The Mechanism Of DC Differentiation Inhibited By UVB

Objective:As complement iC3b has been identified abundantly at the dermis-epidermis junction, and furthermore our previous study has demonstrated inhibition of monocyte-derived dendritic cell differentiation and interleukin-12 production by iC3b via a mitogen-activated protein kinase (MAPK) signaling pathway. The present study was thus supposed to investigate the roles of ERK1/2 and p38 MAPK cascades in the development of iC3b-coated-Mo into CD1a+DC, furthermore to test whether the disappearance of CD1a+DC can be reversed via their inhibitors, in addition, to explore the role of iC3b-coated-Mo with inhibitors pretreatment upon CD4+T proliferation compared with normal Mo.Methods:1) Mo was isolated with CD14+microbeads first, following incubation with EAiC3b (IgM-plus iC3b-coated sheep erythrocyte), IL-4 and GM-CSF for 2 days, then followed by detections of expression of CD14+and CD1a+via Flow cytometry, expression of phoso-ERK1/2 MAPK and phoso-p38 MAPK via Western blot, expression of IL-10 and IL-12 p70 via ELISA, compared with EA.2) When pretreatment with PD98059 (inhibitor for ERK1/2 MAPK) or SB203580 (inhibitor for p38 MAPK) Mo, prior to EaiC3b, also following 2-day incubation with IL-4 and GM-CSF, expression of CD 14+, CD1a+ via Flow cytometry, expression of phoso-ERK1/2 MAPK, phoso-p38 MAPK via Western blot, and expression of IL-10, IL-12 p70 via ELISA were all detected.3) PD98059 (or SB203580) pretreatment with Mo prior to EAiC3b, with GM-CSF and IL-4 incubated for 6 days, then cells were irradiated by 60Co as stimulating cells. CD4+T cells were isolated as response cells, and incubated with stimulating cells for 5 days. Then proliferation of CD4+T cells was detected via 3H-TdR. The controls were treated simultaneously.Results:Compared with EA results, maturation of CD1a+DC was inhibited by EAiC3b, that is, expressions of CD1a, phoso-p38 MAPK, IL-12p70 down-regulated, expressions of phoso-ERK1/2 MAPK, IL-10 up-regulated. When PD98059 pretreated, compared with the EAiC3b results, the inhibited maturation of imDC was reversed prominently, that is, expressions of CD1a, IL-12p70 up-regulated, and expressions of phoso-ERK1/2 MAPK, IL-10 down-regulated. When SB203580 pretreated, in contrast to the EAiC3b results, the inhibition of imDC maturation was enhanced, that is, expressions of CD1a, phoso-p38 MAPK, IL-12p70 down-regulated, and IL-10 level light up-regulated. Interestingly, the result of CD4+T proliferation (dpm) in iC3b-combined-Mo decreased dramatically compared with EA result. When PD pretreated, the result of CD4+T proliferation (dpm) was reversed obviously compared with EAC result. When SB pretreated, the result of CD4+T proliferation (dpm) showed enhanced inhibition compared with EAC result.Conclusion:1) Deposit of iC3b induced by UVB plays an important role in the disappearance of CDla+DC via ERK1/2 MAPK cascade; 2) pretreatment of the inhibitor PD98059 for ERK1/2 pathway could reversed the phenomenon dramatically, showing the promotion of imDC maturation, whereas the p38 MAPK specific inhibitor SB203580 pretreatd, the inhibition of imDC maturation was enhanced.3) Mo with iC3b pretreatment showed obvious downregulating potential upon CD4+T proliferation (dpm) compared with normal Mo. When PD pretreated, the potential could reversed to a significant extent.Objective:to explore the role of ERK1/2 MAPK in the mechanism of local UVB-induced immunosuppression, and to test the protective role of its specific inhibitor PD98059 against the phenomenon for the potential of suncreen design. In addition, to explore the anti-inflammatory effect of the p38 MAPK specific inhibitor SB203580 based on the mouse model of contact hypersensitivity.Methods:The mouse model of the local UVB-induced immuno—suppression was established first, that is,8 kJ/m2 of UVB was irradiated on the abdomen depilated skin on Balb/c mouse.3 days later the same area was sensitized by 25μl of 0.5% DNFB. Another 5 days later the baseline ear thickness of the right ear was measured and the dorsal skin of the same ear was challenged by 10μl of 0.2% DNFB. After 24 hours, the ear thickness was remeasured for ear swelling response and the same ear was biopsied for pathological HE stainings. PD study:based on the establised mouse model as described above, PD98059 was applied 1 hour before UVB exposure (group A), simultaneously (group B), and 6 hours after UVB exposure (group C) repectively. Evaluations were by ear swelling response and pathological HE staining. SB study:SB203580 was applied on the abdomen depilated skin on Balb/c mouse first. After 6 hours the same area was sensitized by 25μl of 0.5% DNFB.5 days later the baseline ear thickness of the right ear was measured and the dorsal skin of the same ear was challenged by 10μl of 0.2%DNFB. Another 24 hours later the ear thickness was remeasured for ear swelling response and the same ear was biopsied for pathological HE staining. All the controls were treated simultaneously.Results:PD study:based on the mouse model of local UVB-induced immunosuppression, PD98059 application 6 hours after UVB exposure (group C) showed reversal of immunosuppression, that is, ear swelling response is statistically significant compared with the control group P< 0.05), dermis thicken and edema with infiltration of lymphocytes. Group A and B showed no singificant. SB study:topical pretreatment with different doses of SB203580 based on the contact hypersensitivity mouse model failed to alleviate the degree of contact hypersensivity which were evaluated by ear swelling response and pathological HE staining, compared with the controls.Conclusion:ERK1/2 MAPK pathway plays a critical role in the nature of UVB-induced immunosuppression and its specific inhibitor PD98059 can reverse the immunosuppression prominently, and furthermore, PD98059 can protect against UVB-induced immunosuppression. The anti-inflammatory effect of the p38 MAPK specific inhibitor SB203580 based on contact hypersensivity was not observed in the present study.