Objective:To investigate the effects of pulsed electric field (PEF) combined with gemcitabine (GEM) on the viability and stemness of HCCC-9810 cholangiocarcinoma stem cells.Methods:HCCC-9810 cholangiocarcinoma stem cells were established in serum-free, cytokine-rich medium and divided into four groups: the control group, GEM group, PEF group, and the pulsed electric field combined with gemcitabine (PEF+ GEM) group. Cell proliferation was detected using the Cell Counting Kit-8 (CCK8) assay. Cell viability and apoptosis rate were measured by flow cytometry. Cell invasion ability was assessed using the Transwell assay. The expression of stemness marker proteins CD133 and Octamer-binding transcription factor 4 (OCT4), as well as the expression of β-catenin, was detected by Western blotting.Results:Regarding cell viability, the GEM, PEF, and PEF+ GEM groups showed significantly lower cell viability and higher apoptosis rate than the control group at 24 h, 48 h, and 72 h (all P<0.05). At 48 h and 72 h, the PEF+ GEM group showed significantly lower cell viability (7.2%±0.3% and 5.9%±0.8%, respectively) than the GEM group (50.7%±0.6% and 31.0%±1.2%, respectively) and the PEF group (12.2%±0.2% and 12.8%±0.2%, respectively) (all P<0.05). Regarding stemness inhibition, the PEF+ GEM groups showed significantly lower expression levels of CD133 and OCT4 at 24 h, 48 h, and 72 h compared with the control group (all P<0.05). Notably, at 48 h, the PEF+ GEM group showed a significantly lower expression level of the OCT4 (0.61±0.02) than the GEM group (0.87±0.08) and the PEF group (1.00±0.10) ( P<0.01). Furthermore, at 24 h and 48 h, the GEM, PEF, and PEF+ GEM groups showed significantly lower expression levels of β-catenin compared with the control group (all P<0.05). Conclusion:Pulsed electric field combined with gemcitabine therapy demonstrated more effective anti-proliferation and cancer stemness inhibition effects on HCCC-9810 cholangiocarcinoma stem cells compared with either monotherapy.

Objective:To investigate the effects of pulsed electric field (PEF) combined with gemcitabine (GEM) on the viability and stemness of HCCC-9810 cholangiocarcinoma stem cells.Methods:HCCC-9810 cholangiocarcinoma stem cells were established in serum-free, cytokine-rich medium and divided into four groups: the control group, GEM group, PEF group, and the pulsed electric field combined with gemcitabine (PEF+ GEM) group. Cell proliferation was detected using the Cell Counting Kit-8 (CCK8) assay. Cell viability and apoptosis rate were measured by flow cytometry. Cell invasion ability was assessed using the Transwell assay. The expression of stemness marker proteins CD133 and Octamer-binding transcription factor 4 (OCT4), as well as the expression of β-catenin, was detected by Western blotting.Results:Regarding cell viability, the GEM, PEF, and PEF+ GEM groups showed significantly lower cell viability and higher apoptosis rate than the control group at 24 h, 48 h, and 72 h (all P<0.05). At 48 h and 72 h, the PEF+ GEM group showed significantly lower cell viability (7.2%±0.3% and 5.9%±0.8%, respectively) than the GEM group (50.7%±0.6% and 31.0%±1.2%, respectively) and the PEF group (12.2%±0.2% and 12.8%±0.2%, respectively) (all P<0.05). Regarding stemness inhibition, the PEF+ GEM groups showed significantly lower expression levels of CD133 and OCT4 at 24 h, 48 h, and 72 h compared with the control group (all P<0.05). Notably, at 48 h, the PEF+ GEM group showed a significantly lower expression level of the OCT4 (0.61±0.02) than the GEM group (0.87±0.08) and the PEF group (1.00±0.10) ( P<0.01). Furthermore, at 24 h and 48 h, the GEM, PEF, and PEF+ GEM groups showed significantly lower expression levels of β-catenin compared with the control group (all P<0.05). Conclusion:Pulsed electric field combined with gemcitabine therapy demonstrated more effective anti-proliferation and cancer stemness inhibition effects on HCCC-9810 cholangiocarcinoma stem cells compared with either monotherapy.

