The Investigation For The Patient Suffer Rfom Ear,Nose And Throat Infection In Pirmary Humoral Immunodeficiency Disease And Correlation Analysis Of PLC Gamma2

ObjectivePrimary humoral immunodeficiency disease is most common one ofprimary immunodeficiency diseases (PID) which associated with geneticfactors. The repeated infection is the main clinical manifestation whichthreaten the life of patients and the complications is also threaten thepatient’s health. In recent years, primary humoral immunodeficiencydisease is paid attention by all the medical profession gradually and hadthe gained great achievements. However, study on these diseases in Chinais fewer, especially in Yunnan Province. This study intends to explore theincidence of primary humoral immunodeficiency disease from the ear,nose and throat infection (ENT) in the Yunnan. Then the expression levelof phospholipase C gamma2(PLCγ2), which is a key molecule ofsignaling transduction in B lymphocyte was detected through themolecular biological methods in the primary humoral immunodeficiency disease to analyze the correlation between the disease and PLC gamma γ2.The study is helpful for the etiology diagnosis and treatment of thediseases.Methods1. The patients infected with chronic and recurring infection from TheAffiliated Hospital fo Dali University and the People’s Hospital ofDali State were selected as study subjects. We taked the patients’medical history and recorded the clinical manifestation. The sera fromthe patients were collection and the levels of immunoglobulin G, Aand M were determined by a single agar diffusion test. The diagnosisof primary humoral immunodeficiency disease is according to themedical history, the levels of Ig and the results of flow cytometry ofthe patients.2. Peripheral blood mononuclear cells (PBMC) is isolated fromEDTA-Na2-whole human blood with Ficoll-Hypaque （F/H） using adensity gradient centrifugation procedure.3. Total RNA was isolated from the peripheral blood mononuclear cellsand first-strand cDNA was synthesized from the total RNA.Semi-quantity PCR was performed to analyze PLCγ2gene expressionusing3-fold dilutions of the cDNA template. The results werecompared with the health population. 4. The peripheral blood mononuclear Cells were lysed with the lyticsolution contained several proteinase. The concentration of theproteins were detected by BCA. Cell lysates were subjected toSDS-PAGE, transferred to PVDF membranes by semi-dryelectrophresis transfer, blotted with anti-human PLCγ2or anti-Actinantibodies, and visualized with ECL chemiluminescent substrates.The results were compared with the results of health population.5. The saliva from the chronic and reccuring ENT infected patients werecollected. The levels of SIgA were detected by competitive ELISAwith the specific antibody aganst SIgA. The results were calculatedby ELISA Calc software. The change of SIgA in primary humoralimmunodeficiency diseases were analyzed.Results1.7patients of common variable immunodefficiency disease werediagnosed in these subjects according to the medical history, the agefrom4.9to40years,male5cases,female2cases. Serumimmunoglobulin IgM all decreased. The levels of Ig and theresults of flow cytometry of the patients. The other primary humoraldiseases were not found in this study. 2. The results of semi-quantity PCR show that the levels of PLCγ2genemRNA expression in CVID is not different from the levels of thehealth population.3. On the above seven cases of CVID patients and other four patientsdiagnosed of CVID (2cases of systemic lupus erythematosus,1caserheumatoid arthritis and1case of white blood cells reduce disease), Itfound that PLCγ2expression on protein has difference by the westernblot test, PLCγ2expression on protein have defect among threepatients, compared with normal. And the incidence is3/11(27.27%).While protein level expression has not affected in others.4. The level of SIgA in the patients infected with chronic and recurringENT infection is lower than that of the health population,P=0.009<0.05, t=-2.818, the difference has statistically significant;The results show that the patients with low level of SIgA also havelow level of serum Ig, P=0.002<0.05, t=3.385, the difference hasstatistically significant; The study showed that the level of SIgA inCVID patients is lower than that of the health population,P=0.009<0.05， t=2.816, the difference has statistically significant;That means that the CVID patients also have reducing the level ofSIgA and serum Ig. ConcluTsiaoknesn together, the above data demonstrate that we firstly fonudCVID patients in Yunnan, that cell signaling transduction moleculePLCγ2may play a important role in some CVID patients according toPLCγ2protein expression, and that the CVID patients but have decreasedthe level of SIgA and also have decreased the level of serum Ig.