treatments blunted LPS-induced expression of TNF-α and IL-6 (P<0.05)，and enhanced expression of anti-inflammatory cytokines (P<0.05). Meanwhile，NED significantly promoteed Nrf2 nuclear translocation and downstream antioxidant transcription (P<0.05)，which contributed to inhibition of LPS-induced accumulation of ROS and GSSG (P<0.05). In addition，NED reversed the LPS-induced decreased of mitochondrial membrane potential，mitochondrial complexes I and III activities，as well as ATP contents through promoting mitochondrial biogenesis (P<0.05). CONCLUSION： NED inhibited LPS-induced macrophages pro-inflammatory differentiation through enhancing Nrf2 antioxidant system and mitochondrial biogenesis. Application of NED may be a potential strategy for treatment of inflammatory diseases.

OBJECTIVE： To explore the effect of m⁶A demethylase FTO on m⁶A methylation，proliferation，metastasis，invasion and other biological behaviors of thyroid cancer cells，and its involvement with the MAPK-JNK signaling pathway. METHODS： The FTO knockdown plasmid was transfected into human thyroid papillary carcinoma cells (TPC-1)，thyroid squamous carcinoma cells (SW-579)，and undifferentiated thyroid carcinoma cells (8505C). Expression of m⁶A in the cancer cells were measured using a m⁶A methylation quantification detection kit. Their proliferation，migration，and invasion abilities were evaluated using the colony formation，CCK-8，wound healing，and Transwell assays. Changes in expression levels of proteins in the MAPK-JNK signaling pathway were examined using Western blot analysis. RESULTS： FTO knockdown increased m⁶A expression levels in the TPC-1，SW-579，and 8505C cancer cells，and promoted their proliferation，invasion，and migration abilities (compared to cells without FTO knockdown，P<0.01). Western blot analysis revealed that FTO knockdown led to decreased expression of E-cadherin and β-catenin，increased expression of N-cadherin and vimentin，and upregulated expression of C-JUN and MLK3 in the MAPK signaling pathway (compared with cells without FTO knockdown，all P<0.05). CONCLUSION： FTO knockdown enhanced proliferation，migration，and invasion abilities of thyroid cancer cells in vitro，and activated the MAPK signaling pathway，thereby potentially influenced development of thyroid cancer. These findings provide insights into the role of FTO demethylase in m⁶A methylation and in thyroid cancer progression.

OBJECTIVE： To investigate expressions of microRNA miR-653-5p in the serum of colon cancer patients and their clinical significance. METHODS： From January 2019 to January 2020，76 colon cancer patients who were treated in the First People’s Hospital of Shangqiu City were selected as the experimental group，and 76 healthy physical examiners in the same period were selected as the observation group. From the collection of 5 mL of fasting venous blood from the subjects，serum miRNA was extracted. Real-time quantitative fluorescence PCR (qPCR) was used to detect expression levels of miR-653-5p，and their differences between the two groups were compared. According to the relative expression level of miR-653-5p，the colon cancer patients in the experimental group were divided into high and low expression groups，and the correlation between miR-653-5p expression and the clinicopathological indicators of colon cancer were analyzed. The clinical diagnostic value of miR-653-5p was analyzed by using the receiver operating characteristic curve (ROC). Kaplan-Meier and multivariate COX regression were used to analyze factors influencing colon cancer survival prognosis. RESULTS： The relative expression level of miR-653-5p in the serum of the control group was 0.622±1.319，and was 0.471±1.076 for the patients，which was significantly lower than that in the control group (P<0.01). There were no significant correlations between the expression levels of miR-653-5p and the sex，age，tumor location and size of colon cancer patients (P>0.05)，but they were related to clinical T stages，distant metastasis，lymph node metastasis，vascular cancer thrombus and tumor differentiation (all P<0.01). The area under the curve of miR-653-5p for clinical diagnosis of colon cancer was 0.823 (P<0.01)，the cut-off value was 0.498，the sensitivity was 80.26%，and the specificity was 82.89%. The Kaplan-Meier curve showed that the survival rate of colon cancer patients in the high-expression group of miR-653-5p was significantly higher than that in the low-expression group (P<0.01). COX regression analysis showed that miR-653-5p expression levels，clinical T stages，distant metastasis，lymph node metastasis，and tumor differentiation were independent risk factors affecting the prognosis and survival of colon cancer patients (all P<0.01). CONCLUSION： The expression levels of miR-653-5p in the serum of colon cancer patients were significantly down-regulated. Our results suggest that miR-653-5p may be a useful and novel biomarker for the diagnosis and prognosis evaluation of colon cancer.

