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Objective To investigate the impact of quercetin(Que)on postherpetic neuralgia(PHN)and chemokine ligand 3(CCL3,namely MIP-1α)/C-C chemokine receptor 1(CCR1)/C-C chemokine receptor 5(CCR5)signaling pathway in rats.Methods Sixty rats were divided into the control group(Con),the PHN group(model group),the L-Que(30 mg/kg)group,the M-Que(60 mg/kg)group,the H-Que(120 mg/kg)group and the H-Que+pathway activator MIP-1α(120 mg/kg Que+0.4 mg/kg recombinant MIP-1α)group.The mechanical paw withdrawal threshold(PWT)and thermal pain threshold(TWL)of rats were detected in each group.The kit was used to detect adenosine,Adenine ribonucleotide(AMP),adenosine diphosphate(ADP)and tumor necrosis factor in spinal dorsal horn samples-α(TNF-α),and interleukin-1 β(IL-1 β)levels in spinal dorsal horn samples.HE staining was applied to observe the pathological sections of spinal dorsal horn.Immunofluorescence staining was used to detect the activation of microglia in spinal dorsal horn.Western blot assay was applied to detect MIP-1α/CCR1/CCR5 signaling pathway protein expression.Results In the PHN group,the dorsal horn of the spinal cord was ruptured,the arrangement of nerve bundles was disordered,and inflammatory cell infiltration,edema,and slight atrophy of neurons appeared.Compared with the Con group,the PWT value,adenosine,AMP and ADP levels were obviously decreased in the PHN group(P<0.05),and TWL value,TNF-α,IL-1β levels,the number of Iba1-positive microglia,MIP-1α,CCR1 and CCR5 protein levels were obviously increased(P<0.05).After treatment with Que,the disordered arrangement of nerve bundles was improved,the infiltration of inflammatory cells was reduced,and the phenomenon of neuronal atrophy disappeared.Compared with the PHN group,the PWT value,adenosine,AMP and ADP levels were obviously increased in the L-Que group,the M-Que group and the H-Que group(P<0.05).TWL value,TNF-αand IL-1β levels,the number of Iba1-positive microglia,and MIP-1α,CCR1 and CCR5 protein levels were obviously decreased(P<0.05).The effect of Que was dose dependent.Compared with the H-Que group,PWT value,adenosine,AMP and ADP levels were obviously decreased in the H-Que+MIP-1α group(P<0.05),and TWL value,TNF-α,IL-1β levels,the number of Iba1 positive microglia,MIP-1α,CCR1 and CCR5 protein levels were obviously increased(P<0.05).Conclusion Que may reduce the inflammatory response in rats by inhibiting the MIP-1α/CCR1/CCR5 signaling pathway,thereby reducing PHN.
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Objective To investigate the levels of soluble tyrosine kinase-1(sFlt-1)and chemokine C-C ligand 3(CCL3)in serum of patients with coronary heart disease(CHD)and their correlation with the severity of the disease.Methods A total of 230 elderly CHD patients admitted to the De-partment of Cardiovascular Medicine of Xinxiang Central Hospital from November 2020 to No-vember 2022 were collected as the study subjects(CHD group),and according to their Gensini score,they were divided into mild(n=89),moderate(n=95),and severe(n=46)CHD sub-groups.Another 230 healthy individuals who taking physical examination during the same period served as the control group.ELISA was applied to measure serum levels of sFlt-1 and CCL3.ROC curve was plotted to analyze the diagnostic values of serum sFlt-1 and CCL3 levels for CHD.Pear-son correlation analysis was employed to analyze the relationship between serum sFlt-1 and CCL3 levels and the CHD severity.Results The serum levels of sFlt-1 and CCL3 were obviously higher in the CHD group than the control group(121.71±29.80 ng/L vs 98.70±17.57 ng/L,18.22± 5.41 ng/L vs 13.68±3.89 ng/L,P<0.01).ROC curve analysis showed that the AUC value of the two indicators combined together was significantly greater than that of them alone in diagnosis of CHD(0.886 vs 0.791,0.775,P<0.01).The serum levels of sFlt-1 and CCL3 were increased along with the severity of the disease and Gensini score when the levels and the score were compared among the mild,moderate and severe subgroups(P<0.05).Pearson correlation analysis indicated that the serum levels of sFlt-1 and CCL3 were positively correlated with the Gensini score(r=0.420,r=0.479,P<0.01).Conclusion The levels of serum sFlt-1 and CCL3 are obviously ele-vated in CHD patients,and closely associated with the severity of coronary lesions.
