| Background:Prolactin (PRL) is produced by lactotroph cells of the anterior pituitary.The classic role is to regulate mammalian breast, ovary and the growth anddifferentiation of male genitals subsidiary, and to promote lactation. In addition, PRLhas received increasing attention to the role of immune regulation. The reseach showsthat the PRL not only in physiological regulation of cellular and humoral immuneresponse, but pathological conditions (such as autoimmune diseases), cellular and thehumoral immune function also have a very important role in the regulation. Thefunction on that include:â‘ It is pacticipate in T cells differentiation, and regulates thebalance in TH1/TH2,moreover, it urges a diversion from TH1;â‘¡It activates proteinkinase C and ornithine cyclase;â‘¢It has the function to promote mitogen, and theIL-12 receptor which induces lymphoid cell is subjected to the expression that thebody expresses with various gene with related growth factor;â‘£The passes theexpression that the interferon regulates factor to promote interferon-r;⑤Stimulating the creation of auto-antibody.The activation of lymphoid cells need double signal systems:The first signalsystem is the compound of antigen peptide MHCâ…¡member and TCR-CD3,thesecond signal is co-stimulate molecule, including the CD28/B7(the CD80 and CD86)members and CD40/CD40 ligand(CD154) members. The CD28/B7 mainly hassomething to do with T cell activating, propagating and apoptosis, the CD40/CD154mainly has something to do with B cell activating, propagating and apoptosis.Systemic lupus erythematosus (SLE) is the prototype of human autoimmunedisease.The abnormal immune regulation played a leading role in its pathogenesis,including many immune cells function. The clinical research has shown that theexpression of CD154 were significantly increased in the active SLE patients,and areassociated with the disease activity. The expression of CD28 in the lymphocytes inthe active and quiescent SLE were not significantly difference in the control group,and the expression of CD80 and CD86 were significantly higher than normal, and theactive group was significantly higher than static group, its expression level waspositively correlated with disease activity. The costimulatory molecules(CD28/B7and CD40/CD154) play an important role in the incidence and development in thesystemic lupus erythematosus.The SLE mice will be dead after injected PRL, and themortality rate of mice can be decrease after inhibiting the synthesis and secretion ofprolactin. The clinical study reported that the patients with SLE in high PRLlevels was positively correlated with ANA titers and the clinical indicators of disease.It can speculate that the PRL plays an important role in the pathogenesis of SLE, butthe specific mechanism is not clear yet.Whether it related with the expression of theco-stimulate molecule,and it is the major problem in this reseach.The reseach found that PRL can induce the rapid phosphorylation in variety ofcomplex protein of TCR/CD3,and potentially affect the function of antigen receptor, and involved in the process of the cell activation in the first signal. PRL both at homeand abroad have not report in the role of the second signal in cellactivation.Therefore, we studied the relation with the prolactin and the expression ofco-stimulate molecule on the surface of PBMC in SLE patients,and to explain theprolactin participation in the incidence mechanisms of lupus. The pathway of theintracellular signal transduction in PRL is the research focus in recent years, theconsensus has been reached is the transmembrane of signal transduction which topromote the main way of JAK-STAT after the three dimer in the combination of thePRL and PRLR.It caused the transcription and expression of target gene(interferonregulator factor -1) after introducting the information in nuclear. IRF-1 is amultifunctional transcript factor gene, that belongs to a small factor family oftranscription, and participate in a variety of immune function, including inhibition oftumor, differentiation of bone marrow cells,macrophage activation,antigen-presenting, apoptosis. In particular, the importance is that the IRF-1 canthrough with the cis-regulatory element binding in the interferon (IFN)-induced geneand regulate the expression of the