Cold atmospheric plasma and silymarin nanoemulsion synergistically inhibits human melanoma tumorigenesis via targeting HGF/c-MET downstream pathway

Abstract

Background Recent studies claimed the important role of cold atmospheric plasma (CAP) with nanotechnology in cancer treatments. In this study, silymarin nanoemulsion (SN) was used along with air CAP as therapeutic agent to counter human melanoma. Methods In this study, we examined the combined treatment of CAP and SN on G-361 human melanoma cells by evaluating cellular toxicity levels, reactive oxygen and nitrogen species (RONS) levels, DNA damage, melanoma-specific markers, apoptosis, caspases and poly ADP-ribose polymerase-1 (PARP-1) levels using flow cytometer. Dual-treatment effects on the epithelial–mesenchymal transition (EMT), Hepatocyte growth factor (HGF/c-MET) pathway, sphere formation and the reversal of EMT were also assessed using western blotting and microscopy respectively. SN and plasma-activated medium (PAM) were applied on tumor growth and body weight and melanoma-specific markers and the mesenchymal markers in the tumor xenograft nude mice model were checked. Results Co-treatment of SN and air CAP increased the cellular toxicity in a time-dependent manner and shows maximum toxicity at 200 nM in 24 h. Intracellular RONS showed significant generation of ROS (< 3 times) and RNS (< 2.5 times) in dual-treated samples compared to control. DNA damage studies were assessed by estimating the level of γ-H2AX (1.8 times), PD-1 (> 2 times) and DNMT and showed damage in G-361 cells. Increase in Caspase 8,9,3/7 (> 1.5 times), PARP level (2.5 times) and apoptotic genes level were also observed in dual treated group and hence blocking HGF/c-MET pathway. Decrease in EMT markers (E-cadherin, YKL-40, N-cadherin, SNAI1) were seen with simultaneously decline in melanoma cells (BRAF, NAMPT) and stem cells (CD133, ABCB5) markers. In vivo results showed significant reduction in SN with PAM with reduction in tumor weight and size. Conclusions The use of air CAP using μ-DBD and the SN can minimize the malignancy effects of melanoma cells by describing HGF/c-MET molecular mechanism of acting on G-361 human melanoma cells and in mice xenografts, possibly leading to suitable targets for innovative anti-melanoma approaches in the future. Electronic supplementary material The online version of this article (10.1186/s12964-019-0360-4) contains supplementary material, which is available to authorized users.

Medical Gas Plasma Jet Technology Targets Murine Melanoma in an Immunogenic Fashion

“…Novel treatment modalities require both efficacy and safety. Despite reports with other plasma sources suggesting that antitumor effects of gas plasmas are facilitated via DNA damage, [ 99–101 ] we have no indication of our plasma jet being genotoxic. Our previous studies established that plasma treatment did not cause micronucleus formation in vitro [ 102 ] regardless of the feed gas settings [ 103 ] as well as in vivo.…”

Melanoma Growth Analysis in Blood Serum and Tissue Using Xenograft Model with Response to Cold Atmospheric Plasma Activated Medium

“…Hence, for the present study, we selected Balb/c (CAnN.Cg-Foxn1nu) male nude mice (5 weeks; 22 ± 2 gm), which is the most relevant and successful animal model, in part, to our broad knowledge, for transplantation models, including the xenograft of human cancer cells [23]. The animal is kept in sterile conditions with recommended feed and sterile water inside an animal house and sacrificed in an ethical way after finishing the experiments.…”

Dosing: The key to precision plasma oncology

“…[ 16 ] CAP was lately shown to induce apoptosis in melanoma cells via sestrin2‐mediated nitric oxide synthase signaling [ 33 ] and by targeting the HGF/c‐MET downstream pathway. [ 34 ]…”

“…The selectivity of CAP on cancer cells has been reported in various types of cancer cells such as triple‐negative breast cancer, [ 17 ] melanoma, [ 33,34 ] non‐small‐cell lung cancer, [ 42 ] head and neck carcinoma, [ 15 ] and pancreatic cancer [ 13 ] (Table 1). It was stated that malignant cells have higher baseline ROS levels than normal cells due to elevated ROS creation and inefficient redox scavenging.…”

“…CAP and silymarin nanoemulsion were shown to synergistically inhibit human melanoma tumorigenesis with significantly elevated toxicity via targeting HGF/c‐MET downstream pathways both in vitro and in mice xenografts. [ 34 ]…”