1.Weichang'an Prescription-containing Serum Induces Ferroptosis of Gastric Cancer MKN-45 Cells
Xin LI ; Jinzu YANG ; Jianxin QIAN ; Li TAO ; Ling CHEN
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(4):132-139
ObjectiveTo observe the effect of Weichang'an prescription-containing serum on ferroptosis of human gastric cancer cells and explore the possible mechanism. MethodsSD rats were administrated with 18, 36, 72 g·kg-1·d-1 Weichang'an prescription by gavage for preparation of serum samples containing different doses of Weichang'an prescription, which were then used to treat MKN-45 cells. The cell proliferation was examined by the cell counting kit-8 (CCK-8). In addition, inhibitors of apoptosis, necroptosis, and ferroptosis were added, and the survival of the cells treated with the serum samples was observed. The fluorescent probe dichlorodihydrofluorescein diacetate (DCF-DA) and the lipid peroxidation sensor C11-BODIPY were employed to detect the intracellular levels of reactive oxygen species (ROS) and lipid peroxidation, respectively. The levels of ferrous ion (Fe2+), glutathione (GSH), and malondialdehyde (MDA) were detected by enzyme-linked immunosorbent assay (ELISA). Real-time PCR and Western blotting were employed to determine the expression of nuclear factor erythroid 2-related factor 2 (Nrf2), aldo-keto reductase family 1 member B1 (AKR1B1), glutathione peroxidase 4 (GPX4), acyl-CoA synthetase long-chain family member 4 (ACSL4), signal transducer and activator of transcription 3 (STAT3), and mitogen-activated protein kinase (MAPK). ResultsCompared with the blank group, Weichang'an prescription-containing serum decreased the viability of MKN-45 cells (P<0.05, P<0.01) in a time- and dose-dependent manner. Compared with the Weichang'an prescription group, the apoptosis inhibitor+Weichang'an prescription group and the ferroptosis inhibitor+Weichang'an prescription group showed increased cell viability (P<0.05, P<0.01). Compared with the blank group, Weichang'an prescription elevated the levels of ROS, lipid peroxidation, and intracellular Fe2+ and MDA (P<0.05, P<0.01) and lowered the level of GSH (P<0.05, P<0.01) in a dose-dependent manner. Compared with the blank group, Weichang'an prescription down-regulated the mRNA and protein levels of Nrf2, AKR1B1, and GPX4 (P<0.05, P<0.01) and up-regulated the mRNA and protein levels of ACSL4 (P<0.05, P<0.01) in a dose-dependent manner. Compared with the blank group, Weichang'an prescription down-regulated the protein levels of p-STAT3 and p-ERK (P<0.05, P<0.01) in a dose-dependent manner. ConclusionThe Weichang'an prescription-containing serum can promote the ferroptosis and inhibit the proliferation of MKN-45 cells by regulating the STAT3 and MAPK pathways.
2.Discussion on the Intestinal Immune Mechanism and Clinical Diagnosis and Treatment of Hashimoto Thyroiditis Based on"Wood Depression and Earth Stagnation"
Guijie WANG ; Yukun WU ; Zongliang YU ; Zhenzhen QIAN ; Jianxin WANG ; Rui GAO
Chinese Journal of Information on Traditional Chinese Medicine 2025;32(11):172-176
Hashimoto thyroiditis(HT)is one of the most common autoimmune diseases."Wood depression and earth stagnation"is its important pathogenesis.Liver depression and qi stagnation,wood depression multiplies the earth,spleen deficiency and earth stagnation,phlegm and blood stasis intermingle,resulting in wood depression and earth stagnation.Or middle-earth stagnation insults wood in turn,liver yang deficiency,wood depression,and failure to disperse and discharge,resulting in earth stagnation and wood depression.It may be related to the intestinal immune mechanism in modern medical study.Based on the theory of"wood depression and earth stagnation",this article discussed the pathogenesis of HT from the perspective of wood depression and imbalance of microbe-gut-brain axis,earth stagnation and dysbiosis of intestinal flora,damage to the intestinal barrier,and proposed the treatment principles,i.e.,"disperse stagnated liver qi for relieving qi stagnation,ascend yang and regulate qi""eliminate stagnation and remove turbidity,invigorate spleen and restore normal movement",which could provide the ideas for mechanism research and clinical treatment of HT.
