【Abstract】 Objective To observe the plasma levels of adiponectin and interleukin-17 ( IL-17) in patients with chronic obstructive pulmonary disease ( COPD) at acute exacerbation or stable stage, and analyze their relationship. Methods Sixty male COPD patients with normal weight ( with BMI range of 18. 5-24. 9 kg/m2 ) were enrolled, including 30 patients with acute exacerbations of COPD ( AECOPD) and 30 patients with stable COPD. Twenty healthy nonsmoking male volunteers were included as controls. The plasma levels of adiponectin and IL-17 as well as lung function ( FEV1% pred and RV% pred) were measured in all subjects. Results The concentrations of adiponectin and IL-17 were significantly higher in the AECOPD patients than those of the patients with stable COPD and the contro1s ( P lt; 0. 001) . Theconcentrations of adiponectin and IL-17 were significantly higher in the patients with stable COPD than those of the controls ( P lt;0. 01) . Adiponectin was positively correlated with IL-17 in the AECOPD patients ( r =0. 822, P lt;0. 001) and in the patients with stable COPD ( r =0. 732, P lt;0. 001) . Adiponectin was positivelycorrelated with RV% pred in the AECOPD patients ( rs = 0. 764, P lt;0. 001) and in the patients with stable COPD ( rs =0. 967, P lt;0. 001) . There was no significant relationship between adiponectin and FEV1% pred ( P gt;0. 05) . Conclusions The plasma level of adiponectin in COPD patients is elevated which is relatedwith excessive inflation of lung. Adiponectin may be involved in the process of inflammation in COPD as a new pro-inflammatory cytokine.
Objective To observe the influence of interleukin-1beta; (IL-1beta;) on the expression of phosphorylated signal transducers and activators of transcription 3 (pSTAT 3) in rat retinal Muuml;ller cells.Methods For in vitro study cultured Muuml;ller cells were treated with IL-1beta; of different concentrations (0, 0.1, 1, 5 and 10 ng/ml) for 24 hours. For in vivo study, 32 Sprague-Dawley（SD）rats were divided into 4 groups randomly (control group，100，500 and 1000 ng/ml group) with 8 rats in each group. After 24 hours of injection with phosphate buffered solution (PBS), or 100,500,1000 ng/ml IL-1beta; into the vitreous cavities of the above rats, retinas were harvested. The expressions of pSTAT3 in cultured Muuml;ller cells or treated retinas were evaluated by indirect immunofluorescence and western blotting.Results After 24 hours of incubation without IL-1beta;, pSTAT3 has little expression in cultured Muuml;ller cells, but was upregulated by 1 ng/ml or higher IL-1beta; in a dosagedependent manner (F=46.64, 43.78;P＜0.01). pSTAT3 was not expressed in adult rat retina, but was upregulated by vitreous injection of 100 ng/ml or higher IL-1beta; in a dosagedependent manner (F=73.53,43.70;P＜0.01).pSTAT3 expressed mainly in inner nuclear layer and ganglion cell layer. Doublelabeling showed that there was no costaining of pSTAT3 and glial fibrillary acidic protein (GFAP) in retina of control group, but there were many costained Muuml;ller cells in retinas treated with IL-1beta;.Conclusions Expression of pSTAT3 in Muuml;ller cells could be activated by IL-1beta; which may represent one pathway link to reactive gliosis.
ObjectiveTo study the inhibitory effects of pigment epithelium derived factor (PEDF) on oxygen-induced retinal neovascularization in mice, and to investigate the possible involvement of interleukin-1β (IL-1β) in the neovascular-inhibitory function of PEDF.
Methods A total of 140 postnatal day (P)7 C57BL/6 mice were randomly divided into normal control group, oxygen-induced retinopathy (OIR) model group, PEDF treatment group and PBS treatment control group. All mice except normal control group with their mothers were exposed to (75±2)% oxygen environment for 5 days and then kept in room air for another 5 days to establish the OIR model. Mice in normal control group were kept in room air only. At P12 and P14, respectively, mice in PEDF treatment group received intravitreous injections of 1 μl PEDF (2 μg/μl), while PBS treatment control group received the same volume of PBS (10 mmol/L, pH7.4).All mice were euthanized at P17 and eyes were isolated. The changes of retinal vessels were observed on retinal flat mounts and cryosections by fluorescence microscopy. Retinal specimens were prepared for IL-1β protein and mRNA analysis by Western blot and real time fluorescence quantitative reverse transcription-polymerase chain reaction (Real-time RT-PCR).
