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08 July 2023, Volume 50 Issue 7 Previous Issue    Next Issue

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Guidelines · Consensus

Shandong Provincial Medical Association multidisciplinary standardized diagnosis and treatment guidelines for esophageal carcinoma （2023 Edition） Shandong Medical Association Multidisciplinary Joint Committee on Lung Cancer 2023, 50 (7):  385-397.  doi: 10.3760/cma.j.cn371439-20230612-00077 Abstract ( 159 )   HTML ( 146 )   PDF (891KB) ( 126 )   Save Esophageal carcinoma is at a high incidence in Shandong province，and there are many non-standard aspects in the current diagnosis and treatment. The standardized diagnosis and treatment of multiple disciplines，including medical oncology，surgical oncology，radiotherapy，imaging，endoscopy，pathology，etc.，is crucial to improve local control，overall survival rate，and quality of life. This guideline is hereby formulated based on clinical needs and combined with the progress of multidisciplinary diagnosis and treatment in recent years. Figures and Tables | References | Related Articles | Metrics

Original Articles

Research on BHLHE40 targets HMGA2 to reduce the sensitivity of thyroid cancer cells to cisplatin through activating the oxidative phosphorylation pathway Zhang Jinnan, Liu Bangqing, Li Jun, Liu Xiaohui 2023, 50 (7):  398-406.  doi: 10.3760/cma.j.cn371439-20230217-00078 Abstract ( 150 )   HTML ( 133 )   PDF (2870KB) ( 64 )   Save Objective To explore whether BHLHE40 can affect the sensitivity of thyroid cancer （TC） cells to cisplatin by activating oxidative phosphorylation （OXPHOS） pathway by targeting high mobility group A2 （HMGA2）. Methods The mRNA expression of HMGA2 and its upstream transcription factor BHLHE40 in TC tissues was analyzed by TCGA-THCA and hTFtarget online databases. The si-HMGA2，oe-HMGA2，oe-BHLHE40，negative control si-NC and oe-NC were transfected into TC cells （K1 and SW579） by liposome transfection method. The mRNA expression levels of BHLHE40 and HMGA2 in TC cells （SW579，FTC-133，and K1） and normal thyroid cells （Nthy ori3-1） were detected by real-time quantitative PCR （qRT-PCR）. The cell viability was detected by MTT assay，the half inhibitory concentration （IC50） value of cisplatin was calculated by CCK-8 assay，the apoptosis level was detected by flow cytometry，and the expression of OXPHOS complex was detected by Western blotting. Seahorse XFe 96 was used to analyze the oxygen consumption rate of the TC cells. Dual-luciferase assay and chromatin immunoprecipitation （ChIP） assay were used to analyze the binding relationship between BHLHE40 and HMGA2. Results TCGA database results showed that the mRNA expression levels of HMGA2 and BHLHE40 in TC tissues （10.57±2.58，13.89±1.13） were higher than those in normal thyroid tissues （4.82±1.69，12.28±1.01），with statistically significant differences （t=16.69，P<0.001； t=10.43，P<0.001）. The results of qRT-PCR showed that the relative mRNA expression levels of HMGA2 in normal thyroid cells （Nthy ori3-1） and TC cells （SW579，FTC-133，and K1） were 1.00±0.13，2.94±0.23，4.71±0.41 and 6.29±0.49，while those of BHLHE40 were 1.00±0.12，2.60±0.23，3.39±0.35 and 6.18±0.51 respectively，both with statistically significant differences （F=130.50，P<0.001； F=125.20，P<0.001）. Further pairwise comparison showed that mRNA expression levels of HMGA2 and BHLHE40 in TC cells were significantly higher than those in normal thyroid cells （all P<0.001）. According to MTT experimental results，si-HMGA2 treatment significantly reduced the cell viability of K1 cells compared to the si-NC group （all P<0.05）. In addition，compared to the oe-NC group，oe-HMGA2 treatment significantly increased the cell viability of SW579 cells （all P<0.05）. Compared to the oe-NC+DMSO group，the oe-HMGA2+DMSO group showed enhanced cell viability of SW579 cells，while the OXPHOS pathway inhibitor Gboxin was able to reverse the effect of overexpressing HMGA2 on cell viability （all P<0.05）. The results of flow cytometry and CCK-8 experiments showed that compared to the si-NC group （apoptosis level： 6.19%±0.28%； cisplatin IC50 value： 17.47 μmol/L），knocking down HMGA2 could increase the apoptosis level （11.96%±0.32%； t=19.17，P<0.001） and cisplatin sensitivity （IC50 value： 1.49 μmol/L） of K1 cells. In addition，compared to the oe-NC group （apoptosis level： 9.98%±0.32%； cisplatin IC50 value： 8.17 μmol/L），overexpressing HMGA2 significantly decreased the apoptosis level （4.32%±0.25%； t=19.65，P<0.001） and cisplatin sensitivity （IC50 value： 34.95 μmol/L） of SW579 cells. The results of dual-luciferase assay showed that compared with the si-NC group，knocking down the expression of BHLHE40 in human kidney epithelial 293T cells significantly reduced the luciferase activity of wild-type HMGA2 （0.31±0.02 vs. 1.00±0.11； t=10.69，P=0.004）. However，there was no significant effect on the luciferase activity of mutant-type HMGA2 （1.06±0.11 vs. 1.00±0.07； t=0.80，P=0.470）. ChIP results showed that the mRNA expression level of HMGA2 in K1 cells was significantly increased in the anti-BHLHE40 group （6.57±0.62） compared with the IgG group （1.00±0.10； t=15.36，P<0.001）. Compared to the oe-NC+DMSO group，the oe-HMGA2+DMSO group showed decreased apoptosis level （P<0.05） and cisplatin sensitivity of SW579 cells，with a significant increase in the expression of OXPHOS complexes Ⅰ-Ⅴ and cellular oxygen consumption rates （all P<0.05）. The effect of overexpressing HMGA2 was reversed by treatment with oe-HMGA2+Gboxin （all P<0.05）. The recovery experiment showed that compared to the oe-NC+si-NC group，overexpression of BHLHE40 in SW579 cells increased cell viability and the expression of OXPHOS complexes Ⅰ-Ⅴ，while decreasing apoptosis levels and increasing cellular oxygen consumption rates and cisplatin IC50 values （all P<0.05）. However，simultaneous knockdown of HMGA2 reversed the effect of overexpressing BHLHE40 （all P<0.05）. Conclusion BHLHE40 can activate the OXPHOS pathway by targeting and regulating the expression of HMGA2，thereby affecting the sensitivity of TC cells to cisplatin. Figures and Tables | References | Related Articles | Metrics