3.Discussion on the Intestinal Immune Mechanism and Clinical Diagnosis and Treatment of Hashimoto Thyroiditis Based on"Wood Depression and Earth Stagnation"
Guijie WANG ; Yukun WU ; Zongliang YU ; Zhenzhen QIAN ; Jianxin WANG ; Rui GAO
Chinese Journal of Information on Traditional Chinese Medicine 2025;32(11):172-176
Hashimoto thyroiditis(HT)is one of the most common autoimmune diseases."Wood depression and earth stagnation"is its important pathogenesis.Liver depression and qi stagnation,wood depression multiplies the earth,spleen deficiency and earth stagnation,phlegm and blood stasis intermingle,resulting in wood depression and earth stagnation.Or middle-earth stagnation insults wood in turn,liver yang deficiency,wood depression,and failure to disperse and discharge,resulting in earth stagnation and wood depression.It may be related to the intestinal immune mechanism in modern medical study.Based on the theory of"wood depression and earth stagnation",this article discussed the pathogenesis of HT from the perspective of wood depression and imbalance of microbe-gut-brain axis,earth stagnation and dysbiosis of intestinal flora,damage to the intestinal barrier,and proposed the treatment principles,i.e.,"disperse stagnated liver qi for relieving qi stagnation,ascend yang and regulate qi""eliminate stagnation and remove turbidity,invigorate spleen and restore normal movement",which could provide the ideas for mechanism research and clinical treatment of HT.
4.Analysis of factors affecting the detection of urinary stone using virtual unenhanced images derived from dual-energy CTU
Yannan CHENG ; Yanan LI ; Jingtao SUN ; Qian TIAN ; Jian YANG ; Wei TONG ; Jian YANG ; Jianxin GUO
Journal of Xi'an Jiaotong University(Medical Sciences) 2024;45(4):535-541
Objective To evaluate the factors affecting urinary stone detection rate using virtual unenhanced(VUE)images obtained from triphasic dual-energy CT urography(DECTU)based on Logistic regression analysis.Methods For this study,150 patients who had suspected urinary stone and underwent triphasic DECTU were included.The true unenhanced(TUE)images were reconstructed as 120 kVp-like images,and VUE images at the portal venous phase[VUE(VP)]and excretory phase[VUE(EP)]were obtained using iodine removal technique from portal venous and excretory phase DECTU images,respectively.Two readers independently evaluated the above three types of images,and recorded the number of urinary stones,their anatomical locations,and whether there was residual iodine on the VUE images.Stone size and CT number were recorded only on the TUE images.Stone size,CT number,anatomical location,and iodine contrast agent were included in univariate and multivariate Logistic regression analyses to evaluate the factors affecting urinary stone detection rate using VUE images.Thresholds for detecting urinary stones on VUE images were determined using receiver operating characteristics(ROC)analysis.Results We detected 304 stones on TUE images,while the detection rates were 92.4%and 71.4%when using VUE(VP)and VUE(EP)images,respectively.Stone size and CT number were important factors influencing urinary stone detection rate using VUE(VP)and VUE(EP)images(P<0.01).The area under curve(AUC)of using stone size and CT number for detecting stones using the VUE(VP)images was up to 0.96,and as threshold values,stones with size larger than 3.52 mm and CT number greater than 469 HU were found to have high accuracy.However,the AUC decreased to 0.88 when we combined stone size,CT number and anatomical location using the VUE(EP)images.In addition,different contrast agents did not affect the detection rate of stones on the VUE(EP)images(P=0.57).The stone detection rate in the kidney was significantly lower than those on the VUE(EP)images(P<0.001).Conclusion VUE(VP)images provide better stone detection.Stone size and CT number have significant impacts on the stone detection rate using VUE images.The lower stone detection rate in the kidney on the VUE(EP)images is related to the residual iodine.