typeâ… interferon and the related gene,but theabnormal expression of that plays an important role in the pathogenesis of SLE.Thestudy investigate the expression of IRF-1 gene and its intervented by PRL inSLEpatients.Bomocriptine is a kind of excite dose which be the receptor about the two type ofdopamine, it can reduce the produce of deoxyribonucleic acid and ribonucleic acidmessenger in the lactotroph cells of the anterior pituitary ,consequently, it restrain thecell secretes excessively, and repress the biologic activity of PRL in the outside of theblood circulation.At present,some clinical studies use the BRC to treat certainautoimmune diseases, and it has been reported that the BRC25mg/day significantlyreduced the SLE disease. After oral BRC,the negative impact of high prolactin and high estrogen can be removed quickly in the SLE patients of postpartum, andsignificantly reduced the incidence of the aggravation in SLE. Moreover, the estrogenand the dosage of immunosuppressive agents after postpartum were be significantlydecreased. This experiment mainly passes BRC on the expression of co-stimulatemolecule on the surface of the PBMC in SLE patients,and to explore the majormechanisms for the treatment of SLE.Objective:1. To detect the content of the serus prolactin (PRL) in systemic lupuserythematosus(SLE)patients and to study the correlations among PRL and SLEactivity and clinical manifestations and laboratory indicators.2. To investigate the influence of prolactin on the expression of the CD40,CD154,CD28,CD86 and CD80 on the surface of the peripheral blood mononuclearcells of the patients with systemic lupus erythematosus and to discuss its status inSLE pathogenesy.3. To investigate the change after joined bomocriptine on the CD40,CD154,CD28,CD86 and CD80 on the surface of the peripheral blood mononuclear cells ofthe patients with systemic lupus erythematosus and to judge whether it can be use tocure the SLE.4. To research the IRF-1 gene expression and its intervention by PRL in SLEpatients.Methods:1. The ontent of the serus PRL was examined by electrochemilumescence-meterin 30 cases with SLE and 20 cases healthy human in the morning of quiescentcondition.2. To divide samples as follows:â‘ SLE groups with high PRL (N=18);â‘¡SLEgroups with normal PRL (N=12);â‘¢Normal control groups (N=20);â‘£SLE groups with normal PRL+rhPRL;⑤normal control groups+rhPRL;â‘¥SLE groups withhigh PRL +Brc;⑦SLE groups with normal PRL+rhPRL+Brc. Take the 12 boreplanks, and to make the each bore cells to attain 5×106/ml.The concentration ofrhPRL is 10ng/ml finally, and The concentration of Brc is 2ug/ml finally.3. Stimulated training:The PBMC in the peripheral blood were be separated bydensity gradient centrifugation and separation of training, the following steps:â‘ Inthe calibration (12ml) of the sterile tube. Each tube joined 4mlFicoll lymphocytesisolated liquid-taking the peripheral blood 10ml heparin (including 30 SLE patientsand 20 normal controls females) with the same blending PBS fully diluted. With thePasteur dropper drawed with the anticoagulant 5ml (anti-clotting diluted with PBSequivalent) and along slowly superimposed on the wall of lymphocyte liquid cellseparation (by 4 per sample tube), 20℃, 20min 2000rpm level centrifugal;â‘¡Usingthe suction to reach gray hair cell layer, and drawing the mononuclear cells placed ina separate centrifuge tube, adding more than five times the size of PBS,20℃, 10 min1500rpm level centrifuge, washing the cells number 1, being the samples of PBMC inthe study.â‘¢Sent to the FCM.after each of the cells into 37℃,5% CO2 incubatorwithin 24 hours.4. Cultured cells detected by flow cytometry: five each tube, 1×106cells/tube.After washing twice in PBS,and joining the monoclonal antibody ofanti-CD40,CD154, CD28, CD80,CD86 followed by five control 18ul and two withthe management control Mouse IgG1-FITC and Mouse IgG1-PE 18ul,4℃, darkincubated for 30 min in PBS washed 2 times after adding more than 1%paraformaldehyde fixation 300ul inspection. On the plane before using standardequipment fluorescent microspheres adjusted coefficient of variation of 2.0%. Aftercollecting 10,000 on the cell fluorescence intensity to the number of amplified opticalShanshe measured data to calculate the expression of CD40,CD154, CD28, CD80, CD86 cells, fluorescent cells