Biology subjects

Hemshekhar, M.

Publications and source records attributed to Hemshekhar, M..

3 recordsLinked to original sources

bioRxiv2026

LL-37 and Citrullinated-LL-37 elicited Transcriptome and Cytokine Profiles in Human Bronchial Epithelial Cells: Citrullination dampens the inflammatory biosignature

The human host defence peptide LL-37 mediates pleiotropic immunomodulatory functions that can be both pro- and anti-inflammatory. Under inflammatory conditions in the lungs, LL-37 is susceptible to the post-translational modification (PTM) citrullination. The impact of this PTM on LL-37's immunomodulatory functions is not fully understood. Therefore, we characterized the transcriptome and cytokine profile in response to LL-37 and citrullinated-LL-37 (citLL-37), in human bronchial epithelial cells (HBEC). Cells were stimulated with either LL-37 or citLL-37 (0.5 M), or a scrambled peptide (sLL-37). RNA after 4 hours (h) was analyzed with NanoString nCounter Host Response Panel, and 96 cytokines were examined (after 24 h) in tissue culture supernatants (Luminex platform). Genes and cytokines with a Log2 fold-change [≥]0.5 with p<0.05 compared to unstimulated cells were considered differentially expressed (DE). These studies revealed overlapping and distinct biosignatures. There were 118 DE genes with LL-37 and 71 with citLL-37, of which magnitude of change was significantly different for 32 genes. 16 out of 17 DE genes altered by both peptides were directly related to inflammation, and these were significantly less (by ~45%) with citLL-37 compared to LL-37. Similarly, inflammatory cytokines enhanced in response to LL-37 were significantly less (by 37-94%) with citLL-37. Overall, our findings indicate that citrullination of LL-37 does not abrogate cellular response in HBEC, instead dampens the inflammatory biosignature induced by LL-37.

immunology↗

bioRxiv2025

LL-37 and citrullinated-LL-37 enhance disparate oxylipins: LL-37-mediated chemokine response is dependent on COX-2 and the P2X7 receptor in human bronchial epithelial cells

BackgroundDuring airway inflammation, chemokines, oxylipins (bioactive lipids) and cationic host defence peptides (CHDP) are enhanced in the lungs. However, the interplay of these molecules in the process of airway inflammation is not fully resolved. The human cathelicidin CHDP, LL-37, can enhance the expression of chemokines which is turn facilitates influx of leukocytes into the lungs. Moreover, LL-37 can get citrullinated during inflammation and the effect of this post-translational modification on LL-37-mediated immunomodulatory functions remains unclear. Therefore, in this study we aimed to define the impact of LL-37 and citrullinated-LL-37 (citLL-37) on oxylipins and its association with downstream chemokine production in human bronchial epithelial cells (HBEC), and its functional impact on leukocyte influx. MethodsWe used a lipidomics approach to identify oxylipins that are enhanced in response to LL-37 and citLL-37 in HBEC. We further examined the role of selected oxylipins in LL-37- and citLL-37-mediated chemokine production by ELISA, and related leukocyte migration using a transwell migration assay. ResultsWe showed that LL-37, but not citLL-37, enhances oxylipins that are known to promote inflammation such as prostaglandins regulated by the cyclooxygenase (COX pathway). Although both LL-37 and citLL-37 upregulated COX-2, LL-37-mediated increase in COX-2 expression was significantly higher than that mediated by citLL-37. We showed that upregulation of COX-2 expression was dependent on the P2X7 purinergic receptor. Our mechanistic studies revealed that LL-37-mediated increase in chemokines, GRO, IL-8 and MIP-3, was dependent on the COX-2 pathway. Our results also indicated that COX-2-induced PGE2 may act in an autocrine manner signaling through its EP receptors to facilitate LL-37-induced chemokine production. We functionally confirmed that factors secreted from HBEC in response to LL-37, but not citLL-37, promotes neutrophil migration which is COX-2 dependent. ConclusionThe results of this study indicate that pro-inflammatory responses mediated by LL-37 is alleviated by citrullination of the peptide. These findings suggest that citrullination of LL-37 may be a post-translational regulatory mechanism to control inflammation. Overall, this study underscores the role of LL-37 in influencing the enhancement of bioactive lipids and metabolic pathways such as COX-2 and its link to the peptide-mediated immunomodulatory functions in the lungs.

immunology↗

bioRxiv2024

LL-37 modulates IL-17A/F-mediated airway inflammation by selectively suppressing Lipocalin-2.

BackgroundLevels of the human cationic host defence peptide (CHDP) LL-37 is enhanced in the lungs during neutrophilic airway inflammation. LL-37 drives Th17 differentiation, and Th17 cells produce IL-17A and IL-17F which forms the biologically active heterodimer IL-17A/F. While IL-17 is a critical mediator of neutrophilic airway inflammation, LL-37 exhibits contradictory functions; LL-37 can both promote and mitigate neutrophil recruitment depending on the inflammatory milieu. The impact of LL-37 on IL-17-induced responses in the context of airway inflammation remains largely unknown. Therefore, we examined signaling intermediates and downstream responses mediated by the interplay of IL-17A/F and LL-37, in human bronchial epithelial cells (HBEC). As LL-37 can get citrullinated during airway inflammation, we also examined LL-37-mediated downstream response compared to that with citrullinated LL-37 (citLL- 37), in HBEC. ResultsUsing an aptamer-based proteomics approach, we identified proteins that are altered in response to IL-17A/F in HBEC. Proteins enhanced in response to IL-17A/F were primarily neutrophil chemoattractants, including chemokines and proteins associated with neutrophil migration such as lipocalin-2 (LCN-2) and Elafin. We showed that selective depletion of LCN-2 mitigated neutrophil migration, thus functionally demonstrating LCN-2 as a critical neutrophil chemoattractant. We further demonstrated that LL-37 and citLL-37 selectively suppresses IL- 17A/F-induced LCN-2 production, in bronchial epithelial cells. Mechanistic studies revealed that LL-37 and citLL-37 suppressed IL-17A/F-mediated C/EBP{beta}, a transcription factor required for LCN-2 production. In contrast, LL-37 and citLL-37 enhanced the ribonuclease Regnase-1, which is a negative regulator of IL-17 and LCN-2. In an animal model of neutrophilic airway inflammation with elevated IL-17A/F in the lungs, we demonstrated that CRAMP (mouse ortholog of LL-37) negatively correlates with LCN-2. ConclusionsOverall, our findings showed that LL-37 and citLL-37 can selectively suppress the abundance of IL-17A/F-mediated LCN-2, a protein that is critical for neutrophil migration, in bronchial epithelial cells. These results suggest that LL-37, and its modified citrullinated form, has the potential to negatively regulate IL-17-mediated neutrophil migration to control airway inflammation. To our knowledge, this is the first study to report that the immunomodulatory function of LL-37 engages an RNA binding protein, Regnase-1, indicating post-transcriptional regulation of airway inflammation by the peptide.

immunology↗