5.Treatment status of tyrosine kinase inhibitor for newly-diagnosed chronic myeloid leukemia: a domestic multi-centre retrospective real-world study
Xiaoshuai ZHANG ; Bingcheng LIU ; Xin DU ; Yanli ZHANG ; Na XU ; Xiaoli LIU ; Weiming LI ; Hai LIN ; Rong LIANG ; Chunyan CHEN ; Jian HUANG ; Yunfan YANG ; Huanling ZHU ; Ling PAN ; Xiaodong WANG ; Guohui LI ; Zhuogang LIU ; Yanqing ZHANG ; Zhenfang LIU ; Jianda HU ; Chunshui LIU ; Fei LI ; Wei YANG ; Li MENG ; Yanqiu HAN ; Li'e LIN ; Zhenyu ZHAO ; Chuanqing TU ; Caifeng ZHENG ; Yanliang BAI ; Zeping ZHOU ; Suning CHEN ; Huiying QIU ; Lijie YANG ; Xiuli SUN ; Hui SUN ; Li ZHOU ; Zelin LIU ; Danyu WANG ; Jianxin GUO ; Liping PANG ; Qingshu ZENG ; Xiaohui SUO ; Weihua ZHANG ; Yuanjun ZHENG ; Qian JIANG
Chinese Journal of Hematology 2024;45(3):215-224
Objective:To retrospectively analyze the treatment status of tyrosine kinase inhibitors (TKI) in newly diagnosed patients with chronic myeloid leukemia (CML) in China.Methods:Data of chronic phase (CP) and accelerated phase (AP) CML patients diagnosed from January 2006 to December 2022 from 77 centers, ≥18 years old, and receiving initial imatinib, nilotinib, dasatinib or flumatinib-therapy within 6 months after diagnosis in China with complete data were retrospectively interrogated. The choice of initial TKI, current TKI medications, treatment switch and reasons, treatment responses and outcomes as well as the variables associated with them were analyzed.Results:6 893 patients in CP ( n=6 453, 93.6%) or AP ( n=440, 6.4%) receiving initial imatinib ( n=4 906, 71.2%), nilotinib ( n=1 157, 16.8%), dasatinib ( n=298, 4.3%) or flumatinib ( n=532, 7.2%) -therapy. With the median follow-up of 43 ( IQR 22-75) months, 1 581 (22.9%) patients switched TKI due to resistance ( n=1 055, 15.3%), intolerance ( n=248, 3.6%), pursuit of better efficacy ( n=168, 2.4%), economic or other reasons ( n=110, 1.6%). The frequency of switching TKI in AP patients was significantly-higher than that in CP patients (44.1% vs 21.5%, P<0.001), and more AP patients switched TKI due to resistance than CP patients (75.3% vs 66.1%, P=0.011). Multi-variable analyses showed that male, lower HGB concentration and ELTS intermediate/high-risk cohort were associated with lower cytogenetic and molecular responses rate and poor outcomes in CP patients; higher WBC count and initial the second-generation TKI treatment, the higher response rates; Ph + ACA at diagnosis, poor PFS. However, Sokal intermediate/high-risk cohort was only significantly-associated with lower CCyR and MMR rates and the poor PFS. Lower HGB concentration and larger spleen size were significantly-associated with the lower cytogenetic and molecular response rates in AP patients; initial the second-generation TKI treatment, the higher treatment response rates; lower PLT count, higher blasts and Ph + ACA, poorer TFS; Ph + ACA, poorer OS. Conclusion:At present, the vast majority of newly-diagnosed CML-CP or AP patients could benefit from TKI treatment in the long term with the good treatment responses and survival outcomes.
