Major immune-related cells in psoriasis vulgaris lesions / 中华皮肤科杂志

Psoriasis vulgaris is a recurrent inflammatory skin disease. A variety of factors, such as trauma and infection, can destroy the skin barrier function, thereby breaking the balance of immune homeostasis and tolerance, causing abnormalities in function and/or number of various immune-related cells in local skin, resulting in psoriasis-like skin changes such as abnormal proliferation of keratinocytes and excessive inflammatory reactions in skin lesions. Various immune cells in skin lesions can sense changes in the surrounding environment (autocrine or paracrine) through surface molecules, and then express and secrete a variety of inflammation-related factors; if maintenance mechanisms for immune homeostasis and tolerance become invalid, the positive feedback network of inflammation mediated by inflammation-related factors will be formed locally, leading to the occurrence of psoriasis vulgaris. This review summarizes research progress in the role of immune-related cells in skin lesions in the immunopathological mechanism of psoriasis vulgaris, especially innate immune cells such as γδT cells.

Relationship between serum mannose-binding lectin and Th17/Treg cells in patients with silicosis / 中国职业医学

OBJECTIVE: To explore the relationship between serum mannose-binding lectin( MBL) and T helper cell 17( Th17)/regulatory T cells( Treg) balance in patients with silicosis. METHODS: A total of 101 male patients with silicosis were selected in silicosis group and 62 health individuals in control group using the cross-sectional study. The level of serum MBL was measured by enzyme linked immunosorbent assay. The ratio of Th17/Treg was recorded by flow cytometry.The relative expression of retinoid-related orphan nuclear receptor γt( RORγt) and forkhead box 3( Foxp3) mRNA in peripheral blood mononuclear cell were tested by real-time polymerase chain reaction method. RESULTS: The level of serum MBL in silicosis group was higher than that of control group( P < 0. 01). The ratio of Th17 cells and the relative expression of RORγt mRNA increased in silicosis group( P < 0. 05),while the ratio of Treg cells and the relative expression of Foxp3 mRNA decreased in silicosis group( P < 0. 05) compared to the control group. The level of serum MBL had negative correlation with forced expiratory volume in the first second,forced vital capacity and forced expiratory flow( P < 0. 05) in patients with stage Ⅰ and Ⅱ silicosis. Meanwhile,the level of serum MBL had negative correlation with Th17 ratio and RORγt mRNA relative expression( P < 0. 05),and positive correlation with Treg ratio and Foxp3 mRNA relative expression( P < 0. 05). CONCLUSION: MBL might participate in the development of silicosis through regulating the balance of Th17/Treg cells.

Effect of metformin on proliferation,cell cycle and apoptosis of U937 cells / 中国免疫学杂志

Objective:To study the effect of metformin on proliferation,cell cycle and apoptosis of U937 cells.Methods: U937 cells were treated with different concentrations of metformin,collected cells in 24,48 and 72 hours.Subsequently,cell proliferation was assessed by CCK-8 assay,and the cell cycle and apoptosis were analyzed by flow cytometry (FCM).The expression of Bcl-2,Bax,p-AMPK,p53 were determined by Western blot.Results: The proliferation of U937 cells was inhibited by metformin in a time-and dose-dependent manner.Metformin-treated cells were arrested at G0/G1 phase,the cell frequency at G0/G1 phase was increased in a time-and dose-dependent manner.Metformin also induced cell apoptosis in a dose-dependent manner.It showed that 20 mmol/L metformin induced cell apoptosis in a time-dependent manner.The expression of p-AMPK,p53,Bax was up-regulated while Bcl-2 expression was down-regulated after metformin treatment.Conclusion: Metformin could inhibit the U937 cell proliferation,block the cell cycle at G0/G1 phase,and induce cell apoptosis,which may partially be attribute to the up-regulation of Bax,down-regulation of Bcl-2,activation of AMPK/p53 signaling.

IgG antibodies against Dengue virus non-structure protein 1 mediate passive sys-temic anaphylaxis in mice / 中国免疫学杂志

