[Protective effect of anti-idiopathic pulmonary fibrosis drug Pirfenidone and Sufenidone (SC1011) on pulmonary injury induced by tuberculosis in a mouse tuberculosis model].

Objective: To evaluate the protective effect of anti-idiopathic pulmonary fibrosis (IPF) marketed drug Pirfenidone and its clinical drug Sufenidone (SC1011) against lung injury in a mouse tuberculosis model. Methods: C57BL/6 mouse model of tuberculosis was established. A total of 75 C57BL/6 mice were infected with 1×107 CFU/ml H37Rv suspension by aerosol and randomly divided into untreated (n=9) group, isoniazid+rifampicin+pyrazinamide (HRZ) group (n=22), PFD+HRZ group (n=22), and SC1011+HRZ group (n=22). C57BL/6 mice were infected with H37Rv by aerosol for 6 weeks and then treated. Seven mice in each treatment group were weighed, sacrificed, dissected and observed for lung and spleen lesions at 4 and 8 weeks of treatment. HE staining and Masson staining were used to assess degree of lung injury and fibrosis, respectively. ELISA was used to assess the IFN-γ/TNF-α content in the serum of mice in each treatment group after 4 weeks of treatment. Hydroxyproline (HYP) content in lung tissue was measured by alkaline hydrolysis; meanwhile, CFU counts were used to assess the bacterial load in the lung and spleen of mice in each treatment group and the recurrence of spleen and lung tissue after 12 weeks of drug withdrawal. Results: At 8 weeks, the HYP content in the lung tissue was (630±58), (635±17), and (840±70) µg/mg in the PFD+HRZ, SC1011+HRZ, and HRZ treatment group, respectively (P<0.05).At 8 weeks, the proportion of Masson staining blue-stained area, that was, positive area, in lung tissue was 16.65%±1.82%, 10.01%±2.16%, and 21.36%±3.21%, respectively (F=27.11, P<0.001).The lung injury scores by HE staining at 8 weeks were (5.00±0.50), (5.00±0.47), and (6.89±0.99) points, respectively (F=19.81, P<0.001).The results of 4-week ELISA showed that the levels of TNF-α and IFN-γ in the serum of the SC1011+HRZ-treated group were lower than those of the HRZ-treated group (all P<0.05).The degree of lung injury and fibrosis in PFD+HRZ and SC1011+HRZ treatment groups were lower than those in HRZ treatment group (all P<0.001). The number of viable bacteria in the lung tissue of mice treated with PFD+HRZ, SC1011+HRZ, and HRZ for 4 weeks was lower than that of mice untreated [(1.82±0.10), (1.91±0.05), (1.79±0.17) vs. (5.27±0.07) lg(CFU+1)/ml, all P<0.05)]. And the aseptic transformation of the spleen of mice was achieved in each treatment group at 8 weeks of administration. After 12 weeks of drug withdrawal, the recurrence of lung infection in the SC1011+HRZ treatment group was 3/7 lower than 5/7 in the HRZ treatment group (P>0.05); the recurrence of spleen infection in the SC1011+HRZ treatment group was 1/7 lower than 5/7 in the HRZ treatment group (P>0.05).Pulmonary infection recurred more frequently in PFD+HRZ 6/7 versus HRZ 5/7 (P>0.05). Conclusions: PFD/SC1011, when combined with HRZ, reduced lung injury and reduced secondary fibrosis in pulmonary tuberculosis in C57BL/6 mice. SC1011 combined with HRZ has no significant short-term therapeutic effect on MTB, but may reduce its recurrence rate in long-term treatment, especially in reducing the recurrence rate of mouse spleen.

[Changing trend of incidence and mortality of esophageal cancer during 2010-2016 in Henan Province, China].

