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Mechanism and intervention of mucosal immune regulation based on "lung and large intestine being interior-exteriorly related" theory of traditional Chinese medicine
LOU Zhaohuan,ZHAO Huajun,LYU Guiyuan
J Zhejiang Univ (Med Sci), 2020, 49(6): 665-678.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.01
Abstract( 31 )   HTML( 4 )     PDF(1138KB)( 33 )

The "lung and large intestine being interior-exteriorly related" is one of the classical theories in traditional Chinese medicine, which indicates a close correlation between the lung and large intestine in physiology and pathology, and plays a pivotal role in guiding the treatment of the lung and bowel diseases. Modern medicine has revealed some connections between the lung and large intestine in tissue origin and mucosal immunity, and preliminarily illuminated the material basis and possible regulatory mechanism of the theory. Recently, this theory has been applied to guide the treatment of refractory lung and intestine diseases such as COVID-19 and ulcerative colitis and has obtained reliable efficacy. Existing research results show that the anatomical homogeneity of lung and large intestine promotes the correlation between lung-bowel mucosal immunity, and mucosal immunity and migration and homing of innate lymphocytes are one of the physiological and pathological mechanisms for lung and large intestine to share. Under the guidance of this theory, Chinese medicines with heat-clearing and detoxifying or tonic effects are commonly used in the treatment of the lung and intestinal diseases by regulating lung-bowel mucosal immunity and they can be candidate drugs to treat lung/intestinal diseases simultaneously. However, the existing studies on immune regulation are mainly focused on the expression levels of sIgA and cytokines, as well as the changes in the number of immune cells such as innate lymphocytes and B lymphocytes. While the following aspects need further investigation: the airway/intestinal mucous hypersecretion, the functional changes of pulmonary and intestinal mucosal barrier immune cells, the dynamic process of lung/intestinal mucosal immune interaction, the intervention effect of local pulmonary/intestinal microecology, the correlation and biological basis between the heat-clearing and detoxifying effect and the tonic effect, and its regulation of pulmonary/intestinal mucosal immunity. In this paper, we try to analyze the internal relationship between lung and intestine related diseases from the point of view of the common mucosal immune system of lung and intestine, and summarize the characteristics and rules of traditional Chinese medicine compound and its active ingredients, which have regulatory effect on lung and intestine mucosal immune system, so as to further explain the theoretical connotation of "lung and large intestine being interior-exteriorly related" and provide reference for the research and development of drugs for related diseases.

Astragaloside Ⅳ inhibits inflammation after cerebral ischemia in rats through promoting microglia/macrophage M2 polarization
ZHENG Xintian,GAN Haiyan,LI Lin,HU Xiaowei,FANG Yan,CHU Lisheng
J Zhejiang Univ (Med Sci), 2020, 49(6): 679-686.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.02
Abstract( 20 )   HTML( 3 )     PDF(1244KB)( 14 )