Objective:To ascertain whether the immune complexes (ICs) formed by Dengue virus 1 non-structure protein 1 (DENV1 NS1)and its IgG antibodies could mediate passive systemic anaphylaxis (PSA) and to explain the pathogenesis of Dengue hemorrhagic fever or Dengue shock syndrome (DHF/DSS).Methods:The monoclonal antibodies (mAbs) or mAb cocktails from 20 IgG mAbs of DENV1 NS1 prepared in this lab were screened to initiate PSA and passive cutaneous anaphylaxis (PCA) in mice.Meanwhile, the effects of GdCl3 and platelet activating factor ( PAF) antagonist CV-3988 on PSA induced by the NS1-IgG ICs were observed.Results:Two groups of monoclonal antibody cocktails with purified NS 1 were proved to be capable of provoking PCA and PSA in mice,whereas the other mAbs or mAb cocktails could be not .The murine PSA initiated by NS1-IgG(5D25+3B1) ICs could be sig-nificantly inhibited by in vivo treatment with GdCl3 or PAF antagonist CV-3988.Conclusion: The NS1-IgG ICs formed with DENV1 NS1 and IgG mAb cocktails can mediate PSA and PCA ,but not all of ICs formed by DENV 1 NS1 mAbs or mAb cocktails with DENV 1 NS1 can induce PSA ,indicating that it may be related to the special epitopes of DENV 1 NS1.The monocyte/macrophages and PAF may be as major effector cells and the major mediator for PSA induced by NS 1-IgG ICs,respectively.

Plasma levels of mannan-binding lectin-associated serine protease 2 in children with upper respiratory tract infection / 南方医科大学学报

<p><b>OBJECTIVE</b>To explore the significance of plasma levels of mannan-binding lectin (MBL)-associated serine protease 2 (MASP2) in children with upper respiratory tract infection (URTI).</p><p><b>METHODS</b>A total of 103 children with URTI and 35 healthy children were examined for plasma levels of MASP2 and C-reactive protein (CRP). According to CRP levels, white blood cell count (WBC), stage of infection, and administration of treatments, the children with URTI were divided into the elevated CRP group (n=48) and the normal CRP group (n=54), elevated WBC group (n=61) and normal WBC group (n=40), the early stage of infection without treatment group (n=68) and mid-late stage of infection with treatment group (n=35).</p><p><b>RESULTS</b>Plasma MASP2 levels was significantly higher in URTI group than in the healthy control group (P<0.001) and showed a close correlation with age (r=0.302, P<0.01). Plasma MASP2 level was significantly correlated with CRP level in elevated CRP group (r=0.310, P<0.05) but not in normal CRP group (P>0.05), correlated with WBC in elevated WBC group (r=0.392, P<0.01) but not in normal WBC group (P>0.05), and was significantly higher in early stage infection without treatment group than in mid-late stage of infection with treatment group (P<0.01). MASP2, MBL2 and CRP genes had a common binding site for the transcription factor HNF-4α.</p><p><b>CONCLUSIONS</b>MASP2 may be an acute-phase protein, and its plasma level might serve as a new reference index in the diagnosis of URTI in children.</p>

Cloning of human CD45 gene and its expression in Hela cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To clone human CD45 gene PTPRC and establish Hela cells overexpressing recombinant human CD45 protein.</p><p><b>METHODS</b>The intact cDNA encoding human CD45 amplified using RT-PCR from the total RNA extracted from peripheral blood mononuclear cells (PBMCs) of a healthy donor was cloned into pMD-18T vector. The CD45 cDNA fragment amplified from the pMD-18T-CD45 by PCR was inserted to the coding region of the PcDNA3.1-3xflag vector, and the resultant recombinant expression vector PcDNA3.1-3xflag-CD45 was transfected into Hela cells. The expression of CD45 in Hela cells was detected by flow cytometry and Western blotting, and the phosphastase activity of CD45 was quantified using an alkaline phosphatase assay kit.</p><p><b>RESULTS</b>The cDNA fragment of about 3 900 bp was amplified from human PBMCs and cloned into pMD-18T vector. The recombinant expression vector PcDNA3.1-3xflag-CD45 was constructed, whose restriction maps and sequence were consistent with those expected. The expression of CD45 in transfected Hela cells was detected by flow cytometry and Western blotting, and the expressed recombinant CD45 protein in Hela cells showed a phosphastase activity.</p><p><b>CONCLUSION</b>The cDNA of human CD45 was successfully cloned and effectively expressed in Hela cells, which provides a basis for further exploration of the functions of CD45.</p>