ResultsChanges of retinal vessels had been viewed by fluorescence microscopy on flat-mounted retina, the relative retinal neovascularization areas were significantly increased in OIR model group compared with normal control group (t=15.02, P < 0.01), and the relative retinal neovascularization areas were obviously smaller in PEDF treatment group than those in PBS treatment control group (t=5.96, P < 0.01). Fluorescence staining revealed that retinal vascular tufts were extending from outer plexiform layer (OPL) to ganglion cell layer (GCL) of the retina along with multiple interconnections; Neovascular tufts in OIR model group and PBS treatment control group were presenting distinctly more than those of normal control group and PEDF treatment group. The specific expression levels of IL-1β protein in retinas of OIR mice by Western-blot analysis were higher than those of normal control group(t=3.35, P < 0.05), While these of PEDF treatment group showed a considerable decline in comparison with PBS treatment control group (P < 0.01), and there were no difference in normal control group and PEDF-treated group (F=11.764, P > 0.05). Similarly, expression levels of IL-1β mRNA tested by Real-time RT-PCR were obviously increased in the OIR model group when compared to normal control group(t=4.43, P < 0.01). After treated with PEDF, expression levels of IL-1β mRNA showed a considerable decrease when compared to PBS treatment control group (P < 0.01), and there were no difference in normal control group and PEDF-treated group (F=11.15, P > 0.05).
ConclusionsPEDF can inhibit oxygen-induced retinal neovascularization. The mechanism may be related to that PEDF can downregulate the expression of IL-1β in retina.
Objective To investigate the expression and significance of Fork head /winged helix protein 3 (Foxp3) , retinoic acid-related orphan receptorγt (RORγt) , and interleukin-17 (IL-17) in Guinea pigs with emphysema. Methods Smoking and active immunization with elastin were separately used in guinea pigs to establish emphysema model. Then the destruction of lung tissue was assayed by measurement of the average radius of alveolar. The expressions of Foxp3 , RORγt, and IL-17 in lung tissue of the guinea pigs were detected by immunohistochemical technique. The results were compared with the normal control group by the analysis of variance or kruskal-Wallis test. Spearman rank correlation was used to analyze the correlation between the ratio of Foxp3/RORγt and IL-17, also the correlation between Foxp3/RORγt and the average radius of alveolar. Results In the smoking group and the active immunization group, the average radius of alveolar were significantly longer than the control group (Plt;0.05) . And the expression of Foxp3/RORγt was significantly unbalanced, with the number of Foxp3-positive cells decreased and RORγt-positive cells increased (Plt;0.05) . Meanwhile the level of IL-17 was significantly increased compared with the control group ( Plt;0.05) . The difference between the smoking group and the active immunization group was not significant (Pgt;0.05) . The ratio of Foxp3/RORγt was negatively correlated with the level of IL-17 and the average radius of alveolar. Conclusions Active immunization with elastin can induce emphysema in guinea pigs. The Foxp3/RORγt expression was unbalanced in lung tissue of guinea pigs with emphysema.This imbalance may be an important mechanism attributed to the disordered expression of CD4+ Treg cells and Th17 cells, which may be involved in autoimmune regulation and development of chronic obstructive pulmonary disease.
Objective To evaluate the role of interleukin-10 (IL-10) in acute pancreatitis. Methods Thirty mongrel dogs were divided into three groups based on the severity: acute edematous pancreatitis (AEP) group (n＝11), acute hemorrhagic necrotizing pancreatitis (AHNP) group (n＝12), and control group (n＝7). Serum level of IL-10 was determined with enzyme-linked immuno-sorbent assay (ELISA). Results Within 24 hours, AEP group had serum level of IL-10 significantly higher than that of AHNP group. Control group had no detectable serum IL-10. No significant difference was observed between AEP group and AHNP group at 48 hours. Conclusion The finding of low values of serum IL-10 suggests that there may be more consumption in AHNP group than in AEP group and it may be beneficial to decrease the severity of experimental acute pancreatitis.