were drawn.5. Application of reverse transcriptase-polymerase chain reaction (RT-PCR) withgel image scanning detect the expression of IRF-1 gene. The expression ofβ-actingene were controled, the ratio of IRF-1 andβ-actin PCR scanning value of amplifiedproduct product will be relative quantitative analysis as the relative values of theIRF-1 gene expression.Results:1. The average level of PRL in SLE patients was higher than in normal controlwhich was very obvious in active stage. The content of the serus PRL in SLE patientswas directly related with SLEDAI.2. The renal damage, anti-ds-DNA antibodies, complement C3, C4 decline in theincidence of high PRL with SLE patients was significantly higher than normal, andskin rashes, arthritis, Serositis, and blood was no significant difference between theincidence of involvement.3.â‘ The expression of CD154 level in high PRL patients with SLE weresignificantly higher than the normal PRL patients and normal controls. The normalPRL patients and the normal control group showed no significant differences in theexpression of CD154.â‘¡The expression of CD86 in the high and normal PRL in SLEpatients was significantly higher than the normal control group. But the expression ofCD86 in high PRL and normal PRL in SLE patients were no significant difference.â‘¢The expression of CD40, CD28, CD80 in high PRL and normal PRL with SLEpatients and the control group were no significant difference.4.â‘ After the rhRPL stimulating, the expression of CD154 in normal PRL levelwith SLE patients was significantly higher than before.â‘¡After the rhRPLstimulating, the expression of CD154 in normal control group with no significantdifference in stimulating ago. 5.â‘ After joined the BRC, the expression of CD154 in high PRL with SLEpatients were no significant difference than before..â‘¡The normal PRL in SLEpatients joined rhPRL and BRC,the expression of CD154 were no significantdifference compared with the stimulating ago.6.â‘ 30 SLE patients and 20 normal control group both have detected thespecific fragments of IRF-1 andβ-actin, which IRF-1/-actin ratio was 0.89±0.21 inSLE patients,and high PRL in SLE patients were 1.06±0.26, normal PRL in SLEpatients were 0.82±0.21,whereas the normal control group were 0.78±0.18.Statistically, the relative value of IRF-1 gene expression in SLE patients significantlyhigher than the normal control group. High PRL were significantly higher thannormal PRL in SLE patients, and the normal PRL in SLE patients and healthy controlgroup were not significantly different.â‘¡After rhRPL stimulating, the relative valueof IRF-1 gene expression in normal PRL with SLE patients were 0.99±0.22,significantly higher than rhRPL stimulating before.â‘¢After rhRPL stimulating, therelative value of IRF-1 gene expression in normal control group were 0.79±0.18,and have no significant difference with rhRPL stimulating before.Conclusion:1. Serus PRL may play a certain role in SLE pathogenesyo The increase of thecontent PRL is obviously related with SLE activity which can be used as a index toassess SLE activity.2. PRL play an important role in SLE pathogenesy that through increasing theexpression of CD 154 on the surface of PBMC. PRL in SLE can speculate on therole of immune regulation that may be combined with PBMC surface PRLR ,andthe PRL-PRLR inherently complexed, and increased the co-stimulate signal moleculeby the expression of CD154.3. In SLE, high PRL has not causal relationship between the expression of CD86, it shows that the CD28/B7 is not involved in the pathogenesis of SLE.4. BRC has not reduced endogenous or exogenous prolactin (rhRPL)-inducedexpression of CD154.It confirmed that the BRC may be the main mechanism for thetreatment of SLE inhibiting pituitary secretion of prolactin., thereby reducing serumPRL levels to achieve therapeutic purposes, but has no antagonism or inhibition ofPRL.5. The study confirmed the existence of abnormal IRF-1 gene expression inSLE.. And the high PRL levels in SLE may be the one of the major reasons thatleading to the abnormal IRF-1 gene expression.6. In SLE, PRL can up-regulated the expression of CD154 in the extracellular, inintracellular, it through the JAK-STAT signaling pathway to activate and regulate theIRF-1 gene expression.
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