OBJECTIVE： To investigate involvement of the long non-coding RNA (lncRNA) and messenger RNA (mRNA) co-expression network in chronic low-dose cadmium exposure in esophageal squamous carcinoma (ESCC) cells，and to explore molecular mechanisms of hub lncRNAs and mRNAs in the cadmium-associated cancer progression and chemo-and radio-resistance. METHODS： EC109 cells were continually treated with 5 μmol/L cadmium chloride over 12 weeks to construct the chronic cadmium treated (CCT)-ESCC cell model，named as CCT-EC109. The whole-transcriptome sequencing was performed by the Illumina NovaSeq 6000 system，and then bioinformatics methods were applied to identify the differentially expressed (DE)- lncRNA and DE-mRNA expression profiles. Functions of target genes of DE-lncRNA were then predicted based on gene ontology (GO) and the Kyoto encyclopedia of genes and genomes (KEGG) forecast analysis. The lncRNA-mRNA co-expression network was conducted based on the Pearson correlation coefficient and the screened lncRNAs and mRNAs were verified using quantitative real-time polymerase chain reaction (qPCR). RESULTS： A total of 2 794 DE-lncRNAs and 4 267 DE-mRNAs were identified in the EC109 and CCT-EC109 cells. Intersection of the target genes of DE-lncRNA and DE-mRNAs yielded 1 546 cadmium-related mRNAs targeted by corresponding DE-lncRNA. GO analysis showed these genes were mainly enriched in metabolic process，membrane components and membrane closed cavity，and catalytic reaction，etc. For KEGG analysis，Hippo was shown to be the most significant enrichment pathway. Thirty-eight mRNAs were identified to be co-expressed with the top ten DE-lncRNAs. The results of qPCR assay indicated that both the key lncRNA NONHSAT097388.2 and its co-expressed mRNAs including EMP2，ECE1，ORAI1，and ZNF713，were significantly downregulated in the exposed cell line (all P<0.01)，while the expressions of PGF and MFAP5 were found to be upregulated (both of P<0.05). CONCLUSION： LncRNA-mRNA co-expression networks may play important roles in the mechanisms of progression and resistance in ESCC cells chronically exposed to cadmium. Meanwhile，these hub RNAs may serve as prognostic biomarkers or therapeutic targets for the ESCC patients with cadmium exposure.

OBJECTIVE： To investigate expression of forkhead box protein M1 (FoxM1) in epithelial ovarian cancers and to determine its association with clinicopathological characteristics，and with epithelial mesenchymal transition (EMT)-related factors，E-cadherin and vimentin. METHODS： Expressions of FoxM1，E-cadherin and vimentin in 41 cases of epithelial ovarian cancer，17 cases of borderline ovarian tumors and 20 cases of normal ovarian tissues were detected by immunohistochemistry. χ² test was used to compare differences in the expression of FoxM1 between epithelial ovarian cancer tissues，borderline ovarian tumors and normal ovarian tissues. In addition，correlations were investigated among expressions of FoxM1，E-cadherin and vimentin，and clinicopathological characteristics of epithelial ovarian cancer patients. RESULTS： The positive expression rates of FoxM1，E-cadherin and vimentin in ovarian cancer tissues were 95.12%，82.93% and 70.73% respectively，and the high expression rates were 51.22%，53.66% and 46.34% respectively. The positive expression and high expression rates of FoxM1 in ovarian cancer tissues were significantly higher than those in the borderline ovarian cancer tissues and normal ovarian tissues (P<0.05). The expressions of FoxM1，E-cadherin and vimentin in ovarian cancer tissues were related to the clinical stages and lymph node metastasis of patients. The high expression rates of FoxM1 and vimentin in ovarian cancer tissues of patients with clinical stage III-IV and lymph node metastasis were significantly higher than those of patients with stage I-II and with no lymph node metastasis，respectively. The high expression rates of E-cadherin were significantly lower than those of patients with stage I-II and no lymph node metastasis (P<0.05). Spearman analysis showed that there were significant and positive correlations between FoxM1 and vimentin expression levels in epithelial ovarian cancer tissues(φ=0.562，P<0.05)，but there were significant and negative correlations between them and E-cadherin expression levels (φ=-0.440，-0.366，P<0.05). CONCLUSION： Expression of FoxM1 was significantly up regulated in epithelial ovarian cancers，and was related to clinical stages and lymph node metastasis. FoxM1 may therefore promote the occurrence，progression and metastasis of epithelial ovarian cancer via the EMT process.