Objective: To investigate the effects of astragaloside Ⅳ (AS-Ⅳ) on microglia/macrophage M1/M2 polarization and inflammatory response after cerebral ischemia in rats. Methods: Forty eight male SD rats were randomly divided into sham operation control group, model control group and AS-Ⅳ group with 16 rats in each. Focal cerebral ischemia model was induced by occlusion of the right middle cerebral artery (MCAO) using the intraluminal filament. After ischemia induced, the rats in AS-Ⅳ group were intraperitoneally injected with 40 mg/kg AS-Ⅳ once a day for 3 days. The neurological functions were evaluated by the modified neurological severity score (mNSS) and the corner test on d1 and d3 after modelling. The infarct volume was measured by 2, 3, 5-triphenyl tetrazolium chloride (TTC) staining on d3 after ischemia. The expression of M1 microglia/macrophage markers CD86, inducible nitric oxide synthase (iNOS) and pro-inflammatory factors TNF-α, IL-1β, IL-6, M2 microglia/macrophages markers CD206, arginase-1 (Arg-1), chitinase-like protein (YM1/2) and anti-inflammatory factors interleukin-10 (IL-10) and transforming growth factor beta (TGF-β) was detected by real-time RT-PCR. The expression of CD16/32/Iba1 and CD206/Iba1 was determined by double labeling immunefluorescence method in the peripheral area of cerebral ischemia. Results: Compared with model control group, AS-Ⅳ treatment improved neurological function recovery and reduced infarct volume after ischemia (P < 0.05 or P < 0.01). The qRT-PCR results showed that AS-Ⅳ treatment down-regulated the expression of CD86, iNOS, TNF-α, IL-1β, IL-6 mRNA (all P < 0.01), and up-regulated the expression of CD206, Arg-1, YM1/2, IL-10 and TGF-β mRNA (all P < 0.01). Furthermore, the results of immunefluorescence labeling showed that AS-Ⅳ treatment reduced the number of CD16/32+/Iba1+ cells (P < 0.05) and increased the number of CD206+/Iba1+ cells (P < 0.01) after cerebral ischemia. Conclusion: The findings suggest that AS-Ⅳ ameliorates brain injury after cerebral ischemia in rats, which may be related to inhibiting inflammation through promoting the polarization of the microglia/macrophage from M1 to M2 phenotype in the ischemic brain.

Chinese medicine Buyang Huanwu decoction promotes neurogenesis and angiogenesis in ischemic stroke rats by upregulating miR-199a-5p expression
ZHUGE Lujie,FANG Yan,JIN Huaqian,LI Lin,YANG Yan,HU Xiaowei,CHU Lisheng
J Zhejiang Univ (Med Sci), 2020, 49(6): 687-696.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.03
Abstract( 12 )   HTML( 2 )     PDF(1331KB)( 10 )

Objective: To investigate the mechanism of Chinese medicine Buyang Huanwu decoction (BYHWD) promoting neurogenesis and angiogenesis in ischemic stroke rats. Methods: Male SD rats were randomly divided into sham operation group, model group, BYHWD group, antagonist group and antagonist control group with 14 rats in each. Focal cerebral ischemia was induced by occlusion of the right middle cerebral artery for 90 min with intraluminal filament and reperfusion for 14 d in all groups except sham operation group. BYHWD (13 g/kg) was administrated by gastrogavage in BYHWD group, antagonist group and antagonist control group at 24 h after modeling respectively, and BrdU (50 mg/kg) was injected intraperitoneally in all groups once a day for 14 consecutive days. miR-199a-5p antagomir or NC (10 nmol) was injected into the lateral ventricle at d5 after ischemia in antagonist and antagonist control groups, respectively. The neurological deficits were evaluated by the modified neurological severity score (mNSS) and the corner test, and the infract volume was measured by toluidine blue staining. Neurogenesis and angiogenesis were detected by immunofluorescence double labeling method. The expression level of miR-199a-5p was tested by real-time RT-PCR, and the protein expressions of vascular endothelial growth factor (VEGF) and brain-derived neurotrophic factor (BDNF) were determined by Western blotting. Results: BYHWD treatment significantly promoted the recovery of neurological function (P < 0.05 or P < 0.01), reduced the infarct volume (P < 0.05), increased the number of BrdU+/DCX+, BrdU+/NeuN+ and BrdU+/vWF+ cells (all P < 0.01), upregulated the expression of miR-199a-5p (P < 0.01), and increased the protein expression of VEGF and BDNF at d14 after cerebral ischemia (all P < 0.05). The above effects were reversed by intracerebroventricular injection of miR-199a-5p antagomir. Conclusion: Buyang Huanwu decoction promotes neurogenesis and angiogenesis in rats with cerebral ischemia, which may be related to increased protein expression of VEGF and BDNF through upregulating miR-199a-5p.