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PURPOSE: The purpose of this study was to identify novel plasma biomarkers for distinguishing nasopharyngeal carcinoma (NPC) patients from healthy individuals who have positive Epstein-Barr virus (EBV) viral capsid antigen (VCA-IgA). MATERIALS AND METHODS: One hundred seventy-four plasma cytokines were analyzed by a Cytokine Array in eight healthy individuals with positive EBV VCA-IgA and eight patients with NPC. Real-time polymerase chain reaction, Western blotting, enzyme-linked immunosorbent assay (ELISA), and immunohistochemistry were employed to detect the expression levels of macrophage migration inhibitory factor (MIF) and CC chemokine ligand 3 (CCL3) in NPC cell lines and tumor tissues. Plasma MIF and CCL3 were measured by ELISA in 138 NPC patients, 127 EBV VCA-IgA negative (VN) and 100 EBV VCA-IgA positive healthy donors (VP). Plasma EBV VCA-IgA was determined by immunoenzymatic techniques. RESULTS: Thirty-four of the 174 cytokines varied significantly between the VP and NPC group. Plasma MIF and CCL3 were significantly elevated in NPC patients compared with VN and VP. Combination of MIF and CCL3 could be used for the differential diagnosis of NPC from VN cohort (area under the curve [AUC], 0.913; sensitivity, 90.00%; specificity, 80.30%), and combination of MIF, CCL3, and VCA-IgA could be used for the differential diagnosis of NPC from VP cohort (AUC, 0.920; sensitivity, 90.00%; specificity, 84.00%), from (VN+VP) cohort (AUC, 0.961; sensitivity, 90.00%; specificity, 92.00%). Overexpressions of MIF and CCL3 were observed in NPC plasma, NPC cell lines and NPC tissues. CONCLUSION: Plasma MIF, CCL3, and VCA-IgA combination significantly improves the diagnostic specificity of NPC in high-risk individuals.
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Humans , Biomarkers , Blotting, Western , Capsid , Cell Line , Chemokine CCL3 , Cohort Studies , Cytokines , Diagnosis , Diagnosis, Differential , Enzyme-Linked Immunosorbent Assay , Herpesvirus 4, Human , Immunoglobulin A , Immunohistochemistry , Macrophages , Plasma , Real-Time Polymerase Chain Reaction , Sensitivity and Specificity , Tissue DonorsABSTRACT
AIM:To explore the regulatory effect of chemokine CCL 3 on exosome secretion from human bone marrow mesenchymal stem cells(hBMSCs).METHODS: hBMSCs were stimulated with chemokine CCL 3 at different concentrations in vitro.The proliferation of hBMSCs was measured by CCK-8 assay and viable cell counting.Exosome se-cretion from hBMSCs was qualitatively analyzed by transmission electron microscope(TEM)and flow cytometry, and the quantitative analysis was carried out by flow cytometry and nanoparticle tracking analysis(NTA).RESULTS:Compared with control group,the viability of the hBMSCs detected by CCK-8 assay was increased when hBMSCs were treated with CCL3(P<0.05).The results of viable cell counting demonstrated that the number of hBMSCs was raised in CCL 3 group in a dose-dependent manner(P<0.05).The results of flow cytometry showed that hBMSCs expressed 3 CCL3-related spe-cific receptors,CCR1,CCR5 and CCR9.Compared with control group,the fluorescence intensity of CCR9 in CCL3 group was obviously enhanced.However,no significant difference of fluorescence intensity for CCR 5 and CCR1 was observed be-tween the 2 groups.The results of NTA demonstrated that the secretion capacity of CCL 3-induced hBMSCs was far less than that in control group(P<0.05).However, the microvesicles larger than 100 nm in CCL3 groups were increased(P<0.05).The above results indicated that the higher concentration of CCL 3 induced the lower secretion of exosomes.In addi-tion,the results of flow cytometry demonstrated that CCL 3-induced hBMSCs showed lower quantity of CD 9 +exosomes than those in control group(P<0.01).CONCLUSION:CCL3 promotes the proliferation of hBMSCs but depresses the secre-tion of exosomes in a dose-dependent manner.CCL3 affects the size distribution of exosomes and increases the number of nonfunctional microvesicles of larger than 100 nm in size.CCL3 induces the expression of CCR9 in hBMSCs.