Objective: To estimate the incidence and mortality of esophageal cancer in 2016 and their changing trend during 2010-2016 according to the cancer registration data in Henan province. Methods: The data quality including completeness, validity, and reliability of local registries which submitted the cancer registration data of 2016 were assessed according to the criteria of Guideline on Cancer Registration in China and IARC/IACR. Esophageal cancer cases (ICD10: C15) were extracted from the database, and the incidence and mortality stratified by gender, age, and areas (urban/rural) were calculated, the incidence and mortality of provincial cancer were estimated combined with provincial population data. China's 2000 census population and Segi's population were used to calculate the age-standardized rate. Joinpoint model was used to estimate the changing trend of age standardized incidence and mortality along with the calendar year. Results: Approximately 40.10 thousand new esophageal cancer cases were diagnosed in Henan in 2016, accounting for 13.46% of all new cancer cases, and it ranked the third among cancer of all sites. The crude incidence of esophageal cancer was 37.21/100 000 with an age-standardized incidence rate by China standard population (ASIRC) of 26.74/100 000 and an age-standardized incidence rate by world standard population (ASIRW) of 27.12/100 000. The incidence of esophageal cancer in males was higher than that in females, with the ASIRC of 34.53/100 000 and 19.19/100 000, respectively. It was higher in rural areas than that in urban areas, with the ASIRC of 28.13/100 000 and 20.90/100 000, respectively. About 29.30 thousand deaths of esophageal cancer in Henan in 2016, accounting for 15.61% of all cancer deaths in Henan, which ranked the third among cancer of all sites. The crude mortality rate was 27.14/100 000 with an age-standardized mortality rate by China standard population (ASMRC) of 18.74/100 000 and an age-standardized mortality rate by world standard population (ASMRW) of 18.78/100 000. The mortality in males was higher than that in females, with the ASMRC of 24.78/100 000 and 13.12/100 000, respectively. It was also higher in rural areas than that in urban areas, with the ASMRC of 19.48/100 000 and 15.73/100 000, respectively. The ASIRC and ASMRC were declining with annual percent change (APC) of 3.12% (APC=-3.12%; 95%CI: -5.30%, -0.90%; P=0.015) and 2.47% (APC=-2.47%; 95%CI: -4.70%, -0.20%; P=0.039) during 2010-2016. However, the significant declining trend was only observed in rural areas in Henan, and the changing trend was same between males and females. Conclusions: The incidence and mortality of esophageal cancer are declining since 2010, however, the disease burden remains large in Henan. Therefore, comprehensive prevention and control efforts should be strengthened according to its epidemic characteristics and risk factors.

[Changing trend of incidence and mortality of stomach cancer during 2010-2016 in Henan Province, China].

Objective: To estimate stomach cancer incidence and mortality in Henan, 2016 and analyze the trend of stomach cancer incidence and mortality from 2010 to 2016. Methods: Stomach cancer related data in 2016 was extracted from Henan cancer registration and follow-up system. All data were qualified in validity, reliability and completeness according to the Guideline on Cancer Registration in China and International Agency for Research on Cancer (IARC/IACR). The incidence and mortality of stomach cancer were estimated by areas, gender and age based on the quality data and the registered population data of Henan province in 2016. The epidemic trend of stomach cancer was also been evaluated based on the age-standardized incidence and mortality by Chinese population (ASR China) from 2010 to 2016. Results: In 2016, the estimated incident cases of stomach cancer were 44 311. The incidence was 41.07/100 000, ASR China was 30.17/100 000, ASR by world population (ASR world) was 30.36/100 000, and the cumulative incidence rate was 3.84%. The incidences of male and female were 55.65/100 000 and 25.35/100 000, respectively. Meanwhile, 32 927 people died of stomach cancer in Henan. The mortality was 30.52/100 000, ASR China was 21.45/100 000, ASR world was 21.54/100 000, and the cumulative mortality was 2.53%. From 2010 to 2016, both the ASR China for incidence and mortality of stomach cancer in Henan showed a steady downward trend. In rural, the ASR China for incidence and mortality decreased rapidly, while the stable trend was observed in urban. Nevertheless, the incidence and mortality of stomach cancer in rural were still higher than those in urban. Conclusions: The incidence and mortality of stomach cancer in Henan province showed steadily declining trend from 2010 to 2016, and the geographical distribution difference between rural and urban areas was gradually narrowing. However, the disease burden was still high in 2016.

[Research progress of airway microbiome and airway immune response in asthma].

Bronchial asthma (asthma) is one of the most common chronic airway diseases, with more than 300 million people worldwide suffering from this disease. In recent years, studies have shown that compared with healthy people, the airway microecological structure and relative abundance of various flora of asthmatic patients have changed, and are related to the airway inflammatory phenotype of asthma. Airway microecology can affect the occurrence and development of asthma through immune response. The mechanism of interaction between airway microecology and asthma can provide new ideas for the accurate treatment of asthma. This article mainly reviewed the current research on airway microecology in asthma, and puts forward prospects for the accurate treatment of asthma in the future.

[Compliance rate and impact factor analysis of liver cancer screening in urban areas of Henan Province].

Objective: To evaluate the compliance rate and its impact factors of liver cancer screening for high-risk groups in urban areas of Henan Province from 2013 to 2019. Methods: Residents of 40-74 years old in 8 cities of Henan province were selected to investigate the risk factors and liver cancer risk assessment. Subjects with high risk of liver cancer received AFP combined ultrasonography for screening. Chi-square tests were used to compare the differences in liver cancer screening participation rates between groups. Multivariate logistic regression models were applied to explore the potential factors correlating to the compliance of liver cancer screening. Results: Overall, 3 6781 participants who met the inclusion criteria were included in this analysis, and 17 241 of them took the following liver cancer screening, yielding a participation rate of 46.87%. The participation rate varied greatly across cities, ranging from 62.50% to 38.59%. Moreover, the participation rate varied greatly across periods, ranging from 52.77% in 2014-2015 to 38.14% in 2013-2014. The multivariate Logistic regression analyses showed that: female, older, high education degree, cigarette and alcohol intake, infrequent physical exercise, chronic hepatitis B, chronic hepatitis C, fatty liver, gallstones and a family history of liver cancer were inclined to accept liver cancer screening (P<0.05). Conclusions: The overall participation rate of liver cancer screening among high-risk population is less than 50% in urban areas of Henan Province. Implement of effective interventions targeting the specific high-risk populations might improve the overall compliance rate of liver cancer screening in the future.