RNF43 inhibits PD-L1 expression via β-catenin in melanoma cells and promotes CD8+ T cell-mediated anti-tumor immune reaction Wu Minhang, Sun Wenzheng, Yu Qingzhuo, Guo Rong, Ye Hui, Du Ying, Qiu Jin, An Huazhang, Cao Lili 2023, 50 (7):  407-412.  doi: 10.3760/cma.j.cn371439-20230404-00079 Abstract ( 147 )   HTML ( 129 )   PDF (1251KB) ( 72 )   Save Objective To investigate the regulatory effects of ring finger protein 43 （RNF43） on CD8+ T cell-mediated anti-tumor immune reaction in melanoma. Methods RNF43 gene was over-expressed and knockdown in mouse melanoma cells line B16-OVA by lentivirus infection； In vivo proliferation of mouse melanoma cells line B16-OVA in the Lv-Ctrl-OE，Lv-RNF43-OE，Lv-Ctrl-KD and Lv-RNF43-KD groups was detected by subcutaneous tumorigenesis assay in mice，and the expression levels of CD8+ T cells perforin and interferon γ （IFN-γ） in tumor immune microenvironment of melanoma were detected by flow cytometry； The expression levels of β-catenin and programmed death-ligand 1 （PD-L1） mRNA in cells were detected by quantitative real-time PCR assay； The effect of RNF43 on the transcriptional regulation of PD-L1 was detected by dual-luciferase reporter gene assay. Results Stable RNF43 over-expressing and RNF43 knockdown mouse melanoma cells lines Lv-RNF43-OE and Lv-RNF43-KD were successfully constructed. The results of subcutaneous tumorigenesis experiment in mice showed that the tumor mass of the Lv-RNF43-OE group was （0.08±0.06） g，which was significantly smaller than that of the Lv-Ctrl-OE group [（1.04±0.52） g]，with a statistically significant difference （t=3.71，P=0.032）； The tumor mass of Lv-RNF43-KD group was （1.94±0.29） g，with no statistically significant difference （t=-1.70，P=0.164） compared with that of the Lv-Ctrl-KD group （1.15±0.74） g. The flow cytometry results showed that the fluorescence intensity of CD8+ T cell perforin in the Lv-RNF43-OE group was 9 034 ± 2 628，which was significantly higher than that in the Lv-Ctrl-OE group （3 847 ±1 637），with a statistically significant difference （t=-3.35，P=0.015）； The fluorescence intensity of CD8+ T cell perforin in the Lv-RNF43-KD group was 966±247，which was significantly lower than that in the Lv-Ctrl-KD group （2 226±646），with a statistically significant difference （t=3.16，P=0.034）； The fluorescence intensity of IFN-γ of CD8+ T cell in the Lv-RNF43-OE group was 2 422±429，which was significantly higher than that of 1 688±324 in the Lv-Ctrl-OE group，with a statistically significant difference （t=-2.73，P=0.034）； The fluorescence intensity of IFN-γ of CD8+ T cell in the Lv-RNF43-KD group was 614 （454，863），with a statistically significant difference （Z=-1.96，P=0.050） compared with 1 159 （1 152，2 068） in the Lv-Ctrl-KD group. The results of quantitative real-time PCR showed that the relative expression level of β-catenin mRNA in the Lv-RNF43-OE group was 0.67±0.16，which was significantly lower than that of 1.00±0.11 in the Lv-Ctrl-OE group，with a statistically significant difference （t=2.98，P=0.041）； The relative expression level of PD-L1 mRNA in the Lv-RNF43-OE group was 0.32±0.09，which was significantly lower than that of 1.00±0.09 in the Lv-Ctrl-OE group，with a statistically significant difference （t=9.13，P=0.001）. The results of the dual-luciferase reporter gene assay showed that the PD-L1 promoter luciferase activity in the pCMV6-NC，RNF43，RNF43+β-catenin and β-catenin groups were 1.00±0.00，0.84±0.00，1.49±0.00 and 1.57±0.03 （F=2 218.33，P<0.001）. Further pairwise comparison showed that compared with the pCMV6-NC group，PD-L1 promoter luciferase activity was significantly lower in the RNF43 group （P<0.001） and significantly higher in the RNF43+β-catenin and β-catenin groups （P<0.001； P=0.003）； compared with the RNF43 group，PD-L1 promoter luciferase activity was significantly higher in the RNF43+β-catenin group （P<0.001）. Conclusion RNF43 may reduce the expression of PD-L1 mRNA in melanoma by inhibiting the expression of β-catenin and promote CD8+ T cell-mediated anti-tumor immune reaction. Figures and Tables | References | Related Articles | Metrics