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Objective To investigate the levels of serum macrophage inflammatory protein-1α (MIP-1α),inflammatory factors and pulnonary function in patients with asthma,and the correlation between thenm.Methods Eighty patients with asthma were selected who were treated in our hospital from March 2015 to August 2016,and the data of 80 healthy adults were selected as control.The differences of serum MIP-1α,inflammatory factors,imnmunoglobulin and lung function were observed in two groups.The correlation between MIP-1α and inflammatory factors,lung function and immunoglobulin were analyzed.Results The levels of MIP-1α,interleukin (IL)-6,and tumor necrosis factor-α (TNF-α) in patients with asthma were significantly higher than those in control group (t =-207.04,-33.209,-55.132,P < 0.01);The levels of forced vital capacity (FVC),the forced expiratory volume in one second (FEV1),FEV1/FVC,maximal mid expiratory flow (MMEF),and peak expiratory flow (PEF) in the asthma group were significantly lower than those in the control group (t =17.100,39.154,22.791,25.391,19.356,P < 0.01).The IgG and IgM of asthma group were significantly lower than those of the control group (t =9.564,7.528,P < 0.01).The IgE level was significantly higher than that of the control group (t =-82.683,P <0.01).The level of MIP-1α was positively correlated with the levels of IL-6,TNF-0 and IgE,and negatively correlated with FVC,FEV1,FEV1/FVC,MMEF,PEF,IgG and IgM in patients with asthma.Conclusions The level of serum MIP-1 α in asthmatic patients is high,and is closely related to the inflammatory cytokines and lung function in patients with asthma.
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Objective This study aimed to investigate whether the copy numbers of the CCL3L1 (Chemokine C-C-Motif Ligand 3 Like Protein 1) gene were associated with susceptibility to ankylosing spondylitis (AS). Methods A total of 806 Chinese individuals including 405 AS patients and 401 healthy controls were enrolled. The CCL3L1 gene copy number was measured by a custom-by-design Multiplex AccuCopyTM Kit based on a multiplex fluorescence competitive polymerase chain reaction (PCR) principle, and 50 samples were randomly selected using the fluorescent quantitative PCR method to verify copy number. Main statistical method was t test, chi-square test and logistic regression model. Results There were no statistically significant differences between the case group and control group in age and gender ( t=1.77, P=0.076, χ2=1.14, P=0.289). The copy number of CCL3L1 gene ranged from 0 to 13 in both AS patients and the controls. After copy numbers were classified into 3 categories by 3, we did not find significant difference between the two groups ( χ2=0.591, P=0.669). And regression analyses also did not support the hypothesis that CCL3L1 gene copy number variation (CNV) could be an impact factor to the severity or function indexes of AS patients ( χ2=0.341, P=0.804 and χ2=0.472, P=0.774, respectively). Conclusion We suggest that the copy number of the CCL3L1 gene does not have a role in the susceptibility and the severity or function to AS.