[Clinical characteristics of 21 infertile women with non-classic 21-hydroxylase deficiency].

Objective: To investigate the clinical features of infertile women with non-classic 21-hydroxylase deficiency (21-OHD). Methods: The study enrolled 21 infertile women with non-classic 21-OHD in Sun Yat-sen Memorial Hospital of Sun Yat-sen University from January 2009 to December 2018. The clinical presentation, endocrine hormone, glucolipid metabolism and treatment outcome were retrospectively analyzed. The diagnosis of non-classic 21-OHD was comprehensively based on clinical and hormonal characteristics. Results: Among 21 cases, the age was (29.9±2.9) years, the mean age at menarche was (13.6±2.0) years, body mass index was (22.1±2.9) kg/m2, and 38% (8/21) had oligomenorrhea. Hirsutism was diagnosed in 3 cases (14%, 3/21). Clitoromegaly was seen in 14% (3/21) and polycystic ovarian morphology was found in 33% (7/21) of the patients. The mean serum level of basal progesterone was (11.3±21.0) nmol/L, with 48% (10/21) having high basal progesterone level; after therapy by glucocorticoid, the level of progesterone was (1.9±2.0) nmol/L. Serum 17-hydroxyprogesterone concentration was (66.4±123.6) nmol/L; after therapy by glucocorticoid, it was (2.4±1.8) nmol/L. In the study increased testosterone, androstenedione and dehydroepiandrosterone sulfate were present in 62% (13/21), 52% (11/21) and 43% (9/21), respectively; and 52% (11/21) of patients manifested androgen excess and basal progesterone elevation; androgen levels decreased after therapy by glucocorticoid. The pregnancy rate was 76% (16/21). Out of 19 pregnancies, 6/19 ended in spontaneous miscarriages. Conclusions: Infertile women with non-classic 21-OHD are characterized by hyperandrogenism and basal progesterone elevation, whereas gonad axis disorder is not apparent. After no response to conventional therapy, adult infertile women with non-classic 21-OHD could achieve a desirable pregnancy outcome with proper treatment of glucocorticoid.

[Analysis of detection rate and compliance of colorectal cancer screening with colonoscopy in urban areas of Henan Province, 2013-2019].

Objective: To evaluate the compliance of colonoscopy screening and the detection rate of colorectal lesions with colonoscopy in urban areas of Henan province from 2013 to 2019. Methods: The study objects were derived from the Cancer Screening Program in Urban Henan Province, China. From October 2013 to October 2019, 282 377 residents, who had lived in the local area for more than three years, were recruited from Zhengzhou, Zhumadian, Anyang, Luoyang, Nanyang, Jiaozuo, Puyang and Xinxiang. The cancer risk assessment questionnaire was used to collect basic demographic characteristics, eating habits, living environment and habits, psychology and emotions, past disease history and family history of cancer, and women's physiological and reproductive history. The cancer risk assessment model was used for evaluating the risk of colorectal cancer. The data related to colonoscopy screening was obtained from the participating hospitals. Chi-square test was used to analyze the compliance rate among different groups. Results: The 282 377 subjects were (55.26±8.68) years old, of which 44.80% (126 505) were males, and 67.63% (190 694) had junior/senior/tertiary education background. A total of 39 834 (14.11%) subjects were assessed to be at high risk for colorectal cancer, of which 7 454 took the following colonoscopy screening, yielding a participation rate of 18.71%. The screening rate was higher in those aged 50 to 54 (20.42%) and 55 to 59 (20.43%) years, and lowest in those aged 70 years and older (12.30%) (P<0.001). A total of 17 colorectal cancer cases were detected, with a detection rate of 0.23% (17/7 454). Conclusion: Colonoscopy as a means of screening for colorectal cancer is helpful for early detection of colorectal lesions, but the participation rate is poor.

Effect of dietary fibre and grit on performance, gastrointestinal tract development, and grit pattern of goose.

1. This study aimed to investigate effects of dietary fibre and grit on growth performance, gastrointestinal tract development, and gizzard grit retention of geese. 2. The trial had a 3 × 2 factorial design consisting of three levels of dietary crude fibre (CF, 4%, 7% and 10%, adjusted by grass powder), with or without grit addition (1-4 mm river sand). 3. In total, 648, 22-d-old male goslings were randomly allotted to six treatments (six pens/treatment). At 49 d and 70 d of age, one goose per pen was euthanised to collect samples. 4. The birds fed 10% CF had decreased feed conversion ratio (FCR) during 22-49 d, but this effect was less in older geese. Increasing dietary CF levels increased relative weights of gizzards for geese aged 49 d and 70 d. Grit addition decreased the relative weights of gizzard and duodenum of geese aged 49 d. The gizzard of geese could selectively retain grit from feed even with no grit added. With adequate supply, most grit in gizzard was 0.45-3 mm in size. 5. In conclusion, supplement of CF and grit mainly affected gastrointestinal tract, and the amount of CF affected FCR. Geese aged 22-49 d could utilise dietary CF levels of 4%-7% and older birds could feed on diets with up to 10% CF. The gizzard of goose selectively retained grit of a particle size of 0.45-3 mm.