OBJECTIVE： To investigate biological functions and mechanism of ribonucleotide reductase subunit M2 (RRM2) in medulloblastoma. METHODS： Relationships between survival and the levels of RRM2 expression in GEO database (GSE85217) were analyzed. Expressions of RRM2 in medulloblastoma and normal cerebellar tissues were analyzed using immunohistochemistry and database. CCK-8 proliferation experiments，transwell experiments，and flow cytometry were used to study the effects of RRM2 on proliferation，migration，cell cycle，and apoptosis of medulloblastoma cell lines Daoy and D283. RNA sequencing (RNA-seq) was used to explore downstream molecular mechanisms. Paired t-test and independent sample t-test were used for statistical analysis. RESULTS： Through the analysis of GEO database，survival of medulloblastoma patients was worse in the RRM2-high group than that in low group (P<0.05). The results of immunohistochemistry and database showed that expressions of RRM2 were significantly higher in medulloblastoma than that in normal cerebellar tissues (P<0.05). Compared to the control groups，overexpressions of RRM2 promoted tumor cell proliferation and migration，while knockdown of RRM2 reduced tumor cell proliferation and migration. In addition，overexpressions of RRM2 significantly increased the proportions of cells in the S phase (P<0.05) and reduced apoptosis (P<0.05). After knockdown of RRM2，RNA-seq results showed that 3 454 genes were up-regulated and 3 332 genes were down-regulated. KEGG analysis showed that multiple signal pathways were affected，including FOXO，cell cycle，ribosome biosynthesis，etc. CONCLUSION： RRM2 was highly expressed in medulloblastoma and the expression was negatively related to prognosis. In addition，RRM2 expression promoted proliferation and migration of medulloblastoma cells and reduced apoptosis.

OBJECTIVE： To investigate expression levels of the AMPK in Kazakh esophageal squamous cell carcinoma，its role in lipid metabolism reprogramming，and its relationship with clinicopathological features and prognosis. METHODS： Sixty-five Kazakh patients with esophageal squamous cell carcinomas were selected. AMPK mRNA expression levels in the cancer cells and the adjacent normal tissues were detected using qPCR. AMPK protein expressions were detected using immunohistochemistry. Kaplan-Meier method was used to draw the survival curve，and to analyze the correlation between AMPK expression and the prognosis of the carcinomas. Cox proportional hazards regression analysis was used to analyze the risk factors affecting the poor prognosis of the cancer patients. AMPK was stably knocked down in the esophageal cancer cell line，KYSE150，and lipid qualitative and quantitative analyses were performed using ultra performance liquid chromatography-mass spectrometry. RESULTS： The mRNA levels and protein positive rates of AMPK in the carcinoma tissues were significantly higher than those in adjacent tissues (P<0.05). The expression level of AMPK was correlated with the degree of differentiation，lymph node metastasis and TNM stage (P<0.05). The expression levels of AMPK were correlated with the degrees of differentiation，lymph node metastasis and TNM stages (P<0.05). The positive expression of AMPK was a reliable risk factor for poor prognosis of esophageal squamous cell carcinoma among Kazakhs. Through the absolute quantitative analysis of lipid content in esophageal cancer cells，it was found that the lipid content and its changing trend were different in different lipid grades. A total of 10 lipid subclasses were identified，consisting of fatty acids (7.88%)，glycerides (60.77%)，glycerides sphingolipids (21.35%)，sphingolipids (9.62%) and sterol lipids (0.38%). AMPK induced significant changes in lipid metabolism in esophageal cancer cells. A total of 76 differential lipid metabolites were found，among which 37 were up-regulated and 29 were down-regulated. The up-regulation of triglyceride (TAG) and the down-regulation of sphingolipid (SM)，diglyceride (DAG) and phosphatidyl ethanolamine (PE) were observed. Targeted lipid quantitative analysis results suggest that AMPK might be involved in lipid metabolic reprogramming in Kazakh patients with esophageal squamous cell carcinoma. CONCLUSION： The mRNA level and protein positive rate of AMPK in esophageal squamous cell carcinoma tissues were significantly higher than those in adjacent tissues (P<0.05). The expression level of AMPK was correlated with the degree of differentiation，lymph node metastasis and TNM stage (P<0.05). The positive expression of AMPK was a reliable risk factor for poor prognosis of Kazakh esophageal squamous cell carcinoma.