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Objective To evaluate the changes in the expression of CC-chemokine ligand 3 (CCL3) and CC-chemokine receptor 5 (CCR5) in the spinal cord during hyperalgesia induced by remifentanil in rats with incisional pain.Methods Thirty-two male Sprague-Dawley rats,aged 2-3 months,weighing 240-260 g,were randomly divided into 4 groups (n=8 each) using a random number table:control group (group C),incisional pain group (group Ⅰ),remifentanil group (group R) and remifentanil+incisional pain group (group R+I).A 1-cm longitudinal incision was made in the plantar surface of the left hindpaw in anesthetized rats.While the model of incisional pain was established,remifentanil was infused for 60 min at 1 μg · kg-1 · min-1.At 24 h before infusion of remifentanil (baseline) and 2,6,24 and 48 h after the end of infusion,the mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) were measured.The rats were sacrificed after the last measurement of pain threshold,the lumbar segment (L4-6) of the spinal cord was removed for determination of CL3 and CCR5 mRNA expression (by real-time PCR) and CL3 and CCR5 expression (by Western blot).Results Compared with group C,the MWT was significantly decreased,the TWL was shortened,and the expression of CCL3 and CCR5 mRNA and protein was up-regulated in I,R and R+ I groups.Compared with I and R groups,the MWT was significantly dccreascd,the TWL was shortened,and the expression of CCL3 and CCR5 mRNA and protein was up-regulated in group R+I.Conclusion The mechanism by which remifentanil induces hyperalgesia is related to up-regulated expression of CCL3 and CCR5 in the spinal cord of rats with incisional pain.
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Objective To investigate the correlation between the human chemokine type 1 chemokine ligand 3 (CCL3L1) and chemokine ligand 4 (CCL4L) gene expression and the immune reconstitution of acquired immunodeficiency syndrome (AIDS) patients after antiretroviral therapy.Methods The gene copy numbers of CCL3L1 and CCL4L were detected by real time polymerase chain reaction in 217 AIDS patients before antiretroviral therapy.And the correlation between CCL3L1 and CCL4L gene copy numbers and the level of CD4+ and CD8+ T lymphocytes were analyzed.The changes of CD4+ and CD8+ T lymphocytes were defined as mean change value per month after 48 months treatment.The change rates of CD4+ and CD8+ T lymphocytes were defined as the logarithm of the ratio of the value after 48 month to that at baseline.Comparison between groups was conducted using analysis of variance.Results The median of gene copy numbers of CCL3L1 and CCL4L were 2 (range:0-8) and 3 (range:0-7),respectively.After antiviral treatment,there were significantly different changes of CD8+ T lymphocyte level (F=3.054,P<0.05) and change rate of CD4+/CD8+ (F=3.520,P<0.05) among groups of high (gene copy 4-8),median (gene copy 2-3) and low (gene copy 0-1) CCL3L1 gene copy numbers.The changes of CD8+ T lymphocyte levels (P=0.023) and change rates (P=0.038) in high and low CCL3L1 gene copy groups were both significantly different.There were significant differents changes rate of ratio of CD4+/CD8+ T lymphocyte among high and median (P=0.010),high and low CCL3L1 gene copy numbers (F=4.397,P<0.05).The significant difference of the change rates of CD4+/CD8+ were found between the gene copy 3 group and gene copy 4-7 group CCL4L (P=0.005) and between the gene copy 4-7 group and gene copy 0-2 group of CCL4L (P=0.030).The change ratio of CD4+/CD8+ T lymphocytes increased with the increase of copy numbers of CCL4L gene.Conclusions The gene expressions of CCL3L1 and CCL4L might be associated with the ability of immune reconstitution of AIDS patients after antiretroviral therapy.