Mechanism of Bcl-2 BH4 selective inhibitor BDA-366 on NK/T cell lymphoma cells Wu Jiali, Zhang Jiahui, Zhang Ping, Xiao Xinyue, Li Rui, Zhang Hongyu 2023, 50 (7):  413-418.  doi: 10.3760/cma.j.cn371439-20230227-00080 Abstract ( 119 )   HTML ( 23 )   PDF (1602KB) ( 46 )   Save Objective To investigate the inhibitory effect and killing mechanism of Bcl-2 BH4 selective inhibitor BDA-366 on NK/T cell lymphoma （NK/TCL）. Methods Human NK cell leukemia cell line YT and human NK/TCL cell line NK92 cells were treated with 0，0.05，0.10，0.20，0.30，0.40，0.50 μmol/L BDA-366. CCK-8 assay was used to calculate the half inhibitory concentration （IC50） value of BDA-366 on these cells. The apoptosis levels of cells in control group and IC50 BDA-366 treated group were detected by flow cytometry. Western blotting was used to detect the expression levels of apoptosis-related proteins in cells of control group and 1/2 IC50，IC50，2× IC50 BDA-366 treated groups. TMRE and Fluo-3 fluorescent probe were used to detect mitochondrial membrane potential of control group and IC50 BDA-366 treated group，and the intracellular Ca2+ concentration of control group，IC50，2× IC50 BDA-366 treated groups. NOD-SCID mice in control group and 10 mg/kg BDA-366 intraperitoneal injection group were weighed and HE staining was performed to evaluate the toxicity of BDA-366 in vivo. Results The IC50 of BDA-366 for YT and NK92 cells were 0.065 and 0.086 μmol/L respectively. The apoptosis rates of YT cells in the control group and 0.065 μmol/L BDA-366 group were （6.62±1.59） % and （34.60±3.06） % respectively. The apoptosis rates of NK92 cells in the control group and 0.086 μmol/L BDA-366 group were （5.57±0.88） % and （29.18±0.90） % respectively，both with statistically significant differences （t=14.05，P<0.001； t=32.58，P<0.001）. The relative expression of Bax in NK92 cells of the control group，0.043，0.086 and 0.172 μmol/L BDA-366 groups were 0.85±0.00，1.26±0.04，1.51±0.18，1.15±0.10 （F=20.70，P<0.001），the relative expression of Bax in BDA-366 groups were higher than that in the control group （all P<0.05）. The fluorescence intensity of TMRE of YT cells in the control group and 0.065 μmol/L BDA-366 group were 8 372.00±330.47 and 6 419.67±311.34，and that of NK92 cells in the control group and 0.086 μmol/L BDA-366 group were 9 169.00±535.72 and 7 311.67±295.52 respectively，and there were statistically significant differences （t=7.45，P=0.002； t=5.26，P=0.006）. In YT cells，the intracellular Ca2+ concentrations of 0.065 and 0.130 μmol/L BDA-366 groups were significantly higher than that of the control group （5 791.67±220.45，6 729.33±585.39，4 874.67±112.61，F=19.16，P=0.003） （P=0.039； P=0.002）. In NK92 cells，the intracellular Ca2+ concentrations of 0.086 and 0.172 μmol/L BDA-366 groups were significantly higher than that of the control group （4 553.67±17.62，4 740.33±254.50，4 185.67±17.67，F=10.96，P=0.010） （P=0.039； P=0.007）. There was no statistically significant difference in body weight change on day 12 compared with day 0 of NOD-SCID mice between BDA-366 group and control group [（3.18±0.01） g vs. （2.73±0.58） g，t=0.60，P=0.570]，and HE staining showed no abnormal morphology of heart，liver，spleen，lung and kidney in BDA-366 group. Conclusion BDA-366 promotes NK/TCL cells apoptosis in vitro，but does not cause weight loss and morphological changes of organs by HE staining in vivo. The inhibitory effect of BDA-366 on NK/TCL cells may be achieved by increasing Bax expression，inducing Ca2+ release and reducing mitochondrial membrane potential. Figures and Tables | References | Related Articles | Metrics

