Transcriptome and metabolome changes induced by bitter melon (Momordica charantia)- intake in a high-fat diet induced obesity model

Abstract

Background and aim: Metabolic syndrome (MetS) is a complex disease of physiological imbalances interrelated to abnormal metabolic conditions, such as abdominal obesity, type II diabetes, dyslipidemia and hypertension. In the present pilot study, we investigated the nutraceutical bitter melon (Momordica charantia L) -intake induced transcriptome and metabolome changes and the converging metabolic signaling networks underpinning its inhibitory effects against MetS-associated risk factors.

Experimental procedure: Metabolic effects of lyophilized bitter melon juice (BMJ) extract (oral gavage 200 mg/kg/body weight-daily for 40 days) intake were evaluated in diet-induced obese C57BL/6J male mice [fed-high fat diet (HFD), 60 kcal% fat]. Changes in a) serum levels of biochemical parameters, b) gene expression in the hepatic transcriptome (microarray analysis using Affymetrix Mouse Exon 1.0 ST arrays), and c) metabolite abundance levels in lipid-phase plasma [liquid chromatography mass spectrometry (LC-MS)-based metabolomics] after BMJ intervention were assessed.

Results and conclusion: BMJ-mediated changes showed a positive trend towards enhanced glucose homeostasis, vitamin D metabolism and suppression of glycerophospholipid metabolism. In the liver, nuclear peroxisome proliferator-activated receptor (PPAR) and circadian rhythm signaling, as well as bile acid biosynthesis and glycogen metabolism targets were modulated by BMJ (p < 0.05). Thus, our in-depth transcriptomics and metabolomics analysis suggests that BMJ-intake lowers susceptibility to the onset of high-fat diet associated MetS risk factors partly through modulation of PPAR signaling and its downstream targets in circadian rhythm processes to prevent excessive lipogenesis, maintain glucose homeostasis and modify immune responses signaling.

Keywords: AMPK, adenosine monophosphate-activated protein kinase; BMJ, bitter melon juice; Bitter melon; DIO, diet-induced obese; Diet intervention; HDL, high density lipoprotein (cholesterol); HFD, high fat diet; HMDB, Human Metabolome Database; High fat diet-induced obesity; KEGG, Kyoto Encyclopedia of Genes and Genomes; LC-MS, liquid-chromatography mass spectrometry; LDL, low density lipoprotein (cholesterol); MetS, Metabolic syndrome; Metabolic syndrome; Momordica charantia; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PPARs, Peroxisome proliferator-activated receptors.

Graphical abstract

Fig. 1

Body and liver weight changes, and biochemical profile in Bitter Melon Juice (BMJ)-treated high fat diet-induced obese (DIO) C57BL/6J mice. A) Relative body weight changes in DIO vs. BMJ + DIO mice. B) Relative liver weights of DIO vs. BMJ + DIO mice (left panel); Representative pictographs (x100) of hematoxylin and eosin (H&E) stained hepatic tissue; magnified images depicted at x400 magnification highlight the appearance of fatty globules/steatosis (right panel). Arrows indicate bulging cells with large globules filled with lipid content; scale bar: 50 μM C) Relative serum levels of metabolic parameters and adipokines in DIO vs. BMJ + DIO mice. BMJ was given for 40 days as oral gavage: 200 mg/kg body wt). DIO, diet-induced obesity; Glc, glucose; TGA, triglycerides; AdipoQ, Adiponectin. ∗∗p < 0.02; ∗p < 0.05. n = 4 per group.

Fig. 2

Volcano plot of differential hepatic gene expression in Bitter Melon Juice (BMJ)-treated high fat diet-induced obese (DIO) C57BL/6J mice. Over 20,000 transcripts were profiled from mouse liver via microarray. A total of 768 genes were differentially expressed by ± 1.2-ratio (p ≤ 0.05). Transcriptome profiling showed 535 were up-regulated (red) and 223 were down-regulated (green) in BMJ + DIO mice compared to DIO controls. n = 4 per group.

Fig. 3

Bitter Melon Juice (BMJ) targeted signaling pathways in Metabolic Syndrome (MetS). Transcriptome and metabolome analyses identified several signaling pathways that contribute to metabolic syndrome (MetS). Major risk factors of metabolic syndrome include cardiovascular disease, obesity, type II diabetes, dyslipidemia, hypertension and vitamin D deficiency. BMJ treatment changed hepatic gene expression (∗p ≤ 0.05) and plasma metabolites (∗FDR≤0.05) involved in circadian rhythmic regulation, peroxisome proliferator-activated receptor (PPAR) signaling, apoptosis, insulin resistance, glycerophospholipid, cholesterol and vitamin D metabolism that affect MetS risk factors (∗FDR≤0.05).

Fig. 4

Network map of predicted metabolite and associated protein of differentially expressed hepatic genes in Bitter Melon Juice (BMJ)-treated high fat diet-induced obese (DIO) C57BL/6J mice. Network mapping of significant lipid phase metabolites [PC(20:4_20:4), PC(22:6_20:4), PE(18:1), PE (22:6), 25-azavitamin D₃, 2alpha-(benzyloxy)-1alpha,25-dihydroxy-19-norvitamin D3,13-Deoxytedanolide, Fertaric acid, Diacylglycerol, and Neurine] detected in the plasma of BMJ + DIO mice, and metabolites (Coenzyme A, l-Carnitine, Inositol 1,4,5-trisphosphate/I3P) associated with human genes (ACOT8, ACOT12, ACSL1, ACY1, CPT1A, DGAT2, HGSNAT, HMGCS1, NAT2, PIGL, PLA2G7, SAT2, SLC22A5, SLC25A20, SLC27A5) were predicted to have associations based on experimental or in silico data using the STITCH database (http://stitch.embl.de). Nodes (sphere) represent a protein and splice isoform or post-translational modification. Experimental and/or predicted interactions between proteins (sphere) and chemical (cubiod) are collapsed in the map. Each node represents all the proteins produced by a single, and protein-coding gene locus. Network edges represent specific protein-protein associations jointly contribute to a shared function-but does not mean proteins physically binding each other. A small node represents that the protein 3D structure is unknown, and a large node represents a partial or full 3D structure is known or predicted. A colored node is the first shell interactor, and a white node is the second shell interactor. Connectors represent predicted associations between proteins and chemicals at high confidence (0.700). A red colored connector indicates the presence of fusion evidence. A green connector indicates neighborhood evidence. A black connector indicates co-expression evidence.

Fig. 5

Scheme depicting Bitter Melon Juice (BMJ) targeted pathways and their implications in Metabolic Syndrome (MetS). The metabolic effect of BMJ in diet-induced obese (DIO) C57BL/6J mice were determined by transcriptomics (n = 4 per group) and metabolomic analyses (n = 3 per group). Transcripts and metabolites differentially expressed in the liver and plasma lipid-phase of BMJ + DIO mice identified several common metabolic mechanisms. In the liver transcriptome and plasma metabolites, BMJ induced expression of genes and modulated levels of metabolites involved in adaptive immunity, steroid (Vitamin D), and glycerophospholipid metabolism. In addition to these pathways, peroxisome proliferator-activated receptor (PPAR), circadian rhythmic, and adipocytokine signaling which are associated with metabolic syndrome (MetS) and associated conditions (shown on the right above) are also modulated by bitter melon. Based on in vitro and in vivo models in the literature, it could be inferred that BMJ may exerts its anti-diabetic, anti-MetS and anti-inflammatory activity and other beneficial health effects via the activation of PPARγ and AMPK signaling.