Effects of mannan-binding lectin on the functions of human polymorphonuclear cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effects of mannan-binding lectin (MBL) on the functions of human polymorphonuclear cells (PMNs).</p><p><b>METHODS</b>ELISA and Dot blot were performed to examine the binding between MBL and the microorganisms. Flow cytometry and fluorescence microscopy were employed to analyze the phagocytosis of FITC-labeled microorganisms by the PMNs. Real-time quantitative PCR was used to detect the expression levels of IL-1β, TNF-α and CD11b mRNA in the PMNs, and ELISA used to detect the levels of TNF-α and IL-6 in the supernatants of PMN culture. Nitro-blue tetrazolium reduction assay was used to estimate the levels of superoxide production.</p><p><b>RESULTS</b>MBL bound to the microorganisms in a dose-dependent manner. MBL had no significant effect on phagocytosis of C. albicans and E.coli by the PMNs in the absence of human serum, but in presence of mixed MBL-deficient human sera, MBL promoted the phagocytosis of C. albicans, which could be blocked by mannan. Mannan treatment increased the expressions of IL-1β, TNF-α, IL-6 and CD11b and enhanced superoxide production in the PMNs.</p><p><b>CONCLUSION</b>MBL can promote phagocytosis of microorganisms by PMNs and increase the expressions of proinflammatory cytokines from PMNs in a complement lectin pathway-dependent manner.</p>

Preparation of the trimeric subunits of recombinant human mannan-binding lectin and analysis of its bioactivity / 南方医科大学学报

<p><b>OBJECTIVE</b>To prepare the trimeric subunits of recombinant human mannan-binding lectin (MBL) with biological activities.</p><p><b>METHODS</b>A prokaryotic expression vector containing human MBL N-terminal deletant (rhMBLδN) gene we previously constructed was transformed into E. coli for efficient expression of rhMBLδN fusion protein. Based on the principle that the collagen polypeptides tend to self-assembly into the tertiary structure of proteins by forming a triple helix due to the characteristic properties of the collagen proteins, rhMBLδN fusion protein was limitedly hydrolyzed with thrombin. The obtained rhMBLδN polypeptide was repeatedly dialyzed in 50 mmol/L PBS (pH7.2) and ddH(2)O, and the final product was analyzed for its bioactivities using a ligand-binding assay and a C4d deposition assay.</p><p><b>RESULTS</b>rhMBLδN polypeptide with a relative molecular mass of about 20 000 was obtained by limited proteolysis of rhMBLδN fusion protein with thrombin. Repeated dialyses of rhMBLδN polypeptides in 50 mmol/L PBS and ddH(2)O resulted in the isolation of the trimeric subunit trhMBLδN (with a relative molecular mass of about 50 000), which contained a collagen-like helix. The trhMBLδN protein had a higher ligand-binding activity than rhMBLδN polypeptide, and acquired the activity to initiate the lectin pathway of complement activation, but the activities were lower than those of natural MBL.</p><p><b>CONCLUSION</b>We have successfully obtained the bioactive trimeric subunit of rhMBL, trhMBLδN, and this structural subunit is also the functional subunit of the MBL molecule.</p>

Change and significance of urinary excretion of aquaporin-2 detected by enzyme linked immunosorbent assay in rat models of heart failure / 中国组织工程研究

AIM: To detect the change of urinary concentration of aquaporin-2 (AQP-2) by enzyme linked immunosorbent assay in rat models of different degrees of heart failure and make a comparison with sham-operation group.METHODS: This experiment was carried out between January 2000 and January 2002 in the animal laboratory of Nanfang Hospital of Southern Medical University. Forty-two male adult Sprague-Dawley rats were involved. Twenty-six rat models of chronic heart failure were prepared by ligation of left coronary artery. When left ventricle infarct area was≥20%, the rat models of congestive heart failure were successful (heart failure group, n =13); When left ventricle infarct area was＜20%, the rat models of congestive heart failure were unsuccessful (compensation group, n =13). The other 16 rats were not ligated at coronary rtery (control group). Serum sodium concentration was determined with BeckmanC×3 equipment and urine osmole by cryoscopic method. Urine volume of 24 hours was monitored. Urinary concentration ofAQP-2 level of rats was determined by double antibodies sandwich enzyme linked immunosorbent assay (DABs-ELISA).RESULTS: Forty-two rats were involved in the result analysis. The 24-hour urine volume and serum sodium concentration in the heart failure group and compensation group were significantly lower than those in the control group (P＜0.05-0.01), while urine osmole in two groups was significantly higher than that in the control group (P＜0.05-0.01).②At postoperative 4 and 6 weeks, urinary concentration of AQP-2 level of rats in the control group was significantly lower than that in the other two groups (P＜0.05-0.01), and urinary concentration of AQP-2 level of rats in the compensation group was significantly lower than that in the heart failure group (P＜0.05, 0.01).In the compensation group and heart failure group, urinary concentration of AQP-2 level of rats was significantly higher at postoperative 6 weeks than at postoperative 4weeks (P＜0.05).CONCLUSION:①AQP-2 is the key target protein of water retention and hyponatremia at heart failure.②Detection of urinary concentration of AQP-2 by ELISA can effectively reflect water retention and hyponatremia when heart failure occurs.