[Analysis of the efficacy of lung cancer screening in urban areas of Henan Province by low-dose computed tomography from 2013 to 2017].

Objective: To evaluate the efficacy of lung cancer screening in urban areas of Henan province by low-dose computed tomography (LDCT) from 2013 to 2017. Methods: A cluster sampling method was used to select the residents of 40-74 years old in Henan province to investigate the risk factors and conduct lung cancer risk assessment. Subjects with high risk of lung cancer received LDCT for screening. Results: A total of 179 002 residents completed the lung cancer risk assessment, and 35 672 subjects were identified as high risk of lung cancer, with a high risk rate of 19.93%. A total of 13 383 subjects with high risk received LDCT, and the screening rate was 37.52%. There were 786 cases diagnosed as positive nodules, and the detection rate was 5.87%. Among them, 755 cases of solid/partial solid nodule were ≥5 mm, 23 cases of non-solid nodules were ≥8 mm, 8 cases were intratracheal nodules, and 115 cases were diagnosed as suspicious lung cancer. The detection rate in males was 6.74%, which was higher than 5.02% in females. The detection rate was positively related with age (P<0.05). Conclusions: The application of LDCT is a useful screening method which can elevate the early detection rate of positive nodules and other related diseases in lungs. In the future, males and older populations should be paid more attention to improve screening efficacy.

[Analysis of endoscopic screening compliance and related factors among high risk population of upper gastrointestinal cancer in urban areas of Henan Province from 2013 to 2017].

Objective: To study the compliance of endoscopic screening for high-risk population of upper gastrointestinal cancer and relevant factors in urban areas of Henan Province, 2013-2017. Methods: The study participants were from the Cancer Screening Program in Urban Henan Province, China. From October 2013 to October 2017, 43 423 residents, who were evaluated as high-risk population for upper gastrointestinal cancer, were recruited from Zhengzhou, Zhumadian and Anyang. The cancer risk assessment questionnaire was used to collect basic demographic characteristics, dietary habits, living environment and habits, psychology and emotions, disease history and family history of cancer, and women's physiological and reproductive history. The data of endoscopic screening was obtained from hospitals participating in the Cancer Screening Program. Multivariate logistic regression model was applied to explore potential factors related to the compliance of endoscopic screening. Results: The age of study participants was(55.49±8.15) years old, and 44.00% (19 105) were male. About 18.41% of study subjects (7 996) took the endoscopic screening. The multivariate logistic regression analysis showed that females, individuals aged 45-64 years old, with junior high school education or above, unmarried/divorced/widowed, previous smoking, alcohol drinking, infrequent physical exercise, history of reflux esophagitis, history of superficial gastritis, history of gastric ulcer, history of duodenal ulcer, history of gastric polyps and family history of upper gastrointestinal cancer were more likely to accept endoscopic screening. Conclusion: The overall participation rate of endoscopic screening among high-risk population of upper gastrointestinal cancer was still low in urban areas of Henan Province. Gender, age, education, marital status, smoking, alcohol consumption, physical activity, history of upper gastrointestinal disease and family history of upper gastroin testinal cancer were associated with the compliance of endoscopic screening.

[Aldo-keto reductase family 1 B10 participates in the regulation of hepatoma cell cycle through p27/p-Rb signaling pathway].

Objective: Aldo-keto reductase family 1 member B10 (AKR1B10) pathogenesis, early diagnosis and prognosis are closely related with hepatoma. Therefore, this study explores the effect and mechanism of AKR1B10 on cell cycle in hepatoma cells. Methods: HepG2 cells were infected with lentivirus LV-AKR1B10-shRNA or treated with epalrestat, an AKR1B10 inhibitor. The expression level of AKR1B10 was detected by Western blot assay and real-time fluorescence quantitative PCR (RT-qPCR). Decreased AKR1B10 activity was detected by reduced coenzyme II (NADPH) absorbance at 340 nm. The low expression of AKR1B10 and the effect of different concentrations of epalrestat on cell proliferation and cell cycle were detected by CCK-8 method and flow cytometry. The protein expression levels of p-rb, cyclin D1, E1, p27 in HepG2 cells were detected by Western blot. The mean of the two samples was tested using independent sample t-test. Results: AKR1B10 expression level in hepatoma cells was significantly increased compared to normal liver cells, and the relative expression level of AKR1B10 protein in HepG2 cells was 6.71 ± 1.11 (P = 0.012). Epalrestat was significantly inhibited with the enzymatic activity of AKR1B10 in a dose-dependent manner. AKR1B10 gene in HepG2 cells was effectively silenced. HepG2 cells treated with different concentrations of epalrestat (AKR1B10 inhibitor) for 24, 48 and 72 h had inhibited cell proliferation, promoted G0/G1 cell cycle arrest, reduced the expression of p-Rb, cyclin D1, and cyclin E1 and increased the expression of cyclin dependent kinase inhibitor p27 expression. Conclusion: AKR1B10 inhibitory expression and activity can promote G0/G1 cell cycle arrest in HepG2 cells through the p27 / p-Rb pathway.