Diagnostic value of transbronchial lung biopsy and bronchoalveolar lavage in pulmonary complications in patients with hematological tumors Wang Jun, Rong Lei, Huang Jing, Meng Jingye, Guo Zhi 2023, 50 (7):  419-424.  doi: 10.3760/cma.j.cn371439-20220912-00081 Abstract ( 109 )   HTML ( 15 )   PDF (1484KB) ( 48 )   Save Objective To evaluate the diagnostic value and safety of transbronchial lung biopsy and bronchoalveolar lavage （BAL） in pulmonary complications in patients with hematological tumors. Methods A retrospective analysis was performed on 68 patients with hematological tumors combined with lung lesions from The University of Hong Kong-Shenzhen Hospital and The Third People's Hospital of Shenzhen from May 2016 to May 2022，including 37 males，31 females，with a median age of 56 years （age range 21-90 years），among which 20 patients were >65 years old. Diagnostic fiberoptic bronchoscopy was performed with signs including fever，cough，hypoxemia，hemoptysis，unexplained dyspnea，and imaging changes. Patients with pulmonary masses were evaluated for transbronchial lung biopsy，including inner and outer leaf mass and high-density shadow of lung leaves，pathological and special staining of biopsy tissue （Grocott staining），BAL acquisition of bronchoalveolar lavage fluid （BALF） for microbiological smear/culture，cytomegalovirus， Pneumocystis jirovecii and Mycobacterium tuberculosis （TB） smear，TB DNA，TB and fungal culture. Etiological analysis of pulmonary complications and observation of the complications associated with fiberoptic bronchoscopy in patients with hematological tumors were conducted. Results BALF test was performed in all patients after bronchoscopy，bronchoscopic lung tissue biopsy was performed in 46 cases. The total number of confirmed pathogenic infections was 40，including 12 cases of fungal infections，9 cases of bacterial infections （2 cases each of E. faecalis and Pseudomonas aeruginosa，1 case of Staphylococcus aureus，1 case of Klebsiella pneumoniae，1 case of E. coli，1 case of coagulase-negative Staphylococcus，and 1 case of Streptococcus mitis），9 cases of viral infection （5 cases of cytomegalovirus，3 cases of parainfluenza virus type Ⅲ，and 1 case of respiratory syncytial virus），4 confirmed cases of Pneumocystis jirovecii pneumonia，3 cases of suspected mixed infection of Pneumocystis jirovecii and fungi，1 case of Cryptococcus，2 cases of suspected TB infection. No pathogenic organisms were found in 28 cases，including 6 cases of mechanized pneumonia，6 cases associated with a history of hematological tumors，and 16 cases of other unidentified pathogens. All patients did not experience death or other serious complications caused by bronchoscopy complications. Conclusion Pulmonary complications are common in patients with hematological tumors，and the application of transbronchial lung biopsy has good safety. Early examination of fiberoptic bronchoscopy can provide pathogenic diagnostic evidence of bacterial，fungal，Pneumocystis jirovecii and viral infections，thus improving the diagnostic rate. Figures and Tables | References | Related Articles | Metrics