Objective:To analyze the effects of irinotecan combined with XELOX regimen on immune status, intestinal microecology and prognostic risk in elderly patients with colorectal cancer.Methods:A total of 105 elderly patients with advanced colorectal cancer admitted to Qidong People's Hospital of Jiangsu Province from October 2018 to April 2023 were included as the study objects. They were divided into control group ( n=45) and observation group ( n=60) according to different chemotherapy regimen. The control group was treated with XELOX regimen alone, and the observation group was treated with irinotecan combined with XELOX regimen. The short-term efficacy, changes of indexes related to immune status and intestinal microecology before and after treatment were compared between the two groups. The patients were followed up from the end of treatment. With death or recurrence and metastasis as the end event during the follow-up, 105 patients were divided into poor prognosis group ( n=32) and good prognosis group ( n=73). The clinical data of the two groups were compared, and multivariate logistic regression was used to analyze the prognostic factors in elderly patients with advanced colorectal cancer. Results:The total effective rate of the observation group was 53.33% (32/60), which was higher than that of the control group (20.00%, 9/45) ( χ2=12.01, P=0.001). One week after treatment, the ratios of CD4 + T cells and CD4 +/CD8 + in the observation group were (38.59±1.50) % and 1.81±0.20, respectively, higher than those in the control group (36.25±1.82) % and 1.59±0.15 ( t=7.22, P<0.001; t=6.19, P<0.001). The ratio of CD8 + T cells was (21.27±2.70) %, lower than that of the control group (22.80±2.92) % ( t=2.78, P=0.007). The numbers of Bifidobacterium, Lactobacillus and Enterococcus were (9.44±0.82), (9.89±0.79), (9.14±0.66) lg CFU/g, respectively, which were higher than those in the control group (8.20±0.70), (9.05±0.72), (8.25±0.62) lg CFU/g ( t=8.16, P<0.001; t=5.60, P<0.001; t=7.02, P<0.001). There were statistically significant differences in the proportion of smoking and drinking history ( χ2=7.61, P=0.006), the proportion of low differentiation ( χ2=6.54, P=0.011), the proportion of lymph node metastasis ( χ2=5.86, P=0.016) and the level of carcinoembryonic antigen (CEA) before chemotherapy [ (5.80±0.89) μg/L vs. (7.48±1.02) μg/L, t=8.51, P<0.001], the level of carbohydrate antigen 199 (CA199) [ (29.54±1.85) U/ml vs. (34.52±2.50) U/ml, t=11.36, P<0.001] in good prognosis group and poor prognosis group. Multivariate analysis showed that smoking and drinking history ( OR=1.74, 95% CI: 1.53-2.15, P<0.001), low differentiation ( OR=1.80, 95% CI: 1.60-2.15, P<0.001), lymph node metastasis ( OR=1.82, 95% CI: 1.68-2.33, P<0.001), high CEA level before chemotherapy ( OR=1.81, 95% CI: 1.62-2.38, P<0.001), high CA199 level before chemotherapy ( OR=1.80, 95% CI: 1.66-2.37, P<0.001) were risk factors for the prognosis of advanced colorectal cancer in the elderly. Conclusion:Irinotecan combined with XELOX regimen can effectively improve immune function and intestinal microecology in elderly patients with advanced colorectal cancer, but the risk of poor prognosis after chemotherapy is higher. Smoking and drinking history, low differentiation, lymph node metastasis, high CEA level before chemotherapy, and high CA199 level before chemotherapy are risk factors affecting the prognosis of elderly patients with advanced colorectal cancer.

Objective:To provide scientific basis for prevention and control measures of island regions through analyzing the characteristics of malignant tumor incidence and mortality in eastern island areas of China and comparing the data with the national cancer mortality data in the same period. Methods:Using the incidence and mortality data of malignant tumor in Daishan county,Zhoushan collected by the Daishan Center for Disease Control and Prevention(CDC)from 2014 to 2019,which was stratified by gender,the crude incidence rates(CIR)and crude mortality rates(CMR)were calculated,and the top 10 malignant tumors with the highest incidence or mortality rates were then ranked.The Segi's world standard population was used to calculate the age-standardized incidence(ASIR)and age-standardized mortality rate(ASMR).The local data were compared with the national cancer mortality data from 2014 to 2019,and Chi-square test was used to analyze the differences between these two sets of data using the SPSS software.The difference was considered statistically significant when P<0.05. Results:There was a total of 7 305 incidence cases of malignant tumors in Daishan county,Zhoushan from 2014 to 2019.The CIR was 662.39/105 and the ASIR was 306.81/105.Notably,the CIR was the highest in the age group of 75-79 years old.The top 5 malignant tumors with the highest incidence rates were lung cancer(27.15％),gastric cancer(12.76％),liver cancer(10.95％),colorectal cancer(6.92％)and breast cancer(5.42％),whose ASIR were 75.09/105,32.06/105,31.01/105,17.81/105 and 18.36/105,respectively.There was a total of 3 412 mortality cases of malignant tumors in Daishan county,Zhoushan from 2014 to 2019.The CMR was 309.39/105 and the ASMR was 122.73/105.Notably,the CMR was the highest in the age group of 80-85 years old.The top 5 malignant tumors with the highest mortality rates were lung cancer(24.94％),liver cancer(18.64％),gastric cancer(17.00％),colorectal cancer(7.56％)and esophageal cancer(5.72％),whose ASMR were 29.65/105,24.97/105,19.01/105,8.75/105 and 6.60/105,respectively.The total ASMR of malignant tumors in Daishan county,Zhoushan was higher than national total ASMR from 2014 to 2019(100.34/105)(P<0.001).Specifically,the ASMR of gastric cancer,lung cancer and colorectal cancer in Daishan county were significantly higher than national levels(gastric cancer:12.46/105;liver cancer:16.45/105;colorectal cancer:6.63/105)(P<0.01),whereas no significant difference in the ASMR of lung cancer and esophageal cancer between Daishan county and the whole nation(lung cancer:28.06/105;esophageal cancer:7.61/105)was observed. Conclusion:Lung cancer,gastric cancer,liver cancer and colorectal cancer were malignant tumors with higher incidence and mortality rates in Daishan county,Zhoushan from 2014 to 2019.Particularly,the ASMR of gastric cancer,liver cancer and colorectal cancer were significantly higher than the national levels,and these malignant tumors should be considered as the major focus of cancer prevention and control.