Effect of Wubi Shanyao pills on sexual function in mice with kidney-yang-deficiency induced by hydrocortisone
CHEN Qingqing,SHAN Chaowen,SU Jie,CHEN Wei,ZHU Jiaming,CHEN Suhong,LYU Guiyuan
J Zhejiang Univ (Med Sci), 2020, 49(6): 697-704.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.04
Abstract( 20 )   HTML( 2 )     PDF(1207KB)( 8 )

Objective: To investigate the effect of Chinese medicine Wubi Shanyao pills on sexual function of kidney-yang-deficiency mice induced by hydrocortisone. Methods: Male Kunming mice were injected with hydrocortisone for 10 days to prepare the kidney-yang-deficiency model, and administrated with Wubi Shanyao pills (0.91, 1.82, 2.73 g/kg) for 9 weeks. The general behaviors of mice (autonomous activity, grasping power) were observed; sexual behaviors (capture, straddle, ejaculation frequency and incubation period) of mice were detected by mating experiment. The serum levels of cortisol, adrenocorticotropic hormone (ACTH), luteinizing hormone (LH), follicle stimulating hormone (FSH), estradiol (E2) and testosterone were measured by ELIZA. The organ indexes of kidney, epididymis, testis and seminal vesicle were calculated. The histomorphology of testis was observed by HE staining. The apoptosis of testis cells was detected by TUNEL; and the expression level of proliferating cell nuclear antigen (PCNA) in testis was detected by immunohistochemistry. Results: Wubi Shanyao pills increased the number of independent activities, grasping power, capture frequency of model mice and shortened the capture latency (all P < 0.01); and also increased the serum levels of testosterone, cortisol, ACTH, decreased the serum levels of FSH (P < 0.05 or P < 0.01); increased testis index (P < 0.05), improved the pathological changes of testis, inhibited cell apoptosis, and promoted PCNA protein expression in testis. Conclusion: Wubi Shanyao pills can improve the sexual function of mice with kidney-yang-deficiency induced by hydrocortisone, which may be related to regulating the hypothalamus-pituitary-adrenal axis (HPA axis), promoting the proliferation of testicular cells, and inhibiting cell apoptosis.

Protective effect of iridoid glycosides of radix scrophulariae on endoplasmic reticulum stress induced by oxygen-glucose deprivation and reperfusion in vitro model
YE Jiayi,GONG Hengpei,WANG Lingfeng,HUANG Zhen,QIU Fengmei,ZHONG Xiaoming
J Zhejiang Univ (Med Sci), 2020, 49(6): 705-713.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.05
Abstract( 16 )   HTML( 1 )     PDF(1295KB)( 6 )

Objective: To investigate the regulatory effect of iridoid glycoside of radix scrophulariae (IGRS) on endoplasmic reticulum stress induced by oxygen-glucose deprivation and reperfusion in vitro model. Methods: Rat pheochromocytoma PC12 cells were pretreated with IGRS (50, 100, 200 μg/mL) for 24h, and the in vitro model of oxygen-glucose deprivation/reoxygenation (OGD/R) was applied. The cell viability was determined by MTT and lactate dehydrogenase (LDH) assay. The apoptotic rate was detected by flow cytometry. The expression of B-cell lymphoma-2 (Bcl-2), Bcl-2 related X protein (Bax), C/EBP homologous protein (CHOP), caspase-12 protein, and glucose-regulated protein-78(GRP78)were detected by Western blotting. The mRNA expression levels of sarco/endoplasmic reticulum Ca2+-ATPase2 (SERCA2), 1, 4, 5-triphosphate inositol receptor 1 (IP3R1), and ryanodine receptor 2 (RyR2)were detected by real-time RT-PCR. Free Ca2+ concentration [Ca2+]i was determined by using laser scanning confocal microscopy. Results: The damage caused by OGD/R to PC12 cells was significantly reduced by IGRS, with significant effect on increasing survival rate and reducing LDH release (all P < 0.01). The expression of GRP78, CHOP, Bax, and caspase-12 were down-regulated (all P < 0.01), and the expression of Bcl-2 and Bcl-2/Bax ratio was up-regulated (all P < 0.01); IGRS increased the expression of SERCA2 mRNA in PC12 cells after OGD/R injury (P < 0.01), decreased [Ca2+]i and down-regulated the expression of RyR2 mRNA and IP3R1 mRNA. Conclusion: IGRS has neuroprotective effect, which may alleviate cerebral ischemia-reperfusion injury by regulating SERCA2, maintaining calcium balance, and inhibiting endoplasmic reticulum stress-mediated apoptosis.

