1.Clinical Efficacy and Economic Evaluation of 1293 Non-Severe Adult Patients with Community-Acquired Pneumonia Treated by the Jiangsu Traditional Chinese Medicine Diagnosis and Treatment Protocol for Dominant Diseases:A Multicenter,Retrospective Real-World Cohort Study
Ye MA ; Yeqing JI ; Zhichao WANG ; Fanchao FENG ; Mingzhi PU ; Hong LYU ; Xiaodong HU ; Gaohua FENG ; Xiaoqian FANG ; Guicai ZHANG ; Yanfen TANG ; Yeqing ZHANG ; Yao ZHUFU ; Wenpan PENG ; Hao WANG ; Cheng GU ; Zhichao ZHANG ; Shuang YANG ; Xinyu SUN ; Qi ZHAO ; Aojie GUO ; Xin TONG ; Zhuoyue WU ; Xiaoxiao WANG ; Jia LIU ; Hailang HE ; Xianmei ZHOU
Journal of Traditional Chinese Medicine 2026;67(9):966-974
ObjectiveTo evaluate the clinical efficacy and economic value of the Jiangsu Traditional Chinese Medicine (TCM) Diagnosis and Treatment Protocol for Dominant Diseases (abbreviated as the Diagnosis and Treatment Protocol) in adult patients with non-severe community-acquired pneumonia (CAP) based on real-world clinical data. MethodsA retrospective real-world cohort study was conducted using electronic medical records of adult patients hospitalized for non-severe CAP from September 1st, 2023 to December 31st, 2024 across 10 TCM hospitals in Jiangsu province. Patients were classified into an exposure group and a non-exposure group based on whether they received Chinese herbal medicine (CHM) according to the Diagnosis and Treatment Protocol. The non-exposure group received only conventional western medicine, while the exposure group additionally received differentiated CHM for at least five consecutive days. Outcomes were compared between two patient groups, including cough resolution rate, sputum resolution rate (assessed by volume, color, and consistency), incidence of abnormal C-reactive protein (CRP), incidence of abnormal white blood cell (WBC) count, and radiographic resolution rate of pulmonary infiltrates on chest imaging. Multivariable logistic regression was performed to identify factors influencing clinical efficacy. Subgroup analyses were conducted according to age, gender, smoking status, history of hypertension, and pneumonia severity score (CURB-65), and the efficacy of treatment for cough and sputum was analyzed within each subgroup. Cost-effectiveness analysis was conducted using cough resolution rate as the outcome measure, evaluating the pharmacoeconomics of the two groups. ResultsA total of 1688 patients were included with 1293 in the exposure group and 395 in the non-exposure group. Compared to the non-exposure group, the exposure group demonstrated significantly higher resolution rates of cough, sputum volume, color, and consistency, as well as a significantly lower incidence of abnormal CRP (P<0.05). No statistically significant difference was observed between the groups in terms of abnormal WBC count and radiographic resolution rate of pulmonary infiltrates (P>0.05). Logistic regression analysis showed that the cough resolution rate in the exposure group was 1.83 times that of the non-exposure group, while the probabilities of resolution in sputum volume, color, and consistency were 1.37, 2.09, and 1.56 times those of the non-exposure group, respectively (P<0.05). Subgroup analyses showed that the exposure group achieved significantly higher cough resolution rates across most subgroups except for populations with a CURB-65 score ≥2 or those with a history of hypertension (P<0.05). Specifically, among females, patients aged ≥18 and <65 years, non-smokers, those without hypertension, and those with a CURB-65 score of 0, the exposure group showed a higher cough resolution rate than the non-exposure group (P<0.05). From an economic perspective, total hospitalization cost, length of stay, antibiotic cost, and CHM cost all differed significantly between groups (P<0.05). The cost-effectiveness ratio (CER) was 10,788.80 CNY/case in the exposure group, while 22,513.80 CNY/case in the non-exposure group. This implies that, compared with the exposure group, the non-exposure group incurred an additional 17,302.27 CNY to achieve one case of cough resolution. When the willingness-to-pay threshold ranged from 0 to 50,000 CNY, the probability of economic advantage was consistently higher in the exposure group than in the non-exposure group. ConclusionOn the basis of conventional western medicine, the addition of CHM in accordance with the Diagnosis and Treatment Protocol can effectively improve clinical symptoms, reduce inflammatory markers, promote clinical recovery, and is more cost-effective in treating adults with non-severe CAP.