Objective:Establishment of a new model of human primary colon cancer transplantation tumor in normal immune mice and to provide a reliable experimental animal model for studying the pathogenesis of colon cancer under normal immunity.Methods:Human colon cancer cells come from colon cancer patients who underwent surgery in the Affiliated Hospital of Jining Medical College in 2017. The mice in the cell control group were inoculated with phosphate buffered solution (PBS) containing colon cancer cells, the microcarrier control group was inoculated with PBS containing microcarrier 6, and the cell-microcarrier complex group was inoculated with the PBS containing colon cancer cell-microcarrier complex. The cells of each group were inoculated under the skin of the right axilla of mice by subcutaneous injection, and the time, size, tumor formation rate and pathological changes under microscope were recorded. The transplanted tumor tissue was immunohistochemically stained with the EnVisiion two-step method, and the tumor formation rate of the transplanted tumor was judged according to the proportion of positive cells in the visual field. The polymerase chain reaction (PCR) method was used to detect the expression of human-specific Alu sequence in mice tumor tissue.Results:After inoculation with tumor cells, the mice in the cell control group and the microcarrier control group did not die and did not form tumors; the mice in the cell-microcarrier complex group had palpable subcutaneous tumors in the right axillary subcutaneously on the 5th to 7th days after inoculation, and tumor formation rate is 67% (10/15), and the tumor volume can reach about 500 mm 3 2 to 3 weeks after vaccination. The immunohistochemistry results showed that CK20, CDX-2 and carcinoembryonic antigen were all positively expressed. The PCR results showed that the expression of human-specific Alu sequence can be detected in the transplanted tumor tissue of tumor-bearing mice. Conclusion:Human primary colon cancer cells used microcarrier 6 as a carrier to form tumors in normal immunized mice, and successfully established a new model of human colon cancer transplantation tumor in normal immune mice.

Objective:Establishment of a new model of human primary colon cancer transplantation tumor in normal immune mice and to provide a reliable experimental animal model for studying the pathogenesis of colon cancer under normal immunity.Methods:Human colon cancer cells come from colon cancer patients who underwent surgery in the Affiliated Hospital of Jining Medical College in 2017. The mice in the cell control group were inoculated with phosphate buffered solution (PBS) containing colon cancer cells, the microcarrier control group was inoculated with PBS containing microcarrier 6, and the cell-microcarrier complex group was inoculated with the PBS containing colon cancer cell-microcarrier complex. The cells of each group were inoculated under the skin of the right axilla of mice by subcutaneous injection, and the time, size, tumor formation rate and pathological changes under microscope were recorded. The transplanted tumor tissue was immunohistochemically stained with the EnVisiion two-step method, and the tumor formation rate of the transplanted tumor was judged according to the proportion of positive cells in the visual field. The polymerase chain reaction (PCR) method was used to detect the expression of human-specific Alu sequence in mice tumor tissue.Results:After inoculation with tumor cells, the mice in the cell control group and the microcarrier control group did not die and did not form tumors; the mice in the cell-microcarrier complex group had palpable subcutaneous tumors in the right axillary subcutaneously on the 5th to 7th days after inoculation, and tumor formation rate is 67% (10/15), and the tumor volume can reach about 500 mm 3 2 to 3 weeks after vaccination. The immunohistochemistry results showed that CK20, CDX-2 and carcinoembryonic antigen were all positively expressed. The PCR results showed that the expression of human-specific Alu sequence can be detected in the transplanted tumor tissue of tumor-bearing mice. Conclusion:Human primary colon cancer cells used microcarrier 6 as a carrier to form tumors in normal immunized mice, and successfully established a new model of human colon cancer transplantation tumor in normal immune mice.