Objective To observe the alteration of anti-inflammatory cytokines (IL-10 and TGF-β) in acute pancreatitis. MethodsSD male rats were divided into 2 groups: group 1, the normal rats as a control (n＝6); group 2, the acute pancreatitis induced by intraductal injection of 5% sodium cholate sulfur with the volume of 1.0 ml/kg。 The animals were killed at 2(n＝6), 6(n＝6) and 24 hours (n＝8) after operation, the blood samples were taken for measurement of IL-10, TGF-β (by ELISA). The weight of pancreatic tissue and amylase were also observed. Results Serum IL-10 and TGF-β in control group were 32.05±14.87 pg/ml and 66.40±13.20 pg/ml, respectively. Serum IL-10 in group 2 was 36.52±9.76 pg/ml (2 hour), 37.75±6.54 pg/ml (6 hour), and 68.13±19.90 pg/ml (24 hour), respectively. Serum TGF-β in group 2 was 64.58±10.56 pg/ml (2 hour), 72.87±18.34 pg/ml (6 hour), 103.77±28.95 pg/ml (24 hour), respectively. Compared to that of normal rats, the serum level of IL-10 and TGF-β in 24 hours of acute pancreatitis increased significantly (Ｐ＜0.05). Conclusion Anti-inflammatory cytokines, both IL-10 and TGF-β were increased remarkablly in acute pancreatitis. This result indicates that there is a potential tendency of compensatory anti-inflammatory response sydrome in acute pancreatitis.
Objective To construct a regulatable plasmid containing single chain fusion gene of murine interleukin-12 (mIL-12) which was regulated with mifepristone (RU486) and explore its expression in vitro. Methods The p40 and p35 subunit sequence of mIL-20 were respectively obtained from the plasmid GCp35Ep40PN by polymerase chain reaction (PCR) and they were cloned into pCA14 plasmid after introducing a linker by overlap PCR. The single chain mIL-12 gene was comfirmed by sequencing and subcloned into pRS-17 vector which contains RU486 regulator cassette. The positive clone named pRS-RUmIL-12 was identified by restriction endonuclease digestion and PCR. Lipofectamine 2000 was used to transfect the pRS-RUmIL-12 to HEK293 cells followed by manufacturer’s recommendations. The protein concentration of mIL-12 induced with RU486 in supernatant of the transfected HEK293 cells was measured by ELISA. Results The sequence of single chain mIL-12 what we obtained was the same as the expected result. The results of restriction endonuclease digestion and PCR showed that the RU486-inducible regulatory vector (pRS-RUmIL-12) was successfully constructed. No significant mIL-12 protein concentration in supernatant of HEK293 cells activation was measured without the inducer RU486, whereas higher concentration of the mIL-12 protein was observed in the presence of RU486. The relationship of concentration of the mIL-12 protein and RU486 was positive correlated under definite range. Conclusion A regulatable eukaryotic expression plasmid of mIL-12 single chain fusion gene was constructed, which could be used in the further research of gene regulation and gene therapy.
Interleukin-1 (IL-1), interleukin-2(IL-2) and interleukin-6(IL-6) activities and tumor necrosis factor (TNF) contents in plasma from patients with different sites of cancers as well as controls using bioassay technique were studied. The results showed that the levels of IL-1,IL-2,IL-6 s from patients with different sites of cancer were decreased remarkably in comparision with controls and the contents of TNF from patients with different sites of cancers increased significantly. But the difference between different sites of cancer was not statistically significant. The data suggest that the variations in the contents of TNF and the levels of interleukins may be related to the development of these caner patients.