Study on the mechanism of Flos Puerariae and Semen Hoveniae in treatment of alcoholic liver injury based on network pharmacology and molecular docking
WANG Yanan,YAN Xiaoming,ZHANG Qingyu,SONG Aihua,HAN Fei
J Zhejiang Univ (Med Sci), 2020, 49(6): 714-724.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.06
Abstract( 22 )   HTML( 1 )     PDF(1337KB)( 9 )

Objective: To explore the mechanism of Flos Puerariae and Semen Hoveniae in treatment of alcoholic liver injury (ALI) based on network pharmacology and molecular docking. Methods: The information of chemical constituents and targets of Flos Puerariae and Semen Hoveniae was collected from TCMSP and Swiss databases, and the threshold values of oral bioavailability (OB) ≥ 30%, drug likeness (DL) ≥0.18 were used to screen the potential active compounds. The GeneCard and DrugBank databases were used to obtain the targets corresponding to ALI. The common targets were queried using Venn Diagram, and the network of PPI and Gene Ontology (GO) functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were performed through DAVID and Reactome database. Autodock Vina software was used for molecular docking of potential ingredients and key targets. Results: A total of 21 potential active compounds and 431 therapeutic targets were gathered in Flos Puerariae and Semen Hoveniae, which involved 273 biological functions, 90 KEGG pathways and 362 Reactome pathways. The GO functions involved protein binding, ATP binding, etc.; the KEGG pathways mainly included PI3K-Akt signaling pathway and TNF signaling pathway; the Reactome pathways contained signal transduction and immune system, etc. The results of molecular docking showed that 21 potential active ingredients had good affinity with the core targets Akt1, TP53 and IL-6. Conclusion: The network pharmacology and molecular docking analysis demonstrate the synergetic effect of Flos Puerariae and Semen Hoveniae with multi-compounds, multi-targets and multi-pathways in the treatment of ALI; and also predict the possible medicinal substance, key targets and pathways, which provides clues for the new drug development and mechanism research.

Expression of tumor-associated vascular insulin receptor in colorectal cancer and its relationship with tumor pathological features
YANG Zeran,ZHANG Xin,MA Jie,JIN Li,HE Xujun
J Zhejiang Univ (Med Sci), 2020, 49(6): 725-731.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.07
Abstract( 17 )   HTML( 1 )     PDF(1156KB)( 5 )

Objective: To study the expression of tumor associated vascular insulin receptor (TVIR) in colorectal cancer with or without metabolic syndrome (MS) and its relationship with the pathological features of colorectal cancer. Methods: The expression of TVIR in 220 colorectal cancer specimens was detected by tissue microarray and immunohistochemistry. The relationships between the expression of TVIR and the pathological features (pathological subtypes, histological grade, invasion depth, lymph node metastasis and TNM stage) of colorectal cancer with/without MS were analyzed. Results: The insulin receptor expression was observed in colorectal cancer tissue or border area between cancer and normal tissue, but not in normal intestinal tissue. The high-expression rates of TVIR in MS group was remarkably lower than that of non-MS group (21.6%vs. 41.0%, P < 0.05). TVIR high expression was significantly associated with tumor deep invasion, lymph node metastasis and high TNM stage (P < 0.05 or P < 0.01). Univariate logistic regression analysis showed high-expression of TVIR to be significantly associated with risk of deep invasion (OR=2.21, 95% CI: 1.10-4.44, P < 0.05), lymph node metastasis (OR=2.21, 95% CI: 1.26-3.86, P < 0.01) and high TNM stage (OR=2.08, 95% CI: 1.19-3.63, P < 0.05). Such associations can be observed in patients without MS, but not in patients with MS. Conclusions: High-expression of TVIR is associated with aggressive pathological features such as invasion, lymph node metastasis and high TNM stage of colorectal cancer, especially for those patients without MS. TVIR could be a useful biological marker for prognosis of colorectal cancer.

