Journal: Cell death & disease

Article Title: LL37 complexed to double-stranded RNA induces RIG-I-like receptor signalling and Gasdermin E activation facilitating IL-36γ release from keratinocytes.

doi: 10.1038/s41419-025-07537-9

Figure Lengend Snippet: Fig. 3 LL37/Poly(I:C) complexes induce IL-36γ release from keratinocytes independent of inflammasome and GSDMD activity. A HPKs were primed with IL-17A (100 ng/ml) or left unprimed (medium control) for 6 h prior to stimulation with Poly(I:C), Poly(dA:dT), LL37/Poly(I:C) or LL37/Poly(dA:dT) complexes (all 5 μg/ml) for 18 h. Supernatants were analysed for IL-36γ release by ELISA. B, C GSDMD-deficient N/TERT-1 cells (sgGSDMD) were stimulated with IL-17A (100 ng/ml) for 6 h prior to stimulation with LL37/Poly(I:C) complexes (5 μg/ml), VbP (1 μM), or transfected with Poly(I:C) (1 μg/ml) for 18 h. Supernatants were analysed for D IL-36γ or E IL-1β levels by ELISA. D, E HPKs were stimulated with IL-17A (10 ng/ml) and TNF-α (5 ng/ml) for 5 h, followed by inhibitor treatment with SB 203580 (p38i, 20 μM), Belnacasan (Casp1i, 10 μM), or DMSO for 1 h before stimulation with LL37/Poly(I:C) complexes (5 μg/ml), VbP (1 μM), or anisomycin (ANS, 1 μM) for 18 h. Supernatants were measured for D IL-36γ or E IL-1β by ELISA. F HPKs were primed with IL-17A for 5 h, followed by inhibitor treatment with MCC950 or DMSO for 1 h before stimulation with LL37/Poly(I:C) complexes for 18 h. Cells were treated with MCC950 1 h before stimulation. Supernatants were measured for IL-36γ release by ELISA. Data are presented (A–F) as the mean ±S.E.M. of 3 independent experiments and analysed with (A–C, F) two-way ANOVA and Šidák’s or (D, E) two-way ANOVA followed by Dunnett’s multiple comparisons test. *p < 0.05, **p < 0.01, ****p < 0.0001. ns non-significant, Lipo Lipofectamine 2000, M protein marker, sg single-guide RNA.

Article Snippet: HPKs were transfected by the 1 μg/ml HMW Poly(I:C) coupled with biotin (Invivogen) using Lipofectamine 2000, or treated with 1 μg/ml LL37 with and without pre-incubation with 1 μg/ml HMW Poly(I:C) coupled with Biotin (Invivogen).

Techniques: Activity Assay, Control, Enzyme-linked Immunosorbent Assay, Transfection, Marker

Journal: Cell death & disease

Article Title: LL37 complexed to double-stranded RNA induces RIG-I-like receptor signalling and Gasdermin E activation facilitating IL-36γ release from keratinocytes.

doi: 10.1038/s41419-025-07537-9

Figure Lengend Snippet: Fig. 4 LL37/Poly(I:C) complexes activate Caspase-3 and GSDME cleavage in human primary keratinocytes. A, B HPKs were stimulated with LL37/Poly(I:C) complexes (5 μg/ml) or were transfected with Poly(I:C) (1 μg/ml) for indicated times control treatment (−) indicates Medium or Lipo, respectively. Supernatants and cell lysates were subjected to SDS-PAGE and immunoblotting with indicated antibodies. * indicates non- specific band (C, D) GSDME-deficient N/TERT-1 cells (sgGSDME) were (D) primed with IL-17A (100 ng/ml) or (C) left unprimed for 6 h prior to transfection with Lipo/Poly(I:C) (1 μg/ml), stimulation with LL37/Poly(I:C) complexes (5 μg/ml) or one UVB pulse (0.0875 J/cm2) for 18 h. Data are presented as a representative (A, B) of three independent experiments or are presented (C, D) as the mean ±S.E.M. of 3 independent experiments and analysed with two-way ANOVA and Šidák’s multiple comparisons test. **p < 0.01, ***p < 0.001, ****p < 0.0001. ns non- significant, Lipo Lipofectamine 2000, M protein marker, sg single-guide RNA.

Article Snippet: HPKs were transfected by the 1 μg/ml HMW Poly(I:C) coupled with biotin (Invivogen) using Lipofectamine 2000, or treated with 1 μg/ml LL37 with and without pre-incubation with 1 μg/ml HMW Poly(I:C) coupled with Biotin (Invivogen).

Techniques: Transfection, Control, SDS Page, Western Blot, Marker

Journal: Nature Communications

Article Title: Lugdunin amplifies innate immune responses in the skin in synergy with host- and microbiota-derived factors

doi: 10.1038/s41467-019-10646-7

Figure Lengend Snippet: Proposed model of lugdunin-mediated skin protection. Lugdunin acts on different levels to protect against S. aureus skin infection: First it can directly inhibit and kill S. aureus . Secondly, it can cooperate with host-derived antimicrobial peptides (AMPs) such as hCAP18/LL-37 and the dermcidin-derived peptides DCD-1(L) to kill S. aureus . Additionally, on a third level of protection, lugdunin induces an innate immune response of the skin, which leads to the recruitment of phagocytic immune cells, which will clear potentially invading pathogens. Finally, this innate immune response can be highly amplified by factors derived from the skin commensal S. epidermidis . Blue, lugdunin; orange, DCD-1(L); multicolored, commensals

Article Snippet: The next day, the plate was washed three times using PBS + 0.5% BSA + 0.05% Triton X-100, followed by incubation with 100 µL primary antibody against hCAP18/LL-37 (HycultBiotech, Cat#HM2071, 1:120 in PBS + 0.5% BSA + 0.05% Triton X-100) at 37 °C for 1 h. After washing, incubation with 100 µL secondary antibody (Cell Signaling, Cat#7076S, 1:3000 in PBS + 0.5% BSA + 0.05% Triton X-100) followed at 37 °C for 1 h. Subsequently, plates were washed and 100 µL TMB substrate solution (Cell Signaling) was added.

Techniques: Infection, Derivative Assay, Amplification