2.Efficacy and Economic Evaluation of Weishi Qingjin Formula (苇石清金方)in the Treatment of Adult Community-Acquired Pneumonia with Phlegm-Heat Obstructing the Lung Syndrome:A Multicenter Retrospective Real-World Cohort Study
Yeqing JI ; Ye MA ; Zhichao WANG ; Fanchao FENG ; Mingzhi PU ; Hong LYU ; Xiaodong HU ; Gaohua FENG ; Xiaoqian FANG ; Guicai ZHANG ; Yanfen TANG ; Yeqing ZHANG ; Yao ZHUFU ; Wenpan PENG ; Hao WANG ; Cheng GU ; Zhichao ZHANG ; Shuang YANG ; Xinyu SUN ; Qi ZHAO ; Aojie GUO ; Xin TONG ; Zhuoyue WU ; Xiaoxiao WANG ; Jia LIU ; Hailang HE ; Xianmei ZHOU
Journal of Traditional Chinese Medicine 2026;67(9):975-984
ObjectiveTo observe the real‑world effectiveness and economic outcomes of Weishi Qingjin Formula (苇石清金方, WQF) in the treatment of adult community‑acquired pneumonia (CAP) with phlegm‑heat obstructing the lung syndrome. MethodsBased on a multicenter, real-world retrospective cohort study, clinical data were collected from hospitalized adult patients diagnosed with non‑severe CAP and phlegm‑heat obstructing the lung syndrome in 10 traditional Chinese medicine (TCM) hospitals in Jiangsu province. Patients were divided into an exposure group (those who received oral WQF) and a non‑exposure group (those who did not). The following outcomes were compared between the two groups before and after treatment, which were remission rates of clinical symptoms including cough, expectoration (sputum volume, color, consistency), and chest pain, levels of inflammatory markers including C‑reactive protein (CRP) and white blood cell count (WBC), and the rate of pulmonary inflammatory absorption on chest CT. Subgroup analyses were performed based on age, gender, smoking status, presence of hypertension, and the severity of community-acquired pneumonia (CURB‑65) score, comparing the two groups in terms of cough remission rate, chest pain remission rate, and chest CT absorption rate. For health economic evaluation, cost‑effectiveness analysis was used to calculate the cost‑effectiveness ratio (CER) and incremental cost‑effectiveness ratio (ICER). Univariate sensitivity analysis and probabilistic sensitivity analysis were performed to test the robustness of the results. ResultsA total of 647 patients in the exposure group and 1491 patients in the non-exposure group were included in the final statistical analysis. There was no statistically significant difference in length of hospital stay, gender, marital status, smoking history, bronchoscopy history, and comorbidities between the groups (P>0.05), but age, CURB-65 score, and antibiotic use. The exposure group had significantly higher remission rates of cough and sputum consistency than the non-exposure group (P<0.05). After adjusting for confounders using propensity score matching and logistic regression, the cough remission rate in the exposure group was 1.49 times that of the non-exposure group (P<0.01). No significant difference was observed between groups in the reduction rates of CRP and WBC, and in the rate of pulmonary inflammatory absorption on chest CT (P>0.05). Subgroup analyses revealed that the cough remission rate in the exposure group was significantly better than that in the non-exposure group except for patients aged ≥65 years, smokers, hypertensive patients, those using other type antibiotics or not using antibiotics, and those with a CURB-65 score ≥1 (P<0.05). Among smokers, the chest pain remission rate in the exposure group was 4.38 times that of the non-exposure group (P<0.01). No significant difference in chest CT absorption rate was found between groups across subgroups of gender, age, hypertension status, or antibiotic type (P>0.05). In terms of economic evaluation, CER was 10,877.60 CNY/case in the exposure group and 16,773.10 CNY/case in the non-exposure group. Compared to the exposure group, the non-exposure group incurred an additional 15,034.26 CNY to achieve one case of cough resolution, indicating a more favorable cost-effectiveness profile. Probabilistic sensitivity analysis yielded results consistent with the cost-effectiveness analysis, confirming the robustness of the findings. ConclusionWQF demonstrates significant efficacy in improving cough symptoms in the treatment of adult CAP with phlegm-heat obstructing the lung syndrome, and also exhibits favorable economic benefits.
