CD38 triggers inflammation mediated pyroptotic cell death in head and neck squamous cell carcinoma

Abstract

Pyroptosis is a form of inflammatory cell death with the formation of pores on the plasma membrane, eventually leading to osmotic cell rupture. Gasdermin D (GSDMD) is a pivotal pore-forming protein in pyroptosis, which only functions after cleaved by capase-1. It is recognized that inflammasome formed by NLRP3 (nucleotide-binding domain, leucine-rich repeat-containing family, pyrin domain- containing 3), ASC (apoptotic speck-like protein containing caspase-recruitment domain), and pro-caspase-1 is essential in triggering pyroptosis. At present, the mechanism by which head and neck squamous cell carcinoma (HNSCC) inhibits pyroptotic cell death remains undefined. Moreover, studies point out that calcium mobilization could trigger assembly and activation of the NLRP3 inflammasome complex. Whether calcium is involved in NLRP3 mediated pyroptosis remains elusive. Hence, it is reasonable to speculate that CD38, as a well-documented calcium modulator, is involved in triggering NLRP3 inflammasome mediated pyroptotic cell death. In summary, this study aims to delineate the role of calcium regulator CD38 in NLRP3 inflammasome-dependent pyroptosis in HNSCC. As the role of CD38 in head and neck cancer remains undefined, I first examined the clinical relevance of CD38 in TCGA head and neck cancer datasets and found that patients with low expression of CD38 had a significant reduction in overall and disease-free survival. Next, I performed functional studies using CD38- overexpressing HNSCC cell lines. Reduced proliferation and colony formation ability was observed in HNSCC cells with stable CD38-overexpressing. Animal xenograft model results also confirmed the tumor suppressor function of CD38. To further confirm the role of calcium in NLRP3 inflammasome activation, HNSCC cells were incubated with CaCl2 and calcium chelator. The data indicated that calcium ion is a regulator for triggering inflammasome activation. It is also noted that caspase-1 activity and the dead cells were increased in response to exogenous calcium. The results implied that calcium mobilizing signals might be involved in NLRP3 inflammasome-mediated cell death in HNSCC. Given that CD38 is a Ca2+ modulator, I speculated that the CD38 could play a part in calcium-associated cell death. The results demonstrated that NLRP3 expression, caspase-1 activity, GSDMD-NT, and the proportion of dead cells were significantly increased in CD38-overexpressing HNSCC cells compared with the mock cells. These proved that CD38 induced GSDMD cleavage by activating NLRP3 inflammasome and caspase-1 activity, eventually leading to pyroptotic cell death. ChIP assay indicated

that calcium-sensitive transcription factor NFAT1 was possibly involved in the transcriptional upregulation of NLRP3 observed in CD38-overexpressing HNSCC. In conclusion, our data provide new insights into the putative tumor- suppressing functions of CD38 in HNSCC. CD38 is participating in NLRP3 expression by stimulating transcription via calcium-sensitive transcription factor. Given that CD38 has a profound impact on pyroptosis, further study is warranted to evaluate the use of CD38-mediated signaling cascade for HNSCC treatment.