Master genes and co-expression network analysis in peripheral blood mononuclear cells of patients with gram-positive and gram-negative sepsis
LI Lu,FANG Junjun,LI Zhitao,SHEN Leixing,WANG Guobin,FU Shuiqiao
J Zhejiang Univ (Med Sci), 2020, 49(6): 732-742.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.08
Abstract( 15 )   HTML( 2 )     PDF(1532KB)( 7 )

Objective: To investigate the functional pathways enriched and differentially expressed genes (DEGs) in peripheral blood mononuclear cells (PBMCs) of patients with gram-positive and gram-negative sepsis. Methods: Dataset GSE9960 obtained from NCBI GEO database containing PBMC samples from 16 non-infectious systematic inflammatory response syndrome (SIRS) patients, 17 gram-positive septic patients and 18 gram-negative septic patients were included in the study. Functional pathway annotations were conducted by gene set enrichment analysis and weighted gene co-expression network analysis. DEGs were filtered and master DEGs were then validated in PBMCs of gram-positive septic, gram-negative septic and non-infectious SIRS patients. Results: The enriched gene sets in gram-positive sepsis and gram-negative sepsis were significantly different. The results indicated the opposite co-expression networks in SIRS and gram-negative sepsis, and the entirely different co-expression networks in gram-positive and gram-negative sepsis. Furthermore, we validated that TYMS was up-regulated in gram-positive sepsis (P < 0.05), CD3D was down-regulated in gram-negative sepsis (P < 0.01), while IRAK3 was up-regulated in gram-negative sepsis (P < 0.05). Conclusion: The results indicate that there are differences in the mechanism and pathogenesis of gram-positive and gram-negative sepsis, which may provide potential markers for sepsis diagnosis and empirical antimicrobial therapy.

Differentially expressed inflammatory proteins in acute gouty arthritis based on protein chip
SUN Guanghan,LIU Jian,WAN Lei,LIU Wei,LONG Yan,BAO Bingxi,ZHANG Ying
J Zhejiang Univ (Med Sci), 2020, 49(6): 743-749.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.09
Abstract( 13 )   HTML( 1 )     PDF(1196KB)( 6 )

Objective: To detect the differentially expressed inflammatory proteins in acute gouty arthritis (AGA) with protein chip. Methods: The Raybiotech cytokine antibody chip was used to screen the proteomic expression in serum samples of 10 AGA patients and 10 healthy individuals. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were applied to determine the biological function annotation of differentially expressed proteins and the enrichment of signal pathways. ELISA method was used to verify the differential protein expression in 60 AGA patients and 60 healthy subjects. The ROC curve was employed to evaluate the diagnostic value of differential proteins in AGA patients. Results: According to|log2FC|>log2 1.2 and corrected P < 0.01, 4 most differentially expressed proteins in AGA patients were identified, including tumor necrosis factor receptor super family members Ⅱ (TNF RⅡ), macrophage inflammatory protein 1β (MIP-1β), interleukin-8 (IL-8), and granulocyte-macrophage colony stimulating factor (GM-CSF). GO and KEGG enrichment analysis showed that the differentially expressed proteins were related to inflammation, metabolism and cytokine pathways. The ELISA results showed that serum levels of differentially expressed proteins were significantly different between AGA patients and healthy subjects(all P < 0.01). ROC curve analysis showed that the areas under the curve (AUCs) of GM-CSF, IL-8, MIP-1β and TNF RⅡ for predicting AGA were 0.657 (95% CI: 0.560-0.760, sensitivity: 68.33%, specificity: 50.00%), 0.994 (95% CI: 0.980-1.000, sensitivity: 100.00%, specificity: 61.67%), 0.980 (95% CI: 0.712-0.985, sensitivity: 95.00%, specificity: 98.33%) and 0.965 (95% CI: 0.928-1.000, sensitivity: 100.00%, specificity: 10.00%), respectively. Conclusion: Proteomics can be applied to identify the biomarkers of AGA, which may be used for risk prediction and diagnosis of AGA patients.

