Expression Of NLRP3 Inflammasome In Keloids And Expression Difference After Anti-inflammatory Therapy

Purpose1.To observe the expression differences of NLRP1,NLRP3,ASC,Caspase-1,IL-1β in normal skin,single keloid and multiple keloids.2.To investigate the differences of NLRP3 inflammasome expression and the activation of NLRP3-(Caspase-1)/IL-1β pathway in keloid lesions before and after glucocorticoid injection and superficial X-ray radiotherapy and other anti-inflammatory therapy.Methods1.Skin tissues of normal skin,single keloid and multiple keloid groups were collected,with 3 cases of each type of tissue collected.RNA-sequencing was performed on the collected skin tissues to detected the expressions of NLRP1,NLRP3,ASC,Caspase-1 and IL-1β in normal skin,single keloid and multiple keloids.The results of RNA-sequencing were verified by immunohistochemistry.2.Lesions of 5 patients with keloids who were treated with glucocorticoid injection and superficial X-ray radiation and other anti-inflammatory therapy were collected before and after treatment.Immunohistochemistry,qRT-PCR and western blotting(WB)were used to detect the differences in the expression of NLRP3,ASC,Caspase-1 and IL-1β in multiple keloids lesions of the 5 patients before and after treatment.Results1.Differences in the expression of NLRP1,NLRP3,ASC,Caspase-1,IL-1β in normal skin,single keloid and multiple keloids:RNA-sequencing results found that NLRP3 expression was overexpression in keloid compared with normal skin,and ASC and Caspase-1 were down-regulated in keloid compared with normal skin(P<0.05),and there were no significant differences in the expressions of NLRP1 and IL-1β.There were no significant differences in the expressions of NLRP1,NLRP3 and ASC between the multiple keloids and the single keloid.Compare to multiple keloids the expressions of Caspase-1 and IL-1β were lower in the single keloid(P<0.05).Immunohistochemistry showed that the number of NLRP3 positive fibroblasts in the dermis of normal skin(10.53±6.84)was lower than that of keloid(44.57±13.52)(P<0.05).There was no significant difference in NLRP3 positive fibroblasts between multiple keloids(55.54±9.39)and single keloid(33.60±6.09)(P>0.05).2.Expression differences of NLRP3,ASC,Caspase-1 and IL-1β in keloids lesions before and after treatment:Immunohistochemistry showed that the number of NLRP3 positive fibroblasts in keloid after treatment(27.60±10.57)was significantly lower than that before treatment(48.72±13.23)(P<0.05).qRT-PCR results showed that mRNA expressions of NLRP3,ASC,Caspase-1 and IL-1β in keloid tissues were decreased after treatment,but the differences were not statistically significant(P>0.05).The results of WB showed that the expression of NLRP3 in keloids decreased significantly after treatment,but there was no statistical significance(P>0.05);The expression of Caspase-1 and IL-1β decreased significantly after treatment,and the differences were statistically significant(P<0.05).There was no significant difference in the expression of ASC before and after treatment(P>0.05).Conclusion1.In keloid tissues the expression of NLRP3 was up-regulated,but there was no difference in NLRP1 expression,and there was no significant difference in the expression of NLRP1 and NLRP3 between single keloid and multiple keloids;2.The inhibition of NLRP3-(Caspase-1)/IL-1β signaling pathway in keloid lesions is related to the inhibition of inflammatory response.