Significance Upregulation Of CD69and CD107a On PBMC For Delayed-type Drug Hypersensitivity

Background and objective:Nowadays there are a growing number of adverse drug reactions(ADRs) with the emergenceof a large number of new synthetic drugs, and the injection of traditional Chinese medicine andChinese medicine tablet. ADRs mainly include excessive drug reactions, side effects, druginteractions, intolerance, idiosyncratic, false allergy, drug allergy and so on. Drughypersensitivity(DH) is one of an important adverse drug reactions with a rate of15%among allADRs.The most common type is delayed-type drug hypersensitivity(DTH), which includesexanthematous eruption,acute generalized exanthematous pustulosis(AGEP),fixed drug eruption(FDE),toxic epidermal necrolysis (TEN),maculopapular exanthema(MPE),Steven-Johnson syndrome(SJS),drug-induced hypersensitivity syndrome (DIHS) or drug reaction with eosinophilia andsystemic symptoms(DRESS), systematic contacting dermatitis, eczematoid dermatitis and so on.The mechanism of drug hypersensitivity reaction is complex, which involves typeⅠallergy(immediate hypersensitivity), typeIIallergy (cytotoxin type), type Ⅲ allergy(immunity compound type), type IV allergy (delayed type). It has the diversifiedmanifestation, often resulting in the multi-system pathological changes. As the range ofcausative drugs can be great, it is difficult to sort out the pattern of pathological changes. Inaddition, there is no significant change in its histopathological changes, and so identifyingthe culprit drug can be extremely difficult. The diagnosis methods mainly include threetypes: drug provocation test(DPT), the skin test and in vitro test. Among them, the first twoare mainly diagnosed in vivo test. DPT remains the gold standard for the identification of aneliciting drug. However, it is usually not accepted by patients as they must take great riskswhich limits the clinical application. The skin test include sprick test, intradermal testsand patch test,which often do not yield positive results even in patients with a clear historyof DTH and get great possibilities of the false positive and the false negative, the skin test is also not widely applied. In vitro test includes drug specific lymphocyte transformationtest(LTT), enzyme-linked immunospot (ELISPOT), the flow cytometric lymphocyteactivation test(LAT), the cytotoxicity test, which brings no risk to patients, and has highsensitivity and specificity. Thus, it is an effective way to diagnose drug hypersensitivityreaction.Serum specific IgE assays are still the most common in vitro methods for evaluatingtype I allergy reaction. And there is no relevant report about applying for type IV drugallergy. Some researches indicated that the T cell play a crucial role on DTH. T cell duringits stereotypia, the multiplication and the function differentiations’ different period willexpress the different superficial molecules, secrete the different cell factor. And the aboveresults can facilitates the better understanding of the T cell’s activation situation. Anddrug-allergic T cell surface member CD69, CD107a have the high specificity and sensitivity.We gather peripheral blood from20patients with DTH and from10healthy control,examine CD69and the CD107a expression, and evaluate their relevance with DTH.Methods:Peripheral blood of20patients and10healthy controls were collected from May2010to April2011in this study. The20patients suffering for exanthematous eruption, AGEP,FDE, TEN, MPEor SJS. Peripheral blood mononuclear cells (PBMCs) were isolated fromperipheral blood by Ficoll density gradient centrifugation. PBMCs were resuspended at2×10~6cells/ml and incubated with culprit drugs. After72hours, cells were washed twice inice-cold phosphate buffered saline (PBS) and stained for30minutes with anti-humanPE-CD107a, FITC-CD69, the expression of CD69and CD107a was analyzed by flowcytometry. And the SPSS13.0system was applied to analyze the data.Results:1.There are20cases of DTH,17cases with antibiotic-allergies,2cases withnon-steroidal anti-inflammatory allergies,1case with carbamazepine-allergy,7patients hada FDE,3patients exanthematous eruption,2multiformity erythema,4cases MPE(includingone case with breathing difficulties, and one case with breathing difficulty, palpitations,sweating),2cases of AGEP,1case of SJS, and1case of exfoliative dermatitis. The maincausative drugs include antibiotics and antipyretic analgesics; and the common drug allergy types include FDE, MPE, and exanthematic eruptions.2.For control group, there was no significant difference among the expression ofCD107a+,CD69~+,CD107a~+CD69~+on PBMC before or after PBMCs were stimulated. Befores t i m u l a t i o n, t h e f i g u r e s a r e C D107a+(0.41±0.18)%, C D69+(0.33±0.13)%,CD107a~+CD69~+(0.12±0.06)%. After stimulation, the figures are CD107a~+(0.38±0.28)%, C D69~+(0.35±0.12)%, C D107a+C D69+(0.14±0.08)%.3. Before PBMCs were stimulated, there was no significant difference among theexpression of CD107a~+,CD69~+,CD107a~+CD69~+on PBMC of healthy control group andpatients with DH. After stimulation, the expression of CD107a~+,CD69~+,CD107a~+CD69~+onPBMCs of patients with DH were significantly increased, whereas the frequency of thesecells on PBMCs of healthy control group was not changed. There was significant differencebetween culprit drugs stimulated group of patients and un-stimulated group of patients orhealthy control group (P<0.01).4.For patients with DH, we selected cefixime-allergic patients and sulfamethoxazoleto accept the stimulation of PBMCs by culprit drugs, and examine the expression ofCD107a, CD69after72hours. The findings are: after stimulation, the expression ofCD107a and CD69is significantly up-regulated, with the proportion of CD107a~+, CD69~+CD107a+and CD69~+cell significantly increased. And with the increase of theconcentration of culprit drugs, the expression tends to increase.Conclusion:Culprit drugs-stimulated PBMC of patients with DTH induce the expression of CD69and CD107a. And with the increase of the concentration of culprit drugs, the expressiontends to increase. And it is expected to become the biological marker of in vitro detection ofallergenic drugs.