3.Clinical Efficacy and Economic Evaluation of 1293 Non-Severe Adult Patients with Community-Acquired Pneumonia Treated by the Jiangsu Traditional Chinese Medicine Diagnosis and Treatment Protocol for Dominant Diseases:A Multicenter,Retrospective Real-World Cohort Study
Ye MA ; Yeqing JI ; Zhichao WANG ; Fanchao FENG ; Mingzhi PU ; Hong LYU ; Xiaodong HU ; Gaohua FENG ; Xiaoqian FANG ; Guicai ZHANG ; Yanfen TANG ; Yeqing ZHANG ; Yao ZHUFU ; Wenpan PENG ; Hao WANG ; Cheng GU ; Zhichao ZHANG ; Shuang YANG ; Xinyu SUN ; Qi ZHAO ; Aojie GUO ; Xin TONG ; Zhuoyue WU ; Xiaoxiao WANG ; Jia LIU ; Hailang HE ; Xianmei ZHOU
Journal of Traditional Chinese Medicine 2026;67(9):966-974
ObjectiveTo evaluate the clinical efficacy and economic value of the Jiangsu Traditional Chinese Medicine (TCM) Diagnosis and Treatment Protocol for Dominant Diseases (abbreviated as the Diagnosis and Treatment Protocol) in adult patients with non-severe community-acquired pneumonia (CAP) based on real-world clinical data. MethodsA retrospective real-world cohort study was conducted using electronic medical records of adult patients hospitalized for non-severe CAP from September 1st, 2023 to December 31st, 2024 across 10 TCM hospitals in Jiangsu province. Patients were classified into an exposure group and a non-exposure group based on whether they received Chinese herbal medicine (CHM) according to the Diagnosis and Treatment Protocol. The non-exposure group received only conventional western medicine, while the exposure group additionally received differentiated CHM for at least five consecutive days. Outcomes were compared between two patient groups, including cough resolution rate, sputum resolution rate (assessed by volume, color, and consistency), incidence of abnormal C-reactive protein (CRP), incidence of abnormal white blood cell (WBC) count, and radiographic resolution rate of pulmonary infiltrates on chest imaging. Multivariable logistic regression was performed to identify factors influencing clinical efficacy. Subgroup analyses were conducted according to age, gender, smoking status, history of hypertension, and pneumonia severity score (CURB-65), and the efficacy of treatment for cough and sputum was analyzed within each subgroup. Cost-effectiveness analysis was conducted using cough resolution rate as the outcome measure, evaluating the pharmacoeconomics of the two groups. ResultsA total of 1688 patients were included with 1293 in the exposure group and 395 in the non-exposure group. Compared to the non-exposure group, the exposure group demonstrated significantly higher resolution rates of cough, sputum volume, color, and consistency, as well as a significantly lower incidence of abnormal CRP (P<0.05). No statistically significant difference was observed between the groups in terms of abnormal WBC count and radiographic resolution rate of pulmonary infiltrates (P>0.05). Logistic regression analysis showed that the cough resolution rate in the exposure group was 1.83 times that of the non-exposure group, while the probabilities of resolution in sputum volume, color, and consistency were 1.37, 2.09, and 1.56 times those of the non-exposure group, respectively (P<0.05). Subgroup analyses showed that the exposure group achieved significantly higher cough resolution rates across most subgroups except for populations with a CURB-65 score ≥2 or those with a history of hypertension (P<0.05). Specifically, among females, patients aged ≥18 and <65 years, non-smokers, those without hypertension, and those with a CURB-65 score of 0, the exposure group showed a higher cough resolution rate than the non-exposure group (P<0.05). From an economic perspective, total hospitalization cost, length of stay, antibiotic cost, and CHM cost all differed significantly between groups (P<0.05). The cost-effectiveness ratio (CER) was 10,788.80 CNY/case in the exposure group, while 22,513.80 CNY/case in the non-exposure group. This implies that, compared with the exposure group, the non-exposure group incurred an additional 17,302.27 CNY to achieve one case of cough resolution. When the willingness-to-pay threshold ranged from 0 to 50,000 CNY, the probability of economic advantage was consistently higher in the exposure group than in the non-exposure group. ConclusionOn the basis of conventional western medicine, the addition of CHM in accordance with the Diagnosis and Treatment Protocol can effectively improve clinical symptoms, reduce inflammatory markers, promote clinical recovery, and is more cost-effective in treating adults with non-severe CAP.