[AKR1B10 inhibitor enhances the inhibitory effect of sorafenib on liver cancer xenograft].

Objective: To investigate the inhibitory effect of AKR1B10 inhibitor combined with sorafenib on hepatocellular carcinoma (HCC) xenograft growth. Methods: HepG2 xenograft model was established in nude mice. The mice were then randomly divided into four groups: control group, epalrestat monotherapy group, sorafenib monotherapy group and combination treatment group. Tumor volume, tumor weight, T/C ratio and the change in body weight of nude mice in each group were compared to evaluate the curative effect. Immunohistochemistry staining was used to detect the expression of Ki-67 in tumor tissues to evaluate the proliferation status of tumor cells. One-way analysis of variance was used to compare the differences between the groups. Student's t-test was used to test means of two groups and chi-square test was used for multiple samples. Results: The differences of the grafted tumor volume before and after treatment between the control group, epalrestat group, sorafenib group and combined therapy group was 238.940 ± 39.813, 124.991 ± 84.670, -26.111 ± 11.518, and -54.072 ± 17.673(mm(3)), respectively, (F = 37.048, P < 0.001). The tumor mass were 0.273 ± 0.140, 0.158 ± 0.078, 0.079 ± 0.054, 0.045 ± 0.024 (g), (F = 16.594, P < 0.001); T/C ratio were 100%, 57.9%, 28.9%, 16.5%, and Ki-67 positive rate were 23.295 ± 6.218, 13.503 ± 3.392, 7.325 ± 2.257, 4.664 ± 1.189 (%), (χ(2) = 822.203, P < 0.001) . The tumor volume (t = -3.579, P = 0.002) and Ki-67 positive rate (t = -10.003, P < 0.001) in epalrestat monotherapy group were significantly lower than control group. The tumor volume (t = 2.056, P = 0.025), tumor mass (t = 2.101, P = 0.043), and Ki-67 positive rate (t = -2.850, P = 0.005) in combination treatment group were significantly lower than sorafenib monotherapy group. Compared with the control group, the body weight of nude mice in the treatment group decreased to a certain extent, but there was no statistically significant difference between epalrestat monotherapy group and control group (t = -1.599, P = 0.262), and combined therapy and sorafenib monotherapy group (t = -0.051, P = 0.96). Conclusion: AKR1B10 inhibitor enhanced the inhibitory effect of sorafenib on hepatocellular carcinoma xenograft.

[Clinical value of (125)I radioactive seed implantation plus transcatheter arterial chemoembolization combined with radiofrequency ablation in treating patient with sub-capsular hepatocellular carcinoma].

Objective: To analyze the clinical efficacy and safety of (125)I radioactive seed implantation in the treatment of sub-capsular hepatocellular carcinoma (sub-HCC) with sequential radiofrequency ablation and transcatheter arterial chemoembolization (TACE). Methods: The clinical data of 76 cases with advanced HCC with sub-capsular nodules including 68 males and 8 females, with an average age of (58±9) years, ranging from 33 to 78 years, enrolled in Lishui Central Hospital from January 2010 to December 2016 were collected.The average maximum diameter of tumor is (5.7±2.3) cm, ranging from 3.1 cm to 12.0 cm.The patients were divided into TACE+ RFA group and (125)I + TACE+ RFA group with 38 cases in each group.The overall survival (OS) and progression free survival(PFS) were calculated.The clinical efficiency and adverse events were evaluated. Results: The disease control rate were 84.2%(32/38) in (125)I + TACE+ RFA group and 63.2% (24/38) in TACE+ RFA group, χ(2)=4.34, P= 0.04.The median PFS were 18 months in (125)I + TACE+ RFA group and 11 months in TACE+ RFA group, χ(2)=4.84, P=0.03.The FPS cumulative rate in (125)I + TACE+ RFA group were higher than that in TACE+ RFA group at 6 months (94.7%±3.6% vs 81.3%±6.4%, Z=24.1>2.58, P=0.00), 1 year (89.2%±5.1% vs 40.7%±8.3%, Z=13.3>2.58, P=0.00) and 2 year (55.9%±8.6% vs 29.6%±8.2%, Z=7.2>2.58, P=0.00). The median OS were 42 months in (125)I + TACE+ RFA group and 30 months in TACE+ RFA group, χ(2)=4.76, P=0.029.The survival cumulative rate in (125)I+ TACE+ RFA group were higher than that in TACE+ RFA group at 1 year (92.1%±4.4% vs 83.8%±6.1%, Z=23.5>2.58, P=0.00), 2 year (75.8%±7.0% vs 59.8%±8.4%, Z=12.43>2.58, P=0.00), 3 year (59.0%±8.2% vs 41.7%±8.9%, Z=8.3>2.58, P=0.00), 5 year (34.2%±8.2% vs 18.2%±8.1%, Z=5.5>2.58, P=0.00). In addition, there was no statistical difference in liver function and complications between TACE+ RFA group and (125)I+ TACE+ RFA group. Conclusion: (125)I radioactive seed implantation plus TACE combined with RFA treatment is an effective and safe treatment for sub-capsular hepatocellular carcinoma.

