Journal of dermatological science · 2012
Long TSLP transcript expression and release of TSLP induced by TLR ligands and cytokines in human keratinocytes.
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Xie Yang, Takai Toshiro, Chen Xue, Okumura Ko, Ogawa Hideoki
Аннотация
BACKGROUND: Thymic stromal lymphopoietin (TSLP), highly expressed in keratinocytes in atopic dermatitis patients and bronchial epithelial cells in asthma patients, plays a key role in allergic diseases. Two forms of TSLP mRNA (long and short) have been reported.
OBJECTIVE: We compared the expression of the long-form and total TSLP transcripts in primary human keratinocytes.
METHODS: Primary human keratinocytes were stimulated with Toll-like receptor (TLR) ligands, cytokines, and vitamin D receptor agonists. Gene expression was analyzed by quantitative real-time PCR. The amount of TSLP released was measured by ELISA.
RESULTS: PolyI:C (TLR3 ligand), FSL-1 (TLR2-TLR6 ligand) and flagellin (TLR5 ligand) upregulated long-form TSLP expression, which predominantly contributed to upregulation of total TSLP expression. A glucocorticoid or an endosomal acidification inhibitor inhibited the polyI:C-dependent upregulation of total TSLP and the decrease of the total TSLP was due to the decrease of the long-form. An atopic cytokine milieu (TNF-α+IL-4+IL-13) or TNF-α alone also upregulated the long-form. These stimuli also induced the release of TSLP. In contrast, a high concentration of calcitriol (1,25-dihydroxyvitamin D(3), the active form of vitamin D(3)) or its analog MC903 upregulated total TSLP significantly but not the long-form, and did not induce the release of TSLP.
CONCLUSION: TLR ligands or cytokines predominantly upregulate the gene expression of the long TSLP form, which contributes to the TSLP protein production, in primary human keratinocytes. Specific measurement of the long-form rather than total TSLP should be useful for accurate detection of functional human TSLP gene expression.
OBJECTIVE: We compared the expression of the long-form and total TSLP transcripts in primary human keratinocytes.
METHODS: Primary human keratinocytes were stimulated with Toll-like receptor (TLR) ligands, cytokines, and vitamin D receptor agonists. Gene expression was analyzed by quantitative real-time PCR. The amount of TSLP released was measured by ELISA.
RESULTS: PolyI:C (TLR3 ligand), FSL-1 (TLR2-TLR6 ligand) and flagellin (TLR5 ligand) upregulated long-form TSLP expression, which predominantly contributed to upregulation of total TSLP expression. A glucocorticoid or an endosomal acidification inhibitor inhibited the polyI:C-dependent upregulation of total TSLP and the decrease of the total TSLP was due to the decrease of the long-form. An atopic cytokine milieu (TNF-α+IL-4+IL-13) or TNF-α alone also upregulated the long-form. These stimuli also induced the release of TSLP. In contrast, a high concentration of calcitriol (1,25-dihydroxyvitamin D(3), the active form of vitamin D(3)) or its analog MC903 upregulated total TSLP significantly but not the long-form, and did not induce the release of TSLP.
CONCLUSION: TLR ligands or cytokines predominantly upregulate the gene expression of the long TSLP form, which contributes to the TSLP protein production, in primary human keratinocytes. Specific measurement of the long-form rather than total TSLP should be useful for accurate detection of functional human TSLP gene expression.