4.Efficacy and Economic Evaluation of Weishi Qingjin Formula (苇石清金方)in the Treatment of Adult Community-Acquired Pneumonia with Phlegm-Heat Obstructing the Lung Syndrome:A Multicenter Retrospective Real-World Cohort Study
Yeqing JI ; Ye MA ; Zhichao WANG ; Fanchao FENG ; Mingzhi PU ; Hong LYU ; Xiaodong HU ; Gaohua FENG ; Xiaoqian FANG ; Guicai ZHANG ; Yanfen TANG ; Yeqing ZHANG ; Yao ZHUFU ; Wenpan PENG ; Hao WANG ; Cheng GU ; Zhichao ZHANG ; Shuang YANG ; Xinyu SUN ; Qi ZHAO ; Aojie GUO ; Xin TONG ; Zhuoyue WU ; Xiaoxiao WANG ; Jia LIU ; Hailang HE ; Xianmei ZHOU
Journal of Traditional Chinese Medicine 2026;67(9):975-984
ObjectiveTo observe the real‑world effectiveness and economic outcomes of Weishi Qingjin Formula (苇石清金方, WQF) in the treatment of adult community‑acquired pneumonia (CAP) with phlegm‑heat obstructing the lung syndrome. MethodsBased on a multicenter, real-world retrospective cohort study, clinical data were collected from hospitalized adult patients diagnosed with non‑severe CAP and phlegm‑heat obstructing the lung syndrome in 10 traditional Chinese medicine (TCM) hospitals in Jiangsu province. Patients were divided into an exposure group (those who received oral WQF) and a non‑exposure group (those who did not). The following outcomes were compared between the two groups before and after treatment, which were remission rates of clinical symptoms including cough, expectoration (sputum volume, color, consistency), and chest pain, levels of inflammatory markers including C‑reactive protein (CRP) and white blood cell count (WBC), and the rate of pulmonary inflammatory absorption on chest CT. Subgroup analyses were performed based on age, gender, smoking status, presence of hypertension, and the severity of community-acquired pneumonia (CURB‑65) score, comparing the two groups in terms of cough remission rate, chest pain remission rate, and chest CT absorption rate. For health economic evaluation, cost‑effectiveness analysis was used to calculate the cost‑effectiveness ratio (CER) and incremental cost‑effectiveness ratio (ICER). Univariate sensitivity analysis and probabilistic sensitivity analysis were performed to test the robustness of the results. ResultsA total of 647 patients in the exposure group and 1491 patients in the non-exposure group were included in the final statistical analysis. There was no statistically significant difference in length of hospital stay, gender, marital status, smoking history, bronchoscopy history, and comorbidities between the groups (P>0.05), but age, CURB-65 score, and antibiotic use. The exposure group had significantly higher remission rates of cough and sputum consistency than the non-exposure group (P<0.05). After adjusting for confounders using propensity score matching and logistic regression, the cough remission rate in the exposure group was 1.49 times that of the non-exposure group (P<0.01). No significant difference was observed between groups in the reduction rates of CRP and WBC, and in the rate of pulmonary inflammatory absorption on chest CT (P>0.05). Subgroup analyses revealed that the cough remission rate in the exposure group was significantly better than that in the non-exposure group except for patients aged ≥65 years, smokers, hypertensive patients, those using other type antibiotics or not using antibiotics, and those with a CURB-65 score ≥1 (P<0.05). Among smokers, the chest pain remission rate in the exposure group was 4.38 times that of the non-exposure group (P<0.01). No significant difference in chest CT absorption rate was found between groups across subgroups of gender, age, hypertension status, or antibiotic type (P>0.05). In terms of economic evaluation, CER was 10,877.60 CNY/case in the exposure group and 16,773.10 CNY/case in the non-exposure group. Compared to the exposure group, the non-exposure group incurred an additional 15,034.26 CNY to achieve one case of cough resolution, indicating a more favorable cost-effectiveness profile. Probabilistic sensitivity analysis yielded results consistent with the cost-effectiveness analysis, confirming the robustness of the findings. ConclusionWQF demonstrates significant efficacy in improving cough symptoms in the treatment of adult CAP with phlegm-heat obstructing the lung syndrome, and also exhibits favorable economic benefits.
