Gasdermin D-Mediated Neutrophil Pyroptosis drives Inflammation in Psoriasis

Reviewer #1 (Public review):

Summary:

In this study, Liu, Jiang, Diao et.al. investigated the role of GSDMD in psoriasis-like skin inflammation in mice. The authors have used full-body GSDMD knock-out mice and Gsdm floxed mice crossed with the S100A8- Cre. In both mice, the deficiency of GSDMD ameliorated the skin phenotype induced by the imiquimod. The authors also analyzed RNA sequencing data from the psoriatic patients to show an elevated expression of GSDMD in the psoriatic skin.

Overall, this is a potentially interesting study, however, the manuscript in its current format is not completely a novel study.

Strengths:

It has the potential to unravel the new role of neutrophils.

Weaknesses:

The main claims are only partially supported and have scope to improve

Reviewer #2 (Public review):

Summary:

The authors describe elevated GSDMD expression in psoriatic skin, and knock-out of GSDMD abrogates psoriasis-like inflammation.

Strengths:

The study is well conducted with transgenic mouse models. Using mouse-models with GSDMD knock-out showing abrogating inflammation, as well as GSDMD fl/fl mice without neutrophils having a reduced phenotype.

I fear that some of the conclusions cannot be drawn by the suggested experiments. My major concern would be the involvement of other inflammasome and GSDMD bearing cell types, esp. Keratinocytes (KC), which could be an explanation why the experiments in Fig 4 still show inflammation.

Weaknesses:

The experiments do not entirely support the conclusions towards neutrophils.

Specific questions/comments:

Fig 1b: mainly in KC and Neutrophils?

Fig 2a: PASI includes erythema, scaling, thickness and area. Guess area could be trick, esp. in an artificial induced IMQ model (WT) vs. the knock-out mice.

Fig 2d: interesting finding. I thought that CASP-1 is cleaving GSDMD. Why would it be downregulated?

Line 313: as mentioned before (see Fig 1b). KC also show a stron GSDMD staining positivity and are known producers of IL-1b and inflammasome activation. Guess here the relevance of KC in the whole model needs to be evaluated.

Fig 4i - guess here the conclusion would be that neutrophils are important for the pathogenesis in the IMQ model, which is true. This experiment does not support that this is done by pyroptosis.

Author response:

We sincerely appreciate the positive assessment regarding the significance of our study, as well as the valuable suggestions provided by editor and the reviewers.

In response to the reviewers’ comments, we will modify the manuscript to include co-staining of CD66b and GSDMD in the whole skin samples of clinical patients, which will further clarify the expression of GSDMD in neutrophils.

Additionally, we plan to conduct further analyses using publicly available data to elucidate the changes in neutrophil pyroptotic signaling in IMQ-induced psoriatic mice tissue, thereby strengthening our conclusions about the role of neutrophil pyroptosis in the progression of psoriasis.

Moreover, while our research primarily focuses on the role of neutrophil pyroptosis in psoriasis, this does not conflict with existing reports indicating that KC cell pyroptosis also contributes to disease progression. Both studies underscore the significant role of GSDMD-mediated pyroptotic signaling in psoriasis, and the consistent involvement of KC cells and neutrophils further emphasizes the potential therapeutic value of targeting GSDMD signaling in psoriasis treatment. We will expand upon this discussion in the revised manuscript.

In our model, to accurately assess the disease condition in mice, we standardized the drug treatment area on the dorsal side (2*3 cm). Therefore, the area was not factored into the scoring process, and we will include a detailed description of this in the revised manuscript.

Regarding the downregulation of CASP in GSDMD KO mouse skin tissue, existing studies indicate that GSDMD generates a feed-forward amplification cascade via the mitochondria-STING-Caspase axis (PMID: 36065823, DOI: 10.1161/HYPERTENSIONAHA.122.20004). We hypothesize that the absence of GSDMD attenuates STING signaling’s activation of Caspase.

Furthermore, in the revised manuscript, we will address the reviewers’ other comments to enhancing the manuscript quality, such as providing further clarification on relevant issues in the discussion section, refining the key experiments in the methods section, and adding details about the antibodies used, including their associated clones and catalog numbers, as well as including sample sizes (n numbers) in the figure legends.

We believe that the new data and further discussions and clarifications included in the revised manuscript will adequately address all the concerns raised by the reviewers and better support our conclusions.

Finally, we would like to express our gratitude once again to the editor and reviewers for their invaluable feedback on this work!