6.Relationship between cognitive impairment and sleep parameters in stroke patients with obstructive sleep apnea
Hongchun QIAN ; Pingshu ZHANG ; Xiaodong YUAN ; Jianxin YUAN ; Lingyun CAO ; Liqin DUAN
Tianjin Medical Journal 2024;52(6):619-624
Objective To investigate the relationship between cognitive impairment and sleep parameters in acute ischemic stroke patients with obstructive sleep apnea(OSA).Methods A total of 343 patients with acute ischemic stroke and OSA were selected.The cognitive function was assessed using the simple mental state examination scale(MMSE).Patients were divided into the stroke with OSA and cognitive impairment group(MMSE<27 points,n=119)and the stroke with OSA without cognitive impairment group(MMSE≥27 points,n=224).General data,TOAST etiological classification and distribution of cerebral infarction lesions were collected.The intelligent sleep monitoring system was used to calculate apnea hypopnea index(AHI)and evaluate OSA.Objective sleep monitoring parameters were collected at night.Sleep monitoring was conducted within 24 h of admission and continuous monitoring for≥3 nights.Continuous monitoring duration≥7 h every night to obtain night sleep structure parameters.Multifactor Logistics regression was used to analyze the relationship between cognitive impairment and sleep parameters in stroke patients with OSA.Results Compared with the stroke with OSA without cognitive impairment group,the proportion of age,diabetes history and HHcy history,the proportion of patients with infarct lesions located in frontal,temporal,parietal,occipital,thalamus,basal ganglia,brainstem and hemioval center increased in the stroke with OSA and cognitive impairment group,and the number of years of education decreased,the number of waking times,the proportion of light sleep and AHI increased,the nighttime sleep efficiency and deep sleep period decreased(P<0.05).Logistics regression analysis showed that after controlling for years of education,age and other interference factors,nighttime sleep efficiency and AHI were strongly associated with cognitive impairment in acute ischemic stroke patients with OSA(P<0.05).The increased nighttime sleep efficiency was protective factor for cognitive impairment,and increased AHI was risk factor for cognitive impairment.Conclusion Cognitive impairment in acute ischemic stroke patients with OSA is closely related to sleep parameters,in which the increased sleep efficiency at night is a protective factor for cognitive impairment,and the increased AHI is a risk factor.
7.Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients (version 2024)