[Effect of transcatheter arterial chemoembolization combined with thymosin alpha 1 on the autophagy of immune cells from advanced hepatocellular carcinoma].

Objective: To investigate the effect of transcatheter arterial chemoembolization(TACE)combined with thymosin alpha1(Tα1)on the autophagy of immune cells from advanced hepatocellular carcinoma. Methods: A total of 30 patients with advanced liver cancer enrolled in Lishui Central Hospital from September 2015 to June 2016 were collected in this study. The average age of patients was 16-75(56±12) years. All patients were treated with TACE after enrolled in hospital in a week. Patients were divided into TACE group and TACE+ Tα1 treatment group(15 cases in each group). Patients in TACE group received a conventional treatment, without any immunotherapy, while the TACE+ Tα1 treatment group accepted TACE following a subcutaneously injection of 1.6 mg Tα1 twice a week for 4 weeks. Flow cytometry was used to detect the T cell subsets in two groups both before and after TACE treatment for 1, 4 weeks and at 3 months follow-up. Peripheral blood mononuclear cells (PBMC) were isolated by density gradient centrifugation. The expression of Beclin-1, LC3 protein and mRNA were detected by Western blot (WB) and PCR respectively. Results: There was no statistical difference of the percentage of CD3(+) , CD4(+) , CD8(+) T cell subsets and Beclin-1, LC3 protein and mRNA expression between the two groups before TACE treatment (P>0.05). The percentage of CD3(+) , CD4(+) , CD8(+) T cell subsets in TACE+ Tα1 group at 1 week post-TACE treatment (58.45%±16.34%, 38.33%±15.16%, 27.31%±12.54%), at 4 weeks post-TACE treatment (62.38%±18.62%, 43.19%±13.86%, 29.54%±10.33%) and 3 months follow-up (64.15%±13.76%, 41.28%±14.65%, 29.38%±15.65%) were statistically higher than those in TACE group at 1 week post-TACE treatment (53.71%±11.17%, 32.12%±10.53%, 24.45%±13.72%) at 4 weeks post-TACE treatment (52.12%±14.26%, 31.16%±15.43%, 23.39%±15.33%) and 3 months follow-up (54.28%±13.15%, 32.17%±14.98%, 24.34%±14.12%) (P<0.05). The Beclin-1, LC3 protein and mRNA expression in TACE+ Tα1 group at 1 week post-TACE treatment (protein: 0.57±0.08, 2.26±0.36, mRNA: 0.62±0.11, 2.69±0.27), at 4 weeks post-TACE treatment (protein: 0.66±0.09, 3.11±0.45, mRNA: 0.78±0.13, 3.43±0.61) were higher than those in TACE group at 1 week post-TACE treatment (protein: 0.45±0.16, 1.43±0.30, mRNA: 0.52±0.15, 1.15±0.37), at 4 weeks post-TACE treatment (protein: 0.51±0.13, 1.81±0.35, mRNA: 0.56±0.10, 1.98±0.41) ( P<0.05). But there was no statistically significant difference in the expression of Beclin-1 and LC3 in two groups at 3 months follow-up (P>0.05). Conclusions: TACE combined with Tα1 significantly increase the level of autophagy in the immune cells of patients with advanced primary hepatocellular carcinoma.

[The role of FOXO3a-Bim signaling in triptolide induced bladder cancer T24 cells apoptosis].