5.Prokaryotic expression of Echinococcus granulosus Polo-like kinase 2 and immunoprotective efficacy of its recombinant protein
Xue WANG ; Mingzhi YAN ; Wenjing QI ; Chuanchuan WU ; Guowu ZHANG ; An GENG ; Mengxiao TIAN ; Jun LI ; Wenbao ZHANG
Chinese Journal of Schistosomiasis Control 2026;38(2):184-193
Objective To prepare the recombinant Echinococcus granulosus Polo-like kinase 2 (rEgPLK2) protein and evaluate its immunoprotective efficacy against cystic echinococcosis, so as to provide insights into research and development of novel vaccines against echinococcosis. Methods The Polo-like kinase (PLK) protein sequences were retrieved from 12 species in the NCBI protein database, including E. granulosus and E. multilocularis. Multiple sequence alignment was performed using the Clustal Omega program, and structural visualization and homology analysis were conducted using the ESPript 3.2 program. The recombinant plasmid pET-30a-EgPLK2 was transformed into BL21(DE3) competent cells. Protein expression was induced with isopropyl-β-D-thiogalactoside (IPTG), and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was performed to characterize the expression and molecular weight of the rEgPLK2 protein. The purified rEgPLK2 protein was thoroughly emulsified with Freund’s complete adjuvant at a 1 : 1 volume ratio. Two New Zealand white rabbits were immunized with multipoint subcutaneous injection on the back at a dose of 300 μg per rabbit for primary immunization. For booster immunizations, the protein was emulsified with Freund’s incomplete adjuvant at a 1 : 1 volume ratio and administered on days 14, 28, and 42 after the primary immunization at a dose of 150 μg per rabbit. Serum was sampled from the rabbit ear vein on day 7 after the final immunization to yield anti-rEgPLK2 polyclonal antibodies. Antibody titer was determined by indirect enzyme-linked immunosorbent assay (ELISA), and antibody specificity was verified by Western blotting. The tissue localization of the EgPLK2 protein was detected in E. granulosus protoscoleces and adult worms using immunofluorescence assay (IFA). Eighteen 6- to 8-week-old female SPF-grade BALB/c mice were randomly divided into three groups, including the blank control group, rEgPLK2-ISA immunization group, and PBS-ISA adjuvant control group, of 6 mice each group. Mice in the rEgPLK2-ISA immunization group and PBSISA group received three primary immunizations via intramuscular injection, and animals in the rEgPLK2-ISA immunization group was inoculated with immunogens prepared by emulsifying rEgPLK2 protein with ISA 201 adjuvant at a 1 : 1 volume ratio (6 μg per mouse), while mice in the PBS-ISA adjuvant control group received an equal volume of PBS emulsified with ISA adjuvant at a 1 : 1 volume ratio. A fourth booster immunization was administered via intraperitoneal injection. Mice in the rEgPLK2-ISA immunization group received a booster immunization with 8 μg of rEgPLK2 protein per mouse, and animals in the PBS-ISA group received an equal volume of PBS, with immunizations given at 2-week intervals. Mice in the blank control group were given no treatment, and housed under standard conditions. Tail vein blood was collected from all mice 7 days after the final immunization, and levels of specific anti-rEgPLK2 IgG antibody and its subclasses (IgG1, IgG2a, IgG2b, IgG3) were measured by indirect ELISA. E. granulosus infection was modelled in mice through injection with 1 000 E. granulosus protoscoleces via intrahepatic portal vein in the rEgPLK2-ISA immunization group and PBS-ISA adjuvant control group 2 weeks after the last immunization. All mice were sacrificed and dissected. The number of cysts was counted in mouse livers, and the cyst reduction rate was calculated. Liver tissues were processed for paraffin sectioning and stained with hematoxylin and eosin (HE), and histopathological changes were examined under a light microscope. Results Sequence analysis revealed that EgPLK2 shared a high amino acid sequence homology with E. multilocularis PLK2 (EmPLK2) and contained the typical domains of the Polo-like kinase family, including the serine/threonine protein kinase catalytic domain (STKc) and Polo-box. The IPTG-induced rEgPLK2 protein was mainly expressed in the form of inclusion bodies, and the purified rEgPLK2 protein showed a relative molecular mass of approximately 70 kDa. The prepared rabbit anti-rEgPLK2 polyclonal antibody had a titer of 1 : 256 000, and Western blotting assay showed