Analysis of global cancer incidence and mortality from 1990 to 2019 Zhao Qianwen, Peng Danli, Qin Tao, Mao Guofei, Tang Hui, Yang Jun 2023, 50 (7):  425-431.  doi: 10.3760/cma.j.cn371439-20230315-00082 Abstract ( 134 )   HTML ( 23 )   PDF (1379KB) ( 55 )   Save Objective To analyze the global incidence and mortality of cancer from 1990 to 2019. Methods The Global Burden of Disease Study 2019 （GBD2019） database was utilized to analyze the global incidence and mortality of cancer，the order of incidence and mortality of cancer，the incidence and mortality of different age groups，and the trend of incidence and mortality from 1990 to 2019. Standardized incidence and mortality rates were derived by utilizing the world standard population age structure. Results In 1990，global cancer cases numbered 10.295 9 million with an incidence rate of 192.45/100 000，leading to 5.732 6 million deaths and a mortality rate of 107.16/100 000. While in 2019，global cancer cases escalated to 23.568 5 million with an incidence rate of 304.60/100 000，resulting in 10.022 8 million deaths and a mortality rate of 129.54/100 000，all higher than those in 1990. In 2019，lung cancer showed the highest incidence rate of both sexes combined in the world （29.21/100 000），followed by colorectal cancer，breast cancer，prostate cancer and gastric cancer. The incidence of lung cancer was highest among males （39.24/100 000），while the incidence of breast cancer was highest among females （51.27/100 000）. Lung cancer also had the highest mortality rate worldwide in both sexes combined （26.40/100 000），followed by colorectal cancer，gastric cancer，breast cancer and pancreatic cancer. Lung cancer had the highest mortality among males （35.72/100 000），while breast cancer had the highest mortality among females （17.85/100 000）. In 2019，the global cancer incidence rate showed an upward trend with age. The incidence rate was low before the age of 25，and increased rapidly after the age of 25. The incidence rates of both sexes combined，males and females all reached the peak in the age group of over 85 years old，which were 3 084.18/100 000，4 434.81/100 000 and 2 353.07/100 000 respectively； The incidence rate of females in the age group of 20-50 years old was higher than that of males，but the incidence rate of males in the age group of over 55 years old was higher than that of females. Compared with 1990，the incidence rates of both sexes combined in the age group of over 20，of males over 55 years old，as well as of females over 15 years old，were all higher than those in 2019. In 2019，the global tumor mortality rate showed an upward trend with age. The mortality rate was relatively low before the age of 35，and increased rapidly after the age of 35. The mortality rates for both sexes combined，as well as for males and females，reached the peak in the age group of over 85 years old，which were 1 787.84/100 000，2 509.87/100 000，and 1 369.99/100 000 respectively； The mortality rate of females in the age group of 20-40 years old was higher than that of males，and the mortality rate of males in the age group of over 45 years old was higher than that of females； For the age of 0-80 years old，the mortality rates for both sexes combined，males，and females were lower in 2019 than 1990，but higher in the age of 85 years old and above. The global standardized incidence rate of cancer showed an overall upward trend，with an average annual increase of 0.30% from 1990 to 2019. The global standardized mortality rate of cancer showed an overall downward trend，with an average annual decrease of 0.60% from 1990 to 2019. Conclusion From 1990 to 2019，the global standardized incidence rate of cancers shows an overall upward trend，while the global standardized mortality rate of cancers has an overall downward trend，and the global incidence and mortality rate of cancers increases with age. The global burden of cancer disease is still heavy. Lung cancer is the cancer with the highest incidence and mortality rate in the world. The highest incidence rate is lung cancer among males，and breast cancer among females. Different countries or regions need to take corresponding cancer prevention and treatment strategies according to their actual conditions. Figures and Tables | References | Related Articles | Metrics