Objective: To investigate the role of FOXO3a-Bim signaling in triptolide induced bladder cancer T24 cells apoptosis. Methods: T24 cells were used and divided into control group, triptolide group(50 nmol/L), MK2206 group(50 nmol/L triptolide+ 5 µmol/L MK2206), FOXO3a-siRNA group(50 nmol/L triptolide+ 100 nmol/L FOXO3a-siRNA), Bim-siRNA group (50 nmol/L triptolide+ 100 nmol/L Bim-siRNA). MTT assay was used to analyze the cells growth inhibition.Annexin V/PI staining was implemented to detect cell apoptosis rate, the expression of p-Akt, Akt, p-FOXO3a, FOXO3a, Bim, Bax.Cleaved-caspase 3 was analyzed by Western blot. Results: After treatment with triptolide 25, 50, 100, 250 nmol/L, the cell growth inhibition rates at 24 hours(17%±9%, 24%±5%, 43%±8%, 61%±8%), 48 hours (20%±7%, 34%±6%, 56%±7%, 74%±5%) and 72 hours(32%±8%, 41%±7%, 69%±7%, 84%±3%) were significantly higher than control group respectively.The IC(50) at 24, 48, 72 hours were (113±10), (91±8), (68±5) nmol/L; the cell apoptosis rates at 24 hours (10%±4%, 15%±5%, 29%±8%, 46%±8%), 48 hours (16%±5%, 24%±6%, 40%±7%, 55%±9%) and 72 hours (27%±4%, 38%±5%, 50%±9%, 65%±8%) were significantly increased (P<0.05). Western blot showed that triptolide reduced the expression of p-Akt, p-FOXO3a and increased the expression of Bim, Bax, cleaved-caspase 3.The cell inhibition rate in Triptolide group (30%±8%) was significantly higher than that in the control group (P<0.05) and the rates in MK2206 group (54% ±6%), FOXO3a-siRNA group (18%±7%) and Bim-siRNA group (11%±6%) were also higher than the control group.Compared with the triptolide group, the inhibition rate in MK2206 group was significantly increased, but decreased in FOXO3a-siRNA group and Bim-siRNA group(P<0.05). Conclusion: Triptolide induces T24 cells apoptosis through FOXO3a-Bim signaling pathway.

[Effects and mechanism ofitraconazole on prostate cancer PC-3 cell apoptosis].

Objective: To investigate the effects and mechanisms of itraconazole on prostate cancer PC-3 cells proliferation. Methods: The PC-3 cells were divided into four group: control group, itraconazole group, itraconazole+ CerS-1-shRNA group and itraconazole+ PDMP group.3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide(MTT)assay was used to detect the growth of PC-3 cells.Apoptosis was detected by Annexin V-FITC /PI.The intracellular ceramide production was assayed by high performance liquid chromatography(HPLC). The expression of Bax, Bcl-2, cleaved-caspase3 and the expression and phosphorylation of Akt, mTORC1 were detected by Western blot. Results: After treatment with 0, 5, 10, 20 µmol/L itraconazole, apoptosis rate was 3.23%±1.32%, 5.87%±2.45%, 23.22%±5.29%, 48.57%±8.37%.The percentage content of ceramide was 100%, 109%±18%, 156%±12%, 197%±22%.Compared with the control group, there were statistically differences when the concentration of itraconazole were 10 and 20 µmol/L(all P<0.05). Western blot analysis showed that the Bax, cleaved-caspase 3 expression of itraconazole group and itraconazole+ PDMP group was significantly higher than control group, while Bcl-2 expression was significantly lower than the control; the Bax, cleaved-caspase 3 expression ofitraconazole+ CerS1-shRNA group was significantly lower than itraconazole group, while Bcl-2 expression was significantly higher than the itraconazole group.After 5, 10, 20 µmol/L itraconazole treatment, the expression of p-Akt and p-mTORC1 were significantly lower than the control group; the expression of p-Akt and p-mTORC1 in itraconazole+ CerS-1-shRNA group were significantly higher than itraconazole group. Conclusion: Itraconazole induces apoptosis of PC-3 cell through increasing the intracellular ceramide content, which might relate to upregulation of cleavage-caspase 3 and Bax, downregulation of Bcl-2 and inactivation of Akt-mTORC signal pathway.

Isolation and characterization of novel polymorphic microsatellite markers for Epinephelus akaara.

In the present study, ten novel microsatellite markers were developed from an enriched-(CA)13 genomic library of Epinephelus akaara. The mean number of alleles per locus was 21.6, with a range of 12 to 33. Observed heterozygosity ranged from 0.767 to 0.967, and expected heterozygosity ranged from 0.831 to 0.975, with mean values of 0.877 and 0.923, respectively. Among the ten loci, three loci deviated from Hardy-Weinberg equilibrium after sequential Bonferroni's correction. These polymorphic microsatellite markers may be useful for studies on the population genetics of E. akaara.

[Influence of family with sequence similarity 134, member B-mediated reticulophagy on lipopolysaccharide-induced apoptosis of mouse dendritic cells].

