Alpha-linolenic acid stabilizes HIF-1 α and downregulates FASN to promote mitochondrial apoptosis for mammary gland chemoprevention.

Subhadeep Roy,Lakhveer Singh,Dinesh Kumar,Shreesh Raj Sammi,Uma Devi,Abdulaziz S Saeedan,Rakesh Pandey,Manjari Singh,Atul Kumar Rawat,Rajnish Kumar Yadav,Mohd Nazam Ansari,Swetlana Gautam,Jitendra Kumar Rawat,Gaurav Kaithwas

doi:10.18632/oncotarget.19551

Abstract

Alpha linolenic acid is an essential polyunsaturated fatty acid and is reported to have the anti-cancer potential with no defined hypothesis or mechanism/s. Henceforth present study was in-quested to validate the effect of alpha linolenic acid on mitochondrial apoptosis, hypoxic microenvironment and de novo fatty acid synthesis using in-vitro and in-vivo studies. The IC50 value of alpha linolenic acid was recorded to be 17.55μM against ER+MCF-7 cells. Treatment with alpha linolenic acid was evident for the presence of early and late apoptotic signals along with mitochondrial depolarization, when studied through acridine orange/ethidium bromide and JC-1 staining. Alpha linolenic acid arrested the cell cycle in G2/M phase. Subsequently, the in-vivo efficacy was examined against 7, 12-dimethylbenz anthracene induced carcinogenesis. Treatment with alpha linolenic acid demarcated significant effect upon the cellular proliferation as evidenced through decreased in alveolar bud count, restoration of the histopathological architecture and loss of tumor micro vessels. Alpha linolenic acid restored the metabolic changes to normal when scrutinized through 1H NMR studies. The immunoblotting and qRT-PCR studies revealed participation of mitochondrial mediated death apoptosis pathway and curtailment of hypoxic microenvironment after treatment with alpha linolenic acid. With all above, it was concluded that alpha linolenic acid mediates mitochondrial apoptosis, curtails hypoxic microenvironment along with inhibition of de novo fatty acid synthesis to impart anticancer effects.

Highlights

Α-Linolenic acid (ALA) (18: 3, ω-3) is an essential polyunsaturated fatty acid (PUFA)
Fluorescence microscopic observations of the ER+MCF-7cells stained with acridine orange/ethidium bromide (AO/EtBr), revealed the presence of early and late apoptotic signals, including chromatin condensation, apoptotic body formation, membrane blebbing and fragmented nuclei
Treatment with ALA increased the uptake of JC-1 in cells, which was visualized through increase in green fluorescence intensity suggesting early apoptotic mitochondrial depolarization (Δψ) (Figure 3)

Summary

Introduction

Α-Linolenic acid (ALA) (18: 3, ω-3) is an essential polyunsaturated fatty acid (PUFA). ALA is the major plant-based PUFA and is found in walnuts, flaxseeds, hemp seeds and their oils. EPA and DHA are obtained from fish oil or derived from plant lipids rich in ALA. It was reported that overiectomized mice produce a significant reduction in tumor growth, when compared with a diet containing no flaxseed oil (rich source of ALA)[9]. PUFAs are the integral component of cell membranes and are susceptible to peroxidation and degeneration causing genetic mutations, a critical mechanism for tumor growth [10].ALA seems to be a promising chemotherapeutic agent with desirable characteristics. Studies have reported anticarcinogenic potential of ALA and ALA being a PUFA can moderate the cell membrane integrity in multiple ways, the mechanistic pathway behind the same is unanswered. Considering the same, the present study was proposed to explore the effects of ALA on mammary gland carcinoma with concomitant efforts to elucidate the possible mechanism beneath the same

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