that this anti-body specifically recognized the rEgPLK2 protein with a relative molecular mass of approximately 70 kDa. Immunofluorescence assay showed that the EgPLK2 protein was localized in the excretory bladder and rostellum of E. granulosus protoscoleces, as well as the tegument, suckers, and inter-proglottid junctions of adult worms. Immunoprotective assay showed that the serum levels of specific anti-rEgPLK2 IgG, IgG1, IgG2a, and IgG2b antibodies were 2.92 ± 0.49, 0.33 ± 0.10, 0.31 (0.36), and 3.12 (1.73) in mice in the rEgPLK2-ISA immunization group, which were all significantly higher than those in the PBS-ISA adjuvant control group (0.14 ± 0.04, 0.07 ± 0.01, 0.12 ± 0.04, and 0.11 ± 0.04, respectively) (t = 19.28 and 8.46, Z = 3.75 and 4.15; all P values < 0.001); however, there was no significant difference in the serum anti-IgG3 antibody level between the rEgPLK2-ISA immunization group and the PBS-ISA adjuvant control group [0.07 (0.01) vs. 0.073 (0.07); Z = 0.69, P > 0.05)]. In the mouse model of E. granulosus infections, the area of hepatic lesions was reduced and the inflammatory infiltration was alleviated in the rEgPLK2-ISA immunization group than in the PBS-ISA adjuvant control group, and the number of hepatic cysts was higher in the PBS-ISA adjuvant control group than in the rEgPLK2-ISA immunization group [8.00 (2.00) vs. 1.00 (0.75); Z = −2.93, P < 0.01], with a cyst reduction rate of 80.40%. Indirect ELISA assay measured higher serum levels of specific anti-rEgPLK2 IgG (3.28 ± 0.48 vs. 0.11 ± 0.04; t = 15.86, P < 0.01), IgG1 (0.29 ± 0.02 vs. 0.09 ± 0.01; t = 15.67, P < 0.01), IgG2a [3.71 (1.09) vs. 0.08 (0.03); Z = 2.88, P < 0.01], and IgG2b antibodies [3.34 (1.01) vs. 0.08 (0.03); Z = 2.88, P < 0.01] in the rEgPLK2-ISA immunization group than in the PBS-ISA adjuvant control group, and there was no significant difference in the serum level of the specific anti-rEgPLK2 IgG3 antibody between the rEgPLK2-ISA immunization group and the PBS-ISA adjuvant control group (0.07 ± 0.01 vs. 0.07 ± 0.01; t = 1.29, P > 0.05). Conclusions The prokaryotic expression system has been successfully constructed for the EgPLK2 gene and the anti-rEgPLK2 polyclonal antibody has been obtained. The rEgPLK2 protein exhibits a high immunogenicity, and is effective to protect against E. granulosus infection, and inhibits cyst development, which is a promising candidate vaccine target against cystic echinococcosis.
6.Prokaryotic expression, purification and immunogenicity of SARS-CoV-2 omicron variant nucleocapsid protein.
Zewen TU ; Quansheng WANG ; Shiguo LIU ; Haosen LIU ; Chunyan ZENG ; Juanjuan XIE ; Mingzhi LI ; Jingcai LI ; Min WANG ; Shiqi WENG ; Lumei KANG ; Lingbao KONG
Chinese Journal of Cellular and Molecular Immunology 2025;41(8):735-743
Objective The study aims to investigate the immunological functions of the nucleocapsid (N) protein of the novel coronavirus Omicron (BA.1, BA.2) and evaluate the differences among different N proteins of mutant strains in immunogenicity. Methods By aligning sequences, the mutation sites of the Omicron (BA.1, BA.2) N protein relative to prototype strain of the novel coronavirus (Wuhan-Hu-1) were determined. The pET-28a-N-Wuhan-Hu-1 plasmid was used as template to construct pET-28a-BA.1/BA.2-N through single point mutation or homologous recombination. The three kinds of N protein were expressed in prokaryotic system, purified through Ni-NTA affinity chromatography, and then immunized into mice. The titer and reactivity of the polyclonal antibody, as well as the expression level of IL-1β and IFN-γ in mouse spleen cells, were detected using indirect ELISA and Western blot assay. Results The constructed prokaryotic expression plasmids were successfully used to express the Wuhan-Hu-1 N, BA.1 N, and BA.2 N proteins in E.coli BL21(DE3) at 37 DegreesCelsius for 4 hours. The indirect ELISA test showed that the titers of polyclonal antibody prepared by three N proteins were all 1:51 200. All three N proteins can increase the expression of IFN-γ and IL-1β cytokines, but the effect of Omicron N protein in activing two cytokines was more obvious than that of Wuhan-Hu-1 N protein. Conclusion The study obtained three new coronavirus N proteins and polyclonal antibodies, and confirmed that mutations in the amino acid sites of the N protein can affect its immunogenicity. This provides a basis for developing rapid diagnostic methods targeting N protein of different novel coronavirus variants.