Objective: To investigate the influence of family with sequence similarity 134, member B (FAM134B)-mediated reticulophagy on lipopolysaccharide (LPS)-induced apoptosis of mouse dendritic cells (DCs), so as to provide a basis for improving the immune suppression of sepsis caused by wound infection and other factors. Methods: The experimental research methods were used. The DC line DC2.4 of the 3rd to 10th passage in the logarithmic growth stage was collected for experiments. DCs were divided into LPS stimulation 0 h (no stimulation) group, LPS stimulation 6 h group, LPS stimulation 12 h group, LPS stimulation 24 h group, and LPS stimulation 72 h group, which were cultured with 1 µg/mL LPS (the same concentration below) for the corresponding time. The protein expressions of FAM134B, microtubule-associated protein 1 light chain 3B (LC3B), and transporter protein SEC61B were determined by Western blotting, and the ratio of LC3B-Ⅱ/LC3B-Ⅰ was calculated (n=3). DCs were divided into phosphate buffer solution (PBS) group and LPS group for corresponding treatment. After 24 hours of culture, the expression of FAM134B and its co-localization with lysosomal probes and LC3B were detected using immunofluorescence method, while the number of autolysosomes in cells were observed through transmission electron microscope. DCs were divided into the FAM134B-knockdown group that were transfected with lentivirus containing small interfering RNA (siRNA) sequence of FAM134B gene and the empty vector group with empty lentivirus transfected. At post transfection hour 72, the fluorescence expression of cells was observed under the inverted fluorescence phase contrast microscope, meanwhile, the normally cultured DCs were set as blank control group, and the same observation was performed at the corresponding time point. DCs were divided into PBS alone group and LPS alone group, DCs successfully transfected with lentivirus containing siRNA sequence of FAM134B gene were divided into FAM134B-knockdown+PBS group and FAM134B-knockdown+LPS group, and DCs successfully transfected with empty lentivirus were divided into empty vector+PBS group and empty vector+LPS group. These cells were stimulated correspondingly and cultured for 24 hours. The protein expression of FAM134B was detected using Western blotting (n=3); the apoptotic rate of cells was determined by flow cytometry (n=3); the situation of apoptosis was observed by Hoechst staining, and the apoptotic rate was calculated (n=5); the protein expressions of cleaved cysteine aspartic acid specific protease-3 (caspase-3), B cell lymphoma 2 (Bcl-2), and Bcl-2-associated X protein (Bax) were detected using Western blotting, and the ratio of Bax/Bcl-2 was calculated (n=5). Data were statistically analyzed with one-way analysis of variance (ANOVA), least significant difference test, and ANOVA for factorial design. Results: Compared with those in LPS stimulation 0 h group, the protein expressions of FAM134B of cells in LPS stimulation 12 h group and LPS stimulation 24 h group were significantly increased (P<0.05), the protein expressions of SEC61B of cells in LPS stimulation 6 h group, LPS stimulation 12 h group, LPS stimulation 24 h group, and LPS stimulation 72 h group were significantly decreased (P<0.05), and the ratios of LC3B-Ⅱ/LC3B-Ⅰ of cells in LPS stimulation 24 h group and LPS stimulation 72 h group were obviously increased (P<0.05). As the most significant changes of three proteins were seen in the cells of LPS stimulation 24 h group, 24 h was used as the duration of subsequent LPS stimulation. After 24 hours of culture, the expression of FAM134B and its co-localization with LC3B and lysosomal probes in the cells of LPS group were all significantly enhanced, with a significant increase in the number of autolysosomes in comparison with those in PBS group. Both the empty vector group and the FAM134B-knockdown group showed high intensity fluorescence in the cells at post transfection hour 72, but the blank control group showed no fluorescence in the cells at the corresponding time point. After 24 hours of culture, the protein expression of FAM134B of cells in FAM134B-knockdown+PBS group was significantly lower than the expressions in PBS alone group and empty vector+PBS group (with P values all <0.05), the protein expression of FAM134B of cells in FAM134B-knockdown+LPS group was significantly lower than the expressions in LPS alone group and empty vector+LPS group (with P values all <0.05), the protein expression of FAM134B of cells in LPS alone group was significantly higher than that in PBS alone group (P<0.05), while the protein expression of FAM134B of cells in empty vector+LPS group was significantly higher than that in empty vector+PBS group (P<0.05). After 24 hours of culture, flow cytometry assay revealed that the apoptotic rate of cells in PBS alone group, LPS alone group, empty vector+PBS group, empty vector+LPS group, FAM134B-knockdown+PBS group, and FAM134B-knockdown+LPS group were (13.3±0.8)%, (32.6±4.3)%, (17.0±1.5)%, (51.7±3.3)%, (52.4±3.1)%, and (62.3±2.6)%, respectively. After 24 hours of culture, compared with those in LPS alone group and empty vector+LPS group, the protein expression of cleaved caspase-3, the ratio of Bax/Bcl-2, and the apoptotic rates of cells detected by flow cytometry and Hoechst staining were significantly increased in FAM134B-knockdown+LPS group (P<0.05); compared with those in the corresponding PBS treatment group, namely, PBS alone group, empty vector+PBS group, and FAM134B-knockdown+PBS group, the protein expression of cleaved caspase-3, the ratio of Bax/Bcl-2, and the apoptotic rates of cells detected by flow cytometry and Hoechst staining were significantly increased in LPS alone group, empty vector+LPS group, and FAM134B-knockdown+LPS group (P<0.05). Conclusions: The activation of reticulophagy mediated by FAM134B in mouse DCs is enhanced and peaked in 24 hours under LPS stimulation, and the activated reticulophagy has a significant inhibitory effect on cell apoptosis.