Yao LU ; Yang LI ; Leiying ZHANG ; Hao TANG ; Huidan JING ; Yaoli WANG ; Xiangzhi JIA ; Li BA ; Maohong BIAN ; Dan CAI ; Hui CAI ; Xiaohong CAI ; Zhanshan ZHA ; Bingyu CHEN ; Daqing CHEN ; Feng CHEN ; Guoan CHEN ; Haiming CHEN ; Jing CHEN ; Min CHEN ; Qing CHEN ; Shu CHEN ; Xi CHEN ; Jinfeng CHENG ; Xiaoling CHU ; Hongwang CUI ; Xin CUI ; Zhen DA ; Ying DAI ; Surong DENG ; Weiqun DONG ; Weimin FAN ; Ke FENG ; Danhui FU ; Yongshui FU ; Qi FU ; Xuemei FU ; Jia GAN ; Xinyu GAN ; Wei GAO ; Huaizheng GONG ; Rong GUI ; Geng GUO ; Ning HAN ; Yiwen HAO ; Wubing HE ; Qiang HONG ; Ruiqin HOU ; Wei HOU ; Jie HU ; Peiyang HU ; Xi HU ; Xiaoyu HU ; Guangbin HUANG ; Jie HUANG ; Xiangyan HUANG ; Yuanshuai HUANG ; Shouyong HUN ; Xuebing JIANG ; Ping JIN ; Dong LAI ; Aiping LE ; Hongmei LI ; Bijuan LI ; Cuiying LI ; Daihong LI ; Haihong LI ; He LI ; Hui LI ; Jianping LI ; Ning LI ; Xiying LI ; Xiangmin LI ; Xiaofei LI ; Xiaojuan LI ; Zhiqiang LI ; Zhongjun LI ; Zunyan LI ; Huaqin LIANG ; Xiaohua LIANG ; Dongfa LIAO ; Qun LIAO ; Yan LIAO ; Jiajin LIN ; Chunxia LIU ; Fenghua LIU ; Peixian LIU ; Tiemei LIU ; Xiaoxin LIU ; Zhiwei LIU ; Zhongdi LIU ; Hua LU ; Jianfeng LUAN ; Jianjun LUO ; Qun LUO ; Dingfeng LYU ; Qi LYU ; Xianping LYU ; Aijun MA ; Liqiang MA ; Shuxuan MA ; Xainjun MA ; Xiaogang MA ; Xiaoli MA ; Guoqing MAO ; Shijie MU ; Shaolin NIE ; Shujuan OUYANG ; Xilin OUYANG ; Chunqiu PAN ; Jian PAN ; Xiaohua PAN ; Lei PENG ; Tao PENG ; Baohua QIAN ; Shu QIAO ; Li QIN ; Ying REN ; Zhaoqi REN ; Ruiming RONG ; Changshan SU ; Mingwei SUN ; Wenwu SUN ; Zhenwei SUN ; Haiping TANG ; Xiaofeng TANG ; Changjiu TANG ; Cuihua TAO ; Zhibin TIAN ; Juan WANG ; Baoyan WANG ; Chunyan WANG ; Gefei WANG ; Haiyan WANG ; Hongjie WANG ; Peng WANG ; Pengli WANG ; Qiushi WANG ; Xiaoning WANG ; Xinhua WANG ; Xuefeng WANG ; Yong WANG ; Yongjun WANG ; Yuanjie WANG ; Zhihua WANG ; Shaojun WEI ; Yaming WEI ; Jianbo WEN ; Jun WEN ; Jiang WU ; Jufeng WU ; Aijun XIA ; Fei XIA ; Rong XIA ; Jue XIE ; Yanchao XING ; Yan XIONG ; Feng XU ; Yongzhu XU ; Yongan XU ; Yonghe YAN ; Beizhan YAN ; Jiang YANG ; Jiangcun YANG ; Jun YANG ; Xinwen YANG ; Yongyi YANG ; Chunyan YAO ; Mingliang YE ; Changlin YIN ; Ming YIN ; Wen YIN ; Lianling YU ; Shuhong YU ; Zebo YU ; Yigang YU ; Anyong YU ; Hong YUAN ; Yi YUAN ; Chan ZHANG ; Jinjun ZHANG ; Jun ZHANG ; Kai ZHANG ; Leibing ZHANG ; Quan ZHANG ; Rongjiang ZHANG ; Sanming ZHANG ; Shengji ZHANG ; Shuo ZHANG ; Wei ZHANG ; Weidong ZHANG ; Xi ZHANG ; Xingwen ZHANG ; Guixi ZHANG ; Xiaojun ZHANG ; Guoqing ZHAO ; Jianpeng ZHAO ; Shuming ZHAO ; Beibei ZHENG ; Shangen ZHENG ; Huayou ZHOU ; Jicheng ZHOU ; Lihong ZHOU ; Mou ZHOU ; Xiaoyu ZHOU ; Xuelian ZHOU ; Yuan ZHOU ; Zheng ZHOU ; Zuhuang ZHOU ; Haiyan ZHU ; Peiyuan ZHU ; Changju ZHU ; Lili ZHU ; Zhengguo WANG ; Jianxin JIANG ; Deqing WANG ; Jiongcai LAN ; Quanli WANG ; Yang YU ; Lianyang ZHANG ; Aiqing WEN
Chinese Journal of Trauma 2024;40(10):865-881
Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.