Reviews

Gut microbiome and tumor immunotherapy Guo Ciliang, Jiang Chunping, Wu Junhua 2023, 50 (7):  432-436.  doi: 10.3760/cma.j.cn371439-20230413-00083 Abstract ( 157 )   HTML ( 30 )   PDF (759KB) ( 86 )   Save Neoplasms immunotherapy has made a major breakthrough in the clinical practice of refractory tumor. However，there are still individual differences in treatment results and drug resistance in clinical application. Gastrointestinal microbiome is gradually recognized as an immunoregulatory factor in recent years，and more and more studies have focused on its influences on the efficacy of tumor immunotherapy. Targeting gastrointestinal microbiota to improve the response of tumor patients to immunotherapy has potential clinical application value. References | Related Articles | Metrics

Research progress of 4-nitroquinoline-1-oxide-induced esophageal squamous cell carcinoma model in mice Li Jinge, Li Jing, Zhang Zhenhan, Guo Jianxin, Wang Jingpu, Wu Zhongbing 2023, 50 (7):  437-441.  doi: 10.3760/cma.j.cn371439-20230227-00084 Abstract ( 180 )   HTML ( 25 )   PDF (788KB) ( 91 )   Save Esophageal squamous cell carcinoma （ESCC） is one of the most common malignant tumors with a poor prognosis. 4-nitroquinoline-1-oxide （4NQO） is a water-soluble quinoline derivative that can successfully induce the production of squamous cell carcinoma in vivo. Establishing and optimizing experimental methods for 4NQO induced ESCC formation in mice can provide a more suitable in situ model for the study of ESCC. References | Related Articles | Metrics