【Abstract】Objective To investigate the role of interleukin-10（IL-10） and interleukin-18 （IL-18） in the pathogenesis of acute lung injury in experimental severe acute pancreatitis.Methods Forty-eight SD rats were divided into control group and SAP group by the random data table. The model of experimental severe acute pancreatitis was established by injection of 3.5% sodium taurocholate into the bili-pancreatic duct. Lung wet weight index, ascities and level of serum amylase, IL-10 and IL-18 were quantitatively measured in different time. Intrapulmonary expressions of IL-10 mRNA and IL-18 mRNA were detected by semiquantitative RTPCR. The histopathology of pancreas and lung were observed under the light microscope.Results Lung wet weight index, ascities, level of serum amylase, IL-10 and IL-18, intrapulmonary expressions of IL-10 mRNA and IL-18 mRNA were significantly increased in SAP group (P＜0.01). The level of serum IL-18 and intrapulmonary expression of IL-18mRNA are positively correlated with lung wet weight index （r=0.68,P＜0.01; r=0.72,P＜0.01） and lung injury score （r=0.74,P＜0.01; r=0.79,P＜0.01) respectively, whereas the level of serum IL-10 and intrapulmonary expression of IL-10 mRNA are negatively correlated with lung wet weight index（r=-0.62,P＜0.01; r=-0.69,P＜0.01） and lung injury score（r=-0.66,P＜0.01; r=-0.60,P＜0.01）. Conclusion IL-18 may play a key role in the pathogenesis of acute lung injury in experimental severe acute pancreatitis, and IL-10 exerts the protection role in this process.
Objective To investigate the changes of microRNA-150 ( miR-150) in peripheral blood leukocytes in sepsis patients, and their relationship with expression of immune cytokines and sepsis severity. Methods The level of mature miR-150 was quantified by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) and normalized to that of control miRNA, U6, in peripheral blood leukocytes of 40 patients with sepsis, 20 patients with systemic inflammatory response syndrome ( SIRS) , and 20 normal individuals. Serum concentrations of tumor necrosis factor alpha (TNF-α) and interleukin-10 (IL-10) were measured by enzyme-linked immunoabsorbent assay in all subjects. The sequential organ failure assessment ( SOFA) score systemwas used to evaluate the severity of sepsis. The relationships between miR-150 and the white blood cell count ( WBC) , TNF-α, IL-10 and SOFA score of the sepsis patients were analyzed. Results MiR-150 was stable for at least 5 days when specimen stored at 4 ℃ and the determination of miR-150 had a broad linear detecting range ( 6. 97-6. 97 ×104 pg/ μL RNA, the lowest detecting limit: 6. 97 pg/μL RNA,r=0.999) .MiR-150 expression in the peripheral blood leukocytes in the sepsis group was significantly lower than that in the healthy control group ( Plt;0.01) , while WBC, IL-10 and IL-10/TNF-α ratio were significantly higher ( Plt;0.05) . There was no significant difference in levels of miR-150, IL-10, IL-10/TNF-α ratio, and WBC between the sepsis group and the SIRS group (Pgt;0.05) . There was no significant difference in serum concentrations of TNF-α among three groups ( Pgt;0.05) . MiR-150 expression in non-survivor sepsis patients was significantly lower than that in survivor sepsis patients (Plt;0.05) , while serum IL-10 and IL-10/TNF-αratio were significantly higher (Plt;0.01) , but there was no significant difference in serum TNF-α between the non-survivor group and the survivor group ( Pgt;0.05) . There was significantly negative correlation between miR-150 and SOFA score, TNF-α and IL-10( r=-0. 619, - 0.457, -0. 431, Plt;0.05, respectively) , but no correlation between miR-150 and WBC ( r =-0. 184, Pgt;0.05) . There was no relationship between serum TNF-α, IL-10, IL-10 /TNF-α ratio or SOFA score ( Pgt;0.05) . Conclusions MiR-150 expression in the peripheral blood specimens is significantly decreased in sepsis patients. The expression level of miR-150 not only reflect the situation of inflammatory response, but also may be used as a prognostic marker in sepsis, as it can reflect the severity of sepsis in certain degree.
