Kisspeptins inhibit human airway smooth muscle proliferation
Niyati A. Borkar, Nilesh Sudhakar Ambhore, Rama Satyanarayana Raju Kalidhindi, Christina M. Pabelick, Y.S. Prakash, Venkatachalem Sathish
Niyati A. Borkar, Nilesh Sudhakar Ambhore, Rama Satyanarayana Raju Kalidhindi, Christina M. Pabelick, Y.S. Prakash, Venkatachalem Sathish
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Research Article Muscle biology Pulmonology

Kisspeptins inhibit human airway smooth muscle proliferation

Abstract

Sex and gender disparity in asthma is recognized and suggests a modulatory role for sex steroids, particularly estrogen. However, there is a dichotomous role for estrogen in airway remodeling, making it unclear whether sex hormones are protective or detrimental in asthma and suggesting a need to explore mechanisms upstream or independent of estrogen. We hypothesize that kisspeptin (Kp)/KISS1R signaling serves this role. Airway smooth muscle (ASM) is a key structural cell type that contributes to remodeling in asthma. We explored the role of Kp/KISS1R in regulating ASM proliferation. We report potentially novel data indicating that Kp and KISS1R are expressed in human airways, especially ASM, with lower expression in ASM from women compared with men and lower in patients with asthma compared with people without asthma. Proliferation studies showed that cleaved forms of Kp, particularly Kp-10, mitigated PDGF-induced ASM proliferation. Pharmacological inhibition and shRNA knockdown of KISS1R increased basal ASM proliferation, which was further amplified by PDGF. The antiproliferative effect of Kp-10 in ASM was mediated by inhibition of MAPK/ERK/Akt pathways, with altered expression of PCNA, C/EBP-α, Ki-67, cyclin D1, and cyclin E leading to cell cycle arrest at G0/G1 phase. Overall, we demonstrate the importance of Kp/KISS1R signaling in regulating ASM proliferation and a potential therapeutic avenue to blunt remodeling in asthma.

Authors

Niyati A. Borkar, Nilesh Sudhakar Ambhore, Rama Satyanarayana Raju Kalidhindi, Christina M. Pabelick, Y.S. Prakash, Venkatachalem Sathish

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Figure 6

KISS1R activation and PDGF-induced proliferation of human ASM cells.

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KISS1R activation and PDGF-induced proliferation of human ASM cells. KIS...
KISS1R agonist, Kp-10, significantly blunted the mitogenic effect on PDGF in ASM cells from individuals with and without asthma, as evaluated by high-contrast bright-field (A) and MTT (B) assays. KI (Kp-234 trifluoroacetate) showed a significant increase in basal ASM proliferation compared with vehicle and did not show any changes in PDGF-induced proliferation in ASM cells from either individuals with asthma or those without. Human ASM cells transduced with KISS1R-specific shRNA had increased basal ASM proliferation compared with those transduced with negative shRNA. Furthermore, we did not observe any significant reduction in PDGF-induced proliferation after Kp-10 treatment in KISS1R shRNA–transduced cells as measured by high-contrast bright-field (C) and MTT (D) assays. Western blot analysis was performed to confirm the transduction efficacy of KISS1R shRNA in ASM cells from individuals with and without asthma (E). The endogenous Kp secretion of ASM was measured by ELISA using conditioned media. The calibration of ELISA was confirmed by a standard curve of Kp (F, top panel). ASM cells from donors with asthma had reduced Kp secretion in conditioned media compared with those from donors without asthma (F, bottom panel). Data are reported as a minimum to maximum of 5 to 7 individual ASM samples from donors with and without asthma and analyzed using 1- or 2-way ANOVA followed by Tukey’s post hoc test or 1-tailed unpaired t test. *P < 0.05, **P < 0.01, ***P < 0.001 vs. vehicle from donors without asthma or negative shRNA vehicle; ###P < 0.001 vs. respective PDGF-exposed group; $P < 0.05, $$P < 0.01, $$$P < 0.001 vs. respective group without asthma or negative shRNA group.