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Objective To investigate the impact of methamphetamine (Meth) on the expressions of macrophage inflammatory protein (MIP)-1α ,MIP-1β ,interleukin (IL)-6 among human immunodeficiency virus(HIV)-infected patients .Methods The investigation was performed among 15 Meth-abuse and HIV-infected subjects (Meth + HIV ) ,15 non-Meth-abuse and HIV-infected subjects (non-Meth + HIV ) ,15 Meth-abuse and HIV-uninfected subjects (Meth) ,and 15 healthy subjects (HC) .CD4 + T lymphocyte counts in peripheral blood were detected by flow cytometry .The HIV viral loads in HIV-infected patients were detected by standard detection method .The levels of plasma MIP-1α ,MIP-1β and IL-6 from four groups were determined by enzyme-linked immunosorbent assay (ELISA ) .Intergroup difference was compared using t-test and interactive analysis was conducted using analysis of variance .Results In HIV-infected patients ,CD4 + T lymphocyte counts in Meth + HIV group was significant lower than non-Meth +HIV group (t= 5 .431 , P 0 .05) ,neither between HIV infection and the levels of cytokines (P> 0 .05) .Conclusion Meth abuse results in elevated expressions of MIP-1αand MIP-1β ,which indicates that Meth abuse may play a regulating role on promoting HIV infection .
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Objective To detect the expression of macrophage inlfammatory protein 1α(MIP-1α) in the myocardium of viral myocarditis (VMC) mice at different phases. Methods A total of 120 4-week-old male BALB/c mice were randomly divided into 2 groups, 80 in the VMC group and 40 in the control group. Mice in VMC group were inoculated intraperitoneally with coxsackievirus B3 to build VMC models, while mice in control group were treated with DMEM cultivate liquid. Ten mice of each group were sacriifced on days 3, 7, 15 and 30 after treatment and their heart tissues were collected for analysis. The level of MIP-1αin the myocardium was determined by immunohistochemistry. Myocardial histopathology was examined with hematoxylin and eosin stain. In addition, the relationship between the level of MIP-1αand the degree of myocardial lesion was investigated. Results The expression of myocardial MIP-1αprotein in VMC group was up-regulated in myocardium on day 3 after inoculation of virus, and slowly decreased after the peak on day 7, but still sustained a high level on day 30. Compared with the control group, the levels of MIP-1αin VMC group were increased signiifcantly at every phase (P<0.05). Furthermore, positive correlation was found between MIP-1αprotein expression levels and myocardial histopathologic scores in VMC group (r=0.94, P<0.01). Conclusion The up-regulated expression of MIP-1αmay play a critical role in the pathogenic mechanisms of viral myocarditis.
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Objective To investigate the effects of macrophage inflammatory protein-1α (MIP-1α) combined with molecule 4-1BB L on the tumorigenicity of hepatocellular carcinoma cells in vivo. Methods Mouse MIP-1α (mMIP-1α) expressed Hepa 1-6 cells were transfected with m4-1BBL recombinant retrovirus, the anti-histidinol cells clones were selected and amplified. The expression of m4-1BB L was confirmed by flow cytometry. The growth curve of Hepa 1-6 cells transfected with mMIP-1α and m4-1BBL alone or together was drawn and compared. C57B/L Mice were randomly divided into 7 groups, 9 mice in each group, injected with mMIP-1α+m4-1BB L Hepa 1-6 cells, m4-1BB L Hepa 1-6 cells, mMIP-1α Hepa 1-6 cells, Hepa 1-6 cells, pLXSHD Hepa 1-6 cells or PBS respectively. The tumorigenicity of hepatocellular carcinoma cells and the mice survival rate were compared between each groups. Results Hepa 1-6 mMIP-1α+m4-1BB L cells which expressed both mMIP-1α and m4-1BB L were successfully established. The expression of mMIP-1α and m4-1BB L alone or together did not affect the growth curve of Hepa 1-6 cells. Observed for 5 weeks, no tumor developed in Hepa 1-6 mMIP-1α+m4-1BB L injected mice. The tumorigenicity of Hepa 1-6 mMIP-1α+m4-1BB L was lower than that of Hepa 1-6 mMIP-1α or Hepa 1-6 m4-1BB L in vivo. The survival rate of Hepa 1-6 mMIP-1α+m4-1BBL injected mice(9/9) was higher than that of Hepa 1-6 m4-1BB L injected mice (6/9)or Hepa 1-6 mMIP-1α injected mice (1/9). Conclusion Chemokine MIP-1α combined with costimulatory 4-1BB L lowered the tumorigenicity of hepatocellular carcinoma cells in vivo, and prolonged the mice survival period.