Sirt3 gene knockout protects mice from Alzheimer's disease through activating autophagy
SHU Min,ZHANG Wenzhe,JIN Xiangbo,ZENG Linghui,XIANG Yingchun
J Zhejiang Univ (Med Sci), 2020, 49(6): 750-757.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.10
Abstract( 19 )   HTML( 0 )     PDF(1162KB)( 9 )

Objective: To investigate the protective effect of Sirt3 gene knockout on Alzheimer's disease (AD) in mice. Methods: The animal model of AD was established by intraperitoneal injection of D-galactose and brain-localized injection of amyloid β-protein (Aβ)1-40 in wild type C57BL/6 mice and Sirt3 gene knockout mice. Morris water maze, Y maze and tail suspension test were used to assess the cognitive function and anxiety-like behaviors in mice. Aβ deposition in the hippocampus was detected by immunofluorescent staining. Western blotting analysis was conducted to detect the expression of related proteins in the brain. Mouse cortical primary neurons were cultured and AD cell model was established. MTT assay was used to detect cell viability after modeling. Results: Behavioral results showed that cognitive deficits were found in wide type mice after induction of AD as its prolonged escape latency (P < 0.05) and decreased crossing number of platform and target zone duration (all P < 0.05); while the knockout of Sirt3 alleviated cognitive deficit induced by AD (all P < 0.05). Aβ immunofluorescence staining showed that the deposition of Aβ in the hippocampal region and expression of cleaved caspase 3 in the brain in Sirt3 knockout mice was reduced compared with that of wild type mice (all P < 0.05). The expression of LC3-Ⅱ and P62 increased after AD was induced in wild type mice, while the autophagy in Sirt3 knockout mice was activated as the increase expression of LC3-Ⅱ and decrease expression of P62 (all P < 0.05). In the AD cell model, the results of MTT assay were consistent with the animal experiments, and the protective effect of Sirt3 knockdown was eliminated after the treatment of the autophagy inhibitor chloroquine (all P < 0.05). Conclusion: The knockdown of Sirt3 shows a protective effect on AD induced by D-galactose and Aβ1-40 in mice, which may be related to its function of activating autophagy.

IL-17A activates mouse lung fibroblasts through promoting chemokine CXCL12 secretion
WANG Huaying,LYU Jiapei,CHEN Liping,YU Wanjun
J Zhejiang Univ (Med Sci), 2020, 49(6): 758-764.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.11
Abstract( 18 )   HTML( 1 )     PDF(1252KB)( 8 )

Objective: To investigate the role of IL-17A in promoting the activation of lung fibroblasts and the secretion of chemokine CXCL12, and to explore the possible mechanism. Methods: Lung tissues of BALB/c mice were collected after intraperitoneal injection of recombinant mouse IL-17A (rmIL-17A). Real-time RT-PCR and Western blotting were used to detect the expression levels of α-smooth muscle actin (α-SMA) and collagen I in lung tissues, and immunohistochemical staining and real-time RT-PCR were used to determine the expression of CXCL12. Normal mouse primary lung fibroblasts were isolated and cultured, and identified by immunofluorescence staining with optical microscopy. Cells and supernatant of culture medium were collected after stimulation with rmIL-17A at different concentrations. mRNA levels of α-SMA, collagen I, and CXCL12 in the cells were determined by real-time RT-PCR, and the levels of collagen I and CXCL12 in the supernatant of culture medium were determined by ELISA. Results: The mRNA and protein levels of α-SMA and collagen I in the lung tissue of mice injected with rmIL-17A were significantly increased compared with the control group (all P < 0.01). The mRNA levels of α-SMA, collagen I and CXCL12 in mice primary lung fibroblasts were increased after stimulation of rmIL-17A at different concentrations (all P < 0.01), and the concentration of collagen Ⅰ and CXCL12 in the supernatants of culture medium were also increased in a dose-dependent manner (all P < 0.01). Conclusions: IL-17A can promote the activation of lung fibroblasts and translation into myofibroblast. The secretion of collagen is increased, which promote the deposition of extracullular matrix, and leads to the occurrence and development of lung fibrosis. CXCL12, a chemokine secreted by activated fibroblasts, may be involved in this process.