8.Physical activity, long-term fine particulate matter exposure and type 2 diabetes incidence: A prospective cohort study
Qian LI ; Fangchao LIU ; Keyong HUANG ; Fengchao LIANG ; Chong SHEN ; Jian LIAO ; Jianxin LI ; Chenxi YUAN ; Xueli YANG ; Jie CAO ; Shufeng CHEN ; Dongsheng HU ; Jianfeng HUANG ; Yang LIU ; Xiangfeng LU ; Dongfeng GU
Chronic Diseases and Translational Medicine 2024;10(3):205-215
Background::Despite the adverse effects of ambient fine particulate matter (PM 2.5) on type 2 diabetes and the beneficial role of physical activity (PA), the influence of PM 2.5 on the relationship between PA and type 2 diabetes remains unclear. Methods::In this prospective study with 71,689 participants, PA was assessed by a questionnaire and was categorized into quartiles for volume and three groups for intensity. Long-term PM 2.5 exposure was calculated using 1-km resolution satellite-based PM 2.5 estimates. PM 2.5 exposure and PA's effect on type 2 diabetes were assessed by cohort-stratified Cox proportional hazards models, individually and in combination. Results::In 488,166 person-years of follow-up, 5487 incident type 2 diabetes cases were observed. The association between PA and type 2 diabetes was modified by PM 2.5. Compared with the lowest quartile of PA volume, the highest quartile was associated with reduced type 2 diabetes risk in low PM 2.5 stratification (≤65.02 μg/m 3) other than in high PM 2.5 stratification (>65.02 μg/m 3), with the hazard ratio (HR) of 0.75 (95% confidence interval [CI]: 0.66-0.85) and 1.10 (95% CI: 0.99-1.22), respectively. Similar results were observed for PA intensity. High PM 2.5 exposure combined with the highest PA levels increased the risk of type 2 diabetes the most (HR= 1.79, 95% CI: 1.59-2.01 for PA volume; HR = 1.82, 95% CI: 1.64-2.02 for PA intensity). Conclusion::PA could reduce type 2 diabetes risk in low-pollution areas, but high PM 2.5 exposure may weaken or even reverse the protective effects of PA. Safety and health benefits of PA should be thoroughly assessed for long-term polluted residents.
9.Physical activity, long-term fine particulate matter exposure and type 2 diabetes incidence: A prospective cohort study
Qian LI ; Fangchao LIU ; Keyong HUANG ; Fengchao LIANG ; Chong SHEN ; Jian LIAO ; Jianxin LI ; Chenxi YUAN ; Xueli YANG ; Jie CAO ; Shufeng CHEN ; Dongsheng HU ; Jianfeng HUANG ; Yang LIU ; Xiangfeng LU ; Dongfeng GU
Chronic Diseases and Translational Medicine 2024;10(3):205-215
Background::Despite the adverse effects of ambient fine particulate matter (PM 2.5) on type 2 diabetes and the beneficial role of physical activity (PA), the influence of PM 2.5 on the relationship between PA and type 2 diabetes remains unclear. Methods::In this prospective study with 71,689 participants, PA was assessed by a questionnaire and was categorized into quartiles for volume and three groups for intensity. Long-term PM 2.5 exposure was calculated using 1-km resolution satellite-based PM 2.5 estimates. PM 2.5 exposure and PA's effect on type 2 diabetes were assessed by cohort-stratified Cox proportional hazards models, individually and in combination. Results::In 488,166 person-years of follow-up, 5487 incident type 2 diabetes cases were observed. The association between PA and type 2 diabetes was modified by PM 2.5. Compared with the lowest quartile of PA volume, the highest quartile was associated with reduced type 2 diabetes risk in low PM 2.5 stratification (≤65.02 μg/m 3) other than in high PM 2.5 stratification (>65.02 μg/m 3), with the hazard ratio (HR) of 0.75 (95% confidence interval [CI]: 0.66-0.85) and 1.10 (95% CI: 0.99-1.22), respectively. Similar results were observed for PA intensity. High PM 2.5 exposure combined with the highest PA levels increased the risk of type 2 diabetes the most (HR= 1.79, 95% CI: 1.59-2.01 for PA volume; HR = 1.82, 95% CI: 1.64-2.02 for PA intensity). Conclusion::PA could reduce type 2 diabetes risk in low-pollution areas, but high PM 2.5 exposure may weaken or even reverse the protective effects of PA. Safety and health benefits of PA should be thoroughly assessed for long-term polluted residents.