OBJECTIVE： To investigate effects of ulinastatin in combination with metal biliary stent on pancreatic cancers which were caused by extrahepatic biliary obstruction. METHODS： A total of 92 pancreatic cancer patients with extrahepatic biliary obstruction were selected and ewually divided into control and observation groups according to the random number table method. The control group was treated with metal biliary stent implantation alone，and the observation group was treated in combination with ulinastatin for two weeks. Before and after the treatments，concentrations of liver function total bilirubin (TBiL) and aspartate aminotransferase (AST)，and changes of blood uric acid (UA) and urea nitrogen (BUN) were determined. In addition，clinical efficacy and occurrence of adverse reactions after the treatment were compared between the two groups. RESULTS： After the treatment，AST and TBiL concentrations in the two groups were lower than before treatment (P<0.05)，and AST and TBiL concentrations in the observation group were lower than control group (P<0.05). After treatment，blood UA and BUN concentrations in the two groups were reduced (P<0.05)，and the observation group was lower than that in the control group (P<0.05). The total effective rates in the observation group (97.83%) was higher than that of the control group (82.61%) (P<0.05). During treatment，there was 1 case of nausea，1 case of vomiting and 1 case of diarrhea in the observation group，and the incidence of adverse reactions was 6.52% (3/46)，while 2 cases of nausea and 4.35% (2/46) were in the control group，there was no difference in the incidence of adverse reactions (correction value χ²=0.211，P=0.646). After treatment，the concentration of inflammatory factors in the two groups was lower than that before treatment (P<0.05)，and the concentration of inflammatory factors in the observation group was lower than that in the control group (P<0.05). CONCLUSION： Ulinastatin in combination with metal biliary stent implantation improved liver and kidney functions，enhanced efficacy and safety，reduced inflammatory factors，and improved immune functions among pancreatic cancer patients with extrahepatic biliary obstruction.