Animals
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Mice
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SARS-CoV-2/genetics*
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Coronavirus Nucleocapsid Proteins/immunology*
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Nucleocapsid Proteins/isolation & purification*
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COVID-19/immunology*
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Antibodies, Viral/immunology*
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Mice, Inbred BALB C
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Interferon-gamma/metabolism*
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Interleukin-1beta/metabolism*
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Female
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Escherichia coli/metabolism*
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Mutation
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Humans
7.Effect of PK-15 cell line stably expressing UL54 protein on replication efficiency of UL54 gene deletion recombinant pseudo rabies virus
Mingzhi LI ; Hongxia WU ; Yimin WANG ; Yuan SUN
Chinese Journal of Veterinary Science 2025;45(6):1117-1125
The UL54 protein is a nonstructural protein of the pseudorabies virus(PRV).Electron microscopy observations showed that most of particles of recombinant PRV(rPRV-△UL54)with a deletion of the UL54 gene had only a nucleocapsid and lacked an intact vesicle structure.The in-complete structure of the viral particles led to a significant reduction in the infection efficiency of the virus,which could not be applied in the study of the gene function and mechanism of action of UL54.In this study,the UL54 gene was cloned into the lentiviral vector pLVX-IRES-Puro,and af-ter lentiviral transduction,the UL54 gene was integrated into the PK-15 cell genome.A cell line(PK-15-UL54)overexpressing UL54 protein was successfully constructed and characterized by PCR,indirect immunofluorescence assay and protein immunoblotting.Replication was assayed in the PK-15-UL54 cell line after inoculation with UL54 gene deletion virus(rPRV-△UL54-Re).Elec-tron microscopy observation showed that the rPRV-△UL54-Re virus had a complete viral particle structure.Fluorescence microscopy observations showed that rPRV-△UL54-Re virus could normal-ly infect cells and replicate.The results of quantitative PCR assay showed that the replication efficiency of rPRV-△UL54-Re virus increased.Therefore,PK-15-UL54 cell line is an important tool to improve the replication efficiency of UL54 gene deletion recombinant pseudorabies virus and plays an important role in exploring the mechanistic study of UL54 gene regulation of PRV repli-cation.
8.Effect of miR-532-3p on macrophage polarization in rats with chronic kidney disease through targeted inhibition of Notch1 signal pathway
Mingzhi XU ; Na AN ; Yafei BAI ; Ruman CHEN ; Jiqing HE ; Chunli WANG ; Yonghui QI ; Mingjiao PAN
Chinese Journal of Immunology 2025;41(2):310-314,319
Objective:To analyze effect of miR-532-3p on macrophage polarization in rats with chronic kidney disease(CKD)through targeted inhibition of Notch1 signal pathway.Methods:A total of 75 SD rats were divided into control group,CKD group,ago-NC group,ago-miR-532-3p group and FLI-06 group,except for control group,CKD models were constructed and corresponding plas-mids and inhibitors were injected,control group and CKD group were replaced with same amount of normal saline.Serum creatinine(Scr),blood urea nitrogen(BUN),TNF-α,IL-1β and IL-10 were measured by ELISA,HE staining and Masson staining were used to observe renal tissue pathology and renal fibrosis in rats,flow cytometry was used to detect CD11c+and CD206+of macrophages,miR-532-3p expression in rat kidney tissue was detected by qRT-PCR,Notch1 protein was detected by Western blot;target binding of miR-532-3p to Notch1 was determined by double luciferase reporter gene.Results:Structure of glomerulus,renal tubules and epithelial cells was complete,cell boundaries were clear,and cells were arranged neatly in control group;glomerular epithelial cell necrosis,mesangial matrix,glomerulosclerosis,inflammatory cell infiltration and renal fibrosis were increased in CKD group;compared with CKD group,damage degree of glomerulus and tubules,inflammatory infiltration cells and renal fibrosis degree in ago-miR-532-3p group and FLI-06 group were reduced;compared with control group,serum Scr,BUN,TNF-α,IL-1β,proportion of CD11c+,CD11c+/CD206+and Notch1 protein expression in macrophages of renal tissue were increased,serum IL-10 level,CD206+and miR-532-3p in renal tissue were decreased(P<0.05);compared with CKD group,serum Scr,BUN,TNF-α,IL-1β,proportion of CD11c+,CD11c+/CD206+and Notch1 protein expression in macrophages of renal tissue in ago-miR-532-3p group and FLI-06 group were decreased,serum IL-10 level,CD206+and miR-532-3p in renal tissue were increased(P<0.05);miR-532-3p targeted Notch1,and overexpression of miR-532-3p inhibited Notch1 protein expression.Conclusion:Promoting expression of miR-532-3p protects kidney tissue of CKD rats by inhibiting Notch1 pathway,which may be due to regulating polarization of macrophages.
