Hyperglycemia-induced P300/CBP acetyltransferase drives ZEB2-mediated proinflammatory macrophages and delays wound healing
Soumyajit Roy, Debarun Patra, Palla Ramprasad, Shivam Sharma, Parul Katiyar, Ashvind Bawa, Kanhaiya Singh, Kulbhushan Tikoo, Suman Dasgupta, Chandan K. Sen, Durba Pal
Soumyajit Roy, Debarun Patra, Palla Ramprasad, Shivam Sharma, Parul Katiyar, Ashvind Bawa, Kanhaiya Singh, Kulbhushan Tikoo, Suman Dasgupta, Chandan K. Sen, Durba Pal
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Research Article Dermatology Immunology Inflammation

Hyperglycemia-induced P300/CBP acetyltransferase drives ZEB2-mediated proinflammatory macrophages and delays wound healing

Abstract

Chronic hyperglycemia changes the expression of various transcription factors and mRNA transcripts that impair cellular functionality and delay wound healing. Zinc finger E-box–binding homeobox 2 (ZEB2), a key transcription factor, maintains tissue-specific macrophage identities; however, its role in regulating macrophage polarization during wound healing under hyperglycemic conditions remains unclear. Here, we found that persistent hyperglycemia increases ZEB2 expression in wound macrophages via histone acetylation, contributing to chronic inflammation and delayed wound healing. Exposure to high glucose levels activated P300/CBP, a transcriptional coactivator involved in histone acetylation, which enhanced ZEB2 expression in wound macrophages. The forced expression of ZEB2 shifted macrophage polarity toward a proinflammatory state by upregulating myeloid lineage–directed transcription factors. Conversely, silencing Zeb2 at the wound site reduced hyperglycemia-induced macrophage inflammation. Topical application of C646, an inhibitor of P300, at the wound edges of streptozotocin-induced high-fat diet–fed diabetic mice significantly decreased ZEB2 expression, reduced inflammation, and accelerated wound healing. Therefore, targeted inhibition of P300 represents a promising therapeutic strategy for improving diabetic wound healing by modulating ZEB2-driven inflammation in wound macrophages.

Authors

Soumyajit Roy, Debarun Patra, Palla Ramprasad, Shivam Sharma, Parul Katiyar, Ashvind Bawa, Kanhaiya Singh, Kulbhushan Tikoo, Suman Dasgupta, Chandan K. Sen, Durba Pal

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Figure 5

Inhibition of acetyltransferase activity ameliorated proinflammatory burden in hyperglycemic macrophages, even in diabetic wounds.

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Inhibition of acetyltransferase activity ameliorated proinflammatory bur...
Effect of C646 was assessed by (A) HAT1 and P300 by immunostaining (scale bars: 30 μm, n = 3) and (B) Western blot of ZEB2 and p-NF-κB (n = 3) in RAW264.7 cells upon C646 incubation (20 μM) in HG condition. **P < 0.01, #P < 0.0001 by paired, 2-tailed Student’s t test. (C) Inflammatory status was measured by immunostaining of C646 + HG–treated RAW264.7 cells with iNOS and ARG1 antibodies (scale bars: 30 μm, n = 3). **P < 0.01 by paired, 2-tailed Student’s t test. (D) ChIP fold enrichment was assessed by ChIP-PCR for H3K9Ac and H3K27Ac activity at the ZEB2 promoter in PG-, vehicle + HG–, and C646 + HG–treated cells (n = 3). (E) Cytokine expression profile in culture media was analyzed for IL-1β and TNF-α by ELISA (n = 3). *P < 0.05, ***P < 0.001, #P < 0.0001 by 1-way ANOVA followed by Tukey’s post hoc test. (F) Photographs of wound healing at different time points in SD, HFD + vehicle, and HFD + C646 groups (n = 3 mice/group). (G) H&E staining of wound images (scale bars: 1000 μm, n = 3 mice/group) and healing curve showing of original wound size versus time in days (n = 3 mice/group). **P < 0.01, **P < 0.01 by paired, 2-tailed Student’s t test. (H) Immunostaining of mouse wound tissue with ZEB2 (red) and F4/80 (green) antibodies in HFD group treated with or without C646 (10 μg/wound) (scale bars: 100 μm [higher magnification] and 500 μm [lower magnification]; n = 3 mice/group). *P < 0.05 by paired, 2-tailed Student’s t test. Inflammatory status was assessed by immunostaining for (I) TNF-α in d7 and d10, (J) IL-1β in d7, and (K) iNOS in d10 mouse wound tissue (scale bars: 200 μm, n = 3 mice/group). The remodeling phase was examined by immunostaining for (L) αSMA in d7 and (M) CK14 in d10 mouse wound tissue (scale bars: 200 μm, n = 3 mice/group). Data are expressed as mean ± standard deviation. NS, not significant; SD, standard diet; HFD, high-fat diet; PG, physiological glucose level; HG, hyperglycemia.