Preparation of an anti-cotinine monoclonal antibody and its application in immunological detection
LEI Yajing,ZHOU Lifang,WANG Anxing,CHEN Shuqing
J Zhejiang Univ (Med Sci), 2020, 49(6): 765-771.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.12
Abstract( 15 )   HTML( 1 )     PDF(1135KB)( 6 )

Objective: To prepare monoclonal antibody against cotinine (COT) and to establish immunoassay for detecting COT in human urinary samples. Methods: BALB/c mice were immunized with synthesized cotinine-bovine serum albumin (COT-BSA) to screen monoclonal antibody with technique of cell fusion. The monoclonal antibody was used for the indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and colloidal gold immunochromatographic strip assay for the detection of COT in human urine. Results: The monoclonal antibody against COT was identified by ic-ELISA with a 50%inhibitive concentration (IC50) value of 21 ng/mL; and it was also identified by colloidal gold immunochromatographic strip assay with a cut-off value of 100 ng/mL. For ic-ELISA, the range of detection was 0-100 ng/mL with a minimal limit of 0.1 ng/mL; the recovery of assay was 99.41%-117.98%, and the intra-assay and inter-assay coefficient variations were not higher than 15.31%and 15.07%, respectively. For colloidal gold immunochromatographic strip assay, the accuracy of stability and repeatability both were 100%. Conclusion: The ic-ELISA and colloidal gold immunochromatographic strip assay using the prepared monoclonal antibody against COT have been proved to be reliable for the rapid detection of COT in human urines, which may be used for monitoring of environmental tobacco smoke.

Tissue factors and venous thromboembolism in cancer patients
ZHU Huiqi,YING Kejing
J Zhejiang Univ (Med Sci), 2020, 49(6): 772-778.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.13
Abstract( 19 )   HTML( 3 )     PDF(1030KB)( 17 )

Malignant tumor is one of the important acquired risk factors of venous thromboembolism (VTE). As the transmembrane receptor of coagulation factor Ⅶ and activated coagulation factor Ⅶa in vivo, tissue factor is the main initiator of exogenous coagulation. Tissue factor positive particles expressed and released by tumor cells enter the circulation and mediate thrombosis in patients with surgical treatment and distant tumor metastasis; the enhanced procoagulant activity of tissue factor after chemotherapy makes many cancer patients more likely to develop thromboembolic disease. Tissue factors can also be used to predict the risk of VTE in patients with pancreatic cancer, colorectal cancer and ovarian cancer.This article summarizes the role of tissue factor in VTE of cancer patients at different treatment stages, and further clarifies the relationship between tissue factor and the risk of VTE in cancer patients.

Research progress on biomarkers for endometriosis based on lipidomics
LIN Cuicui,CHEN Zhengyun,WANG Chunyan,XI Yongmei
J Zhejiang Univ (Med Sci), 2020, 49(6): 779-784.   https://doi.org/10.3785/j.issn.1008-9292.2020.12.14
Abstract( 19 )   HTML( 1 )     PDF(1022KB)( 9 )

The pathogenesis of endometriosis is not well understood at the moment, and the lack of effective biomarkers often leads to delayed diagnosis of the disease. Lipidomics provides a new approach for the diagnosis and prediction of endometriosis. Sphingomyelin, phosphatidylcholine and phosphatidylserine in peripheral blood, endometrial fluid, peritoneal fluid and follicular fluid have good diagnostic value for endometriosis and disease classification; the lipid metabolites in the eutopic endometrium tissue are expected to be biomarkers of early endometriosis; and the lipid metabolites in peripheral blood are also of great value for predicting endometriosis-related infertility. The development of lipidomics technique will further advance the progress on the pathogenesis, prediction, diagnosis and treatment of endometriosis.

14 articles