9.Network Pharmacological Study on Active Compounds of Astragalus and Magnolia officinalis Against Prostate Cancer
Liyue REN ; Mingzhi ZHAO ; Sijie WANG ; Qin LIU ; Jiajia LIU
Journal of Kunming Medical University 2025;46(9):63-71
Objective To comprehensively analyze the mechanism of Astragalus and Magnolia officinalis in treating prostate cancer based on the principles of network pharmacology.Methods Active molecular targets of Astragalus and Magnolia officinalis were predicted using the TCMSP and SwissTargetPrediction databases.Prostate cancer-related targets were screened via Genecards,DisGeNET,and OMIM databases.A"disease-active ingredient-target"network was constructed using Venny software,identifying 69 candidate key target genes.A protein-protein interaction(PPI)network was built using the STRING database,followed by GO and KEGG enrichment pathway analyses performed with R language.Constructing a subcutaneous tumor model in nude mice through in vivo experiments and intervening with active ingredients from Astragalus membranaceus and Magnolia offi-cinalis.Results Molecular docking analysis revealed that active components such as astragaloside IV(MOL000438)and honokiol(MOL000398)exhibited significant binding activity with the key target proteins of prostate cancer,including AKT1,ESR1,PPARG,PTGS2,and SRC.Notably,Honokiol demonstrated a binding energy of-8.7 kcal/mol with estrogen receptor α(ESR1,PDB:1a52),forming stable hydrogen bond interaction with the LEU-391 residue.The in vivo experiments further confirmed that the Astragalus-Magnolia active component group showed smaller subcutaneous xenograft tumor volumes in nude mice as compared to the model group(P<0.05).Immunohistochemical analysis revealed significant downregulation of PPARG and PTGS2 protein expression in tumor tissues(P<0.05).QPCR results indicated that the formula bidirectionally regulated gene expression:pro-apoptotic factor AKT1 was upregulated(P<0.05),while cancer-associated genes PTGS2,PPARG,SRC,and ESR1 were downregulated(P<0.05).Conclusion The combination of Astragalus and Magnolia may exert anti-prostate cancer effects through multi-target and multi-pathway synergistic mechanisms,demonstrating favorable binding activity and therapeutic potential.
10.Comparison of short-term outcomes and 5-year overall survival between robotic and laparoscopic gastrectomy for gastric cancer
Li ZHANG ; Mingzhi CAI ; Bin LI ; Gang MA ; Xuejing ZHENG ; Ye TIAN ; Xuejun WANG ; Yong LIU ; Han LIANG
Chinese Journal of General Surgery 2025;40(8):601-606
Objective:To compare the short-term clinical outcomes of robotic gastrectomy (RG) and laparoscopic gastrectomy (LG) in treating gastric cancer, as well as the overall 5-year's survival rate.Methods:In this retrospective cohort study, 874 patients undergoing minimally invasive gastrectomy were recruited from Jan 2016 to Jan 2020 (LG: n=719; RG: n=155). A one-to-one propensity score matching analysis was applied to minimize the selection bias due to confounding factors, yielding 150 patients in each of the RG and LG groups. After matching, the short-term outcomes and 5-year overall survival were compared in the two groups. Results:The propensity score matching cohort analysis showed a similar 5-year overall survival between RG and LG groups ( P=0.235). Concerning the short-term outcomes, the RG compared to LG resulted in lower blood loss ( P=0.011), lower postoperative complications ( P=0.001), less postoperative pain ( P=0.014), earlier initiation of soft diet ( P=0.010), shorter hospital stay ( P=0.011), but higher hospitalization expenses ( P=0.004). Conclusions:RG had a similar overall survival outcome compared to LG while offering better safety and efficacy in terms of less blood loss, lower surgery complications, faster recovery, and less postoperative pain. Higher hospital expenses were the main disadvantage of RG that could limit its application.

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