10.Prokaryotic expression, polyclonal antibody preparation, spatio-temporal expression profile and functional analysis of c-Myc of Helicoverpa armigera (Lepidoptera: Noctuidae).
Qian SUO ; Xiaoyan SUN ; Ying ZHANG ; Yujing WANG ; Kaiyu LIU ; Hong YANG ; Huazhu HONG ; Jianxin PENG ; Rong PENG
Chinese Journal of Biotechnology 2023;39(7):2730-2742
c-Myc protein encoded by c-Myc (cellular-myelocytomatosis viral oncogene) gene regulates the related gene expression through the Wnt/β-catenin signaling pathway, and has received extensive attention in recent years. The purpose of this study was to express Helicoverpa armigera c-Myc gene (Ha-c-Myc) by using prokaryotic expression system, prepare the polyclonal antibody, examine the spatio-temporal expression profile of Ha-c-Myc, and investigate the possible function of Ha-c-Myc in regulating H. armigera sterol carrier protein-2 (SCP-2) gene expression. The Ha-c-Myc gene was amplified by PCR and cloned into a prokaryotic expression plasmid pET-32a(+). The recombinant plasmid pET-32a-Ha-c-Myc was transformed into Escherichia coli BL21. IPTG was used to induce the expression of the recombinant protein. Protein was purified by Ni2+-NTA column and used to immunize New Zealand rabbits for preparing the polyclonal antibody. The Ha-c-Myc expression levels in different developmental stages (egg, larva, prepupa, pupa, and adult) of H. armigera and different tissues (midgut, fat body, head, and epidermis) of the prepupa were determined by real-time quantitative reverse transcription PCR (qRT-PCR). Ha-c-Myc siRNA was synthesized and transfected into H. armigera Ha cells. The relative mRNA levels of Ha-c-Myc and HaSCP-2 in Ha cells were detected by qRT-PCR. Results showed that the pET-32a-Ha-c-Myc recombinant plasmid was constructed. The soluble Ha-c-Myc protein of about 65 kDa was expressed in E. coli. The polyclonal antibody was prepared. Western blotting analysis suggested that the antibody had high specificity. Enzyme linked immunosorbent assay (ELISA) showed that the titer of the antibody was high. Ha-c-Myc gene expressed at all developmental stages, with high levels in the early and late instars of larva, and the prepupal stage. Tissue expression profiles revealed that Ha-c-Myc expressed in various tissues of prepupa, with high expression level in the midgut, but low levels in the epidermis and fat body. RNAi results showed that the knockdown of Ha-c-Myc expression significantly affected transcription of HaSCP-2, leading to a 50% reduction in HaSCP-2 mRNA expression level. In conclusion, the Ha-c-Myc was expressed through a prokaryotic expression system, and the polyclonal anti-Ha-c-Myc antibody was obtained. Ha-c-Myc may promote the expression of HaSCP-2 and play an important role in the lipid metabolism of H. armigera. These results may facilitate further study on the potential role and function mechanism of Ha-c-Myc in H. armigera and provide experimental data for exploring new targets of green pesticides.
Animals
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Rabbits
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Escherichia coli/metabolism*
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Enzyme-Linked Immunosorbent Assay
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Moths/genetics*
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Blotting, Western
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Larva/genetics*
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Isoantibodies/metabolism*
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Antibody Specificity

Result Analysis
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