OBJECTIVE： To investigate relationships between expression of TNF-α and FOXA2 proteins in colorectal cancer tissues，and clinicopathologic indexes in the patients. METHODS： Thirty-nine cases of colorectal cancer and paired paracancer normal tissues (more than 5 cm from the tumor margin) were collected，and 42 cases of colorectal adenoma tissues which were confirmed by pathology and colonoscopy were selected. Immunohistochemistry was used to determine expression of TNF-α and FOXA2 proteins in colorectal tissue，using the Spearman method for evaluation. The Chi-square test was used to analyze relationships between the expression of TNF-α and FOXA2 protein，and clinicopathologic indexes in the colorectal cancer patients. RESULTS： The positive expression rates for the TNF-α protein in normal colorectal mucosa，colorectal adenoma and colorectal cancer tissues were 23.08%，50% and 76.92%，respectively，and the differences among the groups were statistically significant. The positive expression rates for the FOXA2 protein in the three tissues were 23.08%，52.38% and 79.49%，respectively，and the differences were statistically significant. There were statistically significant differences in expressions of the TNF-α and of the FOXA2 proteins among different diameter and histological types of adenomas (P<0.05). In colorectal cancer tissues，there were statistically significant differences in TNF-α protein expression among different degrees of differentiation，depth of invasion，lymph node metastasis and TNM staging，while there were statistically significant differences in FOXA2 protein expression among different degrees of differentiation，lymph node metastasis and TNM staging (P<0.05). The expressions of TNF-α and FOXA2 were positively correlated with rs=0.475 (P<0.05). CONCLUSION： The positive expression rates of TNF-α and FOXA2 proteins were different in normal intestinal mucosa，colorectal adenoma and colorectal cancer tissues，with gradually increasing trend，suggesting that TNF-α and FOXA2 proteins were closely related to the occurrence and development of colorectal cancers. Higher positive expression rates of TNF-α were associated with worse tumor differentiation，deeper depth of invasion，higher rates of lymph node metastasis and later TNM stages. The strong associations suggest that TNF-α protein played a promoting role in the occurrence and development of colorectal cancers. Similarly，higher the FOXA2 positive expression rates were associated with worse tumor differentiation，higher lymph node metastasis rates and later TNM stages，suggesting that FOXA2 protein was also involved with the occurrence and development of colorectal cancers.

OBJECTIVE： According to the OECD 487 Guidelines for testing of chemicals，the in vitro mammalian cell micronucleus test was used to determine whether the electronic cigarette liquid was mutagenic or not. METHODS： The doses for the liquid were 1.25，2.50 and 5.00 μL/mL. Solvent and positive controls were included，and replicate cultures were set up per dose in the test. The CHL cells were exposed to the liquid in the presence or absence of an exogenous metabolic activation system (S9)，for 4 and 24 hours of incubation. Then，they were harvested，exposed to hypotonic solution and fixed. The cells were then stained with acridine orange and observed under fluorescence microscopy. For each dose group，2 000 binucleated cells were selected for micronucleus rate analysis. RESULTS： After incubation for 4 hours with the dose levels of 1.25，2.50 and 5.00 μL/mL，the induced micronucleus rates without S9 (4 h，-S9)were 2.50‰，1.50‰ and 1.50‰ respectively；and 1.50‰，2.00‰ and 1.00‰ respectively with S9 (4 h，+S9). After 24 hours treatment without S9 (24 h，-S9)，the rates were 1.50‰，1.00‰ and 1.00‰，respectively.. Compared with the solvent control group，there was no statistically significant difference observed (P>0.05). CONCLUSION： Under the current test conditions，electronic cigarette liquid showed no evidence of mutagenic potential in the CHL at the tested dose level of 1.25-5 mg/mL.

OBJECTIVE： To assess whether there was a teratogenic risk from Cyperus esculentus on SPF grade pregnant SD rats. METHODS： For this study，100 3-month-old SPF grade and pregnant rats were used. They were weighed on the 0^th，6^th，9^th，12^th，15^th，and 20^th days of gestation. On the 6^th to 15^th days of gestation，rats were fed at four concentration levels of Cyperus esculentus：0 (CK)，2，4，and 8 g/kg body weight. The pregnant rats were sacrificed using CO₂ anesthesia on the 20th day of gestation. The uterus was taken out by laparotomy，and the weights of the uteri and fetuses were measured. The numbers of corpora lutea，implantation，absorption，early death，late death，and live fetus were recorded and examined. The fetal weights and body lengths were recorded for live fetal rats，and they were examined for abnormalities. RESULTS： For the pregnant rats，no abnormal physical signs were found，their general activities were normal，and no significant abnormalities were found in gross anatomy at the 20^th day of execution. Results from analysis of the data show that Cyperus esculentus had no effect on the general situation，body weight and weight gain，embryonic development，fetal mouse appearance abnormalities，fetal mouse bone development，skeletal teratogenesis rate，fetal mouse visceral development，teratogenesis rate，and total teratogenesis rate of pregnant rats. CONCLUSION： Under our experimental conditions，there was no difference in abnormalities among the different exposure dose groups and from the controls. Thus，it can be determined that Cyperus esculentus did not cause teratogenic risk under our experimental conditions.