Cell lines

human pulmonary microvascular endothelial cells

Preparation Method

In the hydrogen peroxide (H₂O₂)-induced oxidative stress in lung cell assay, cells were exposed to 100 μM H₂O₂with WKYMVm treatment. After incubation with WKYMVm for 24 hours in 96-well plates, the cell counting kit (CCK)-8 assay was carried out to determine the relative cell proliferation rate (%), according to the manufacturer’s instructions.

Reaction Conditions

100 μM, 24 h

Applications

In human pulmonary microvascular endothelial cells (HULEC-5a) and primary murine pulmonary endothelial and epithelial cells, 1 and 100 μM WKYMVm treatments significantly increased proliferation in both the control and H₂O₂-exposed groups.

Animal models

male C57/BL6J mice

Preparation Method

A total of 20 male C57/BL6 mice, eight‐week‐old, were split randomly into four equal groups: sham, LPS, low‐dose WKYMVm (4 mg/kg bodyweight) and high‐dose WKYMVm (8 mg/kg bodyweight). The sham and LPS group were injected with PBS and LPS (5 mg/kg bodyweight), respectively. Predetermined WKYMVm doses were mixed with LPS and injected into the mice in combination. Injections were performed subcutaneously against the skull altogether 7 times with 48 hours intervals between applications.

Dosage form

4 mg/kg and 8 mg/kg;s.c.

Applications

WKYMVm protected against LPS‐induced bone loss in vivo.

WKYMVm is a potent N-formyl peptide receptor (FPR1) and FPRL1/2 agonist, and also activate phosphoinositide hydrolysis in undifferentiated HL-60 cells.^[1]

In vitro efficacy test indicated that treatment with 0.01, 0.1, 1 and 10 μmol/L WKYMVm for 24 hours or 48 hours obviouly suppressed RANKL‐induced mature osteoclasts.^[3]In vitro, treatment with 10 μM WKYMVm to stimulate FPR2 can induce p47phox phosphorylation in IMR90 fibroblasts and in human lung cancer cells, which is considered the key event for NADPH oxidase-dependent superoxide generation. Moreover, 10 μM WKYMVm to stimulate also induced NADPH oxidase-dependent superoxide generation with maximal production occurring at 2 min.^[6]

In vivo experiment it shown that treatment with 2.5- and 5 mg/kg/d daily over four days intraperitoneally in ALI mice decreased the levels of proinflammatory cytokines TNF-α, IL-6, and IL-1β, while it increased the MPO and NO release by differentiated HL-60 neutrophil-like cells.^[2].

Intramuscular injection of 10–5 mol/l WKYMVm improves blood perfusion and provides protection from tissue damage in the ischemic hind limb.^[4]In vivo, treatment with 8 mg/kg WKYMVm subcutaneously (at 0, 12, 24, 36, 48 and 60 h) effectively attenuated the DSS-induced increase in the bleeding score and the stool score and protects dextran sodium sulfate (DSS)-induced experimental ulcerative colitis.^[5]In addition, administration of 4 mg/kg WKYMVm, to septic mice strongly increased neutrophil number through augmented emergency granulopoiesis^[7].

References:
[1]. Christophe T, et al. The synthetic peptide Trp-Lys-Tyr-Met-Val-Met-NH2 specifically activates neutrophils through FPRL1/lipoxin A4 receptors and is an agonist for the orphan monocyte-expressed chemoattractant receptor FPRL2. J Biol Chem. 2001 Jun 15;276(24):21585-93.
[2]. Lee H, et al. WKYMVm ameliorates acute lung injury via neutrophil antimicrobial peptide derived STAT1/IRF1 pathway. Biochem Biophys Res Commun. 2020 Dec 10;533(3):313-318.
[3]. Hu J, et al. The protective effect of WKYMVm peptide on inflammatory osteolysis through regulating NF-κB and CD9/gp130/STAT3 signalling pathway. J Cell Mol Med. 2020 Jan;24(2):1893-1905.
[4]. Heo SC, et al. WKYMVm-induced activation of formyl peptide receptor 2 stimulates ischemic neovasculogenesis by promoting homing of endothelial colony-forming cells. Stem Cells. 2014 Mar;32(3):779-90.
[5]. Kim SD, et al. The immune-stimulating peptide WKYMVm has therapeutic effects against ulcerative colitis. Exp Mol Med. 2013 Sep 13;45(9):e40.
[6]. Cattaneo F, et al. WKYMVm-induced cross-talk between FPR2 and HGF receptor in human prostate epithelial cell line PNT1A. FEBS Lett. 2013 May 21;587(10):1536-42.
[7]. Kim HS, et al. Activation of formyl peptide receptor 2 by WKYMVm enhances emergency granulopoiesis through phospholipase C activity. BMB Rep. 2018 Aug;51(8):418-423.