Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Suppression of LPS-induced transcription and cytokine secretion by the dietary isothiocyanate sulforaphane

ABSTRACT: Consumption of a diet rich in cruciferous vegetables is associated with decreased risk of developing non-communicable chronic disease. It has been proposed that biologically active isothiocyanates (ITCs) obtained from these vegetables, such as sulforaphane (SF) derived from broccoli, are responsible for mediating health benefits via multiple mechanisms of action. ITCs have been shown to suppress the levels of several LPS-induced pro-inflammatory cytokines that are biomarkers for chronic inflammatory conditions, and which have been associated with the development of both cancer and cardiovascular disease. In patients suffering from chronic inflammatory diseases, LPS is elevated in circulation to levels reaching around 1ng/ml. In the current study, we demonstrate that SF suppresses global changes in gene expression induced by LPS exposure (1ng/ml) in human THP-1 cells at a physiologically achievable concentration of 5 micromolar. These results illustrate the profound effect that SF can have at concentrations achievable through the consumption of broccoli in suppressing TLR4-mediated inflammatory pathways.

ORGANISM(S): Homo sapiens

SUBMITTER: Danielle Folkard

PROVIDER: E-MEXP-3931 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Cruciferous vegetable consumption has been associated with a decreased risk of multiple types of cancers, thus presenting a cost-effective, non-pharmacological approach to cancer prevention through dietary intervention. Broccoli sprouts and Brussels sprouts are among the leading cruciferous vegetables under study and contain some similar and some distinct phytochemicals which can activate different, but complementary, mechanisms to promote health. While the cancer-preventative effects of cruciferous vegetables are typically attributed to glucosinolates and their metabolic products, isothiocyanates and indoles, other components of cruciferous vegetables could play a synergistic role in conferring cancer-protective and health promoting effects. Additionally, metabolism of phytochemicals from cruciferous vegetables by the gut microbiome could further lead to the production, inactivation, or clearance of bioactive dietary components. The gut microbiome is essential to the production of bioactive compounds from various food sources. For example, with soy isoflavones and pomegranate urolithins, the presence or absence of specific microbial taxa directly dictates which metabolites are produced (resulting in a metabotype). A similar paradigm could be extended to cruciferous vegetables in which the gut microbiome may play an important role in driving inter-individual metabolism of glucosinolates and isothiocyanates. We recently reported (Bouranis et. al, 2021, Nutrients) that the gut microbiome composition can influence production of glucosinolate-derived nitriles from cruciferous vegetables, showing that the presence or absence of specific microbes can influence the abundance of a single metabolite. Thus, we sought to take an untargeted approach to investigate other phytochemicals from cruciferous vegetables which the gut microbiome could play a role in generating. To investigate plant- and microbe-derived metabolites of cruciferous vegetable digestion and capture information about the microbiome, we utilized an ex vivo fecal incubation system. Broccoli sprouts and Brussels sprouts were in vitro digested using an oral, gastric, and intestinal phase. For fecal bacterial cultivation a 20% fecal slurry (w/v) was made from fecal material from 10 healthy volunteers (6 female, and 4 male, age 17-51, Lee Biosolutions) and sterile PBS (0.1 M pH 7). 500 µL of fecal slurry was mixed with 10 mL of Brain Heart Infusion Broth (BHI) with hemin and vitamin K, per the manufacturer’s recommendation, and either 500 µl of filter sterilized in vitro digested broccoli sprouts (Broc), 500 µL of filter sterilized in vitro digested Brussels sprouts (Brus), 500 µL of Broc and 500 µL of Brus were added (Combo) or a negative control in vitro digestion (NC). NC contained reverse osmosis water, equivalent in volume to the water content of broccoli sprouts and underwent the same in vitro digestion procedure as described above with the same enzymes, chemicals and equipment. Broc and Brus digests were scaled to be equivalent in concentration to a human consuming ½ cup of broccoli or Brussels sprouts, or in the case of the combination, ½ cup of broccoli sprouts and ½ cup of Brussels sprouts. This combination was included as Broc and Brus contain many similar but also some distinct phytochemicals and thus by combining the vegetables we increased the dose and broadened the range of phytochemicals from cruciferous vegetables which can be achieved in the kitchen as a mixed vegetable dish. Fecal cultures were incubated at 37°C for 24 h in anaerobic conditions.

Project description:Purpose: The goals of this study are to analyze the transcriptome of five time point in broccoli seed germination and sprout development and to find the putative glucosinolate metabolism genes in the stage. Methods: Total mRNA of germinated seeds, 3 day cotyledons, 7 day botyledons, 11 day cotyledons and 11 day euphyllas of wild-type broccoli were harvested. Each sample was harvested in three independent biological replicates with equal weight and subsequently pooled together for sequencing. The sequence reads that passed quality filters were de novo assembled using VELVET followed by OASES. Then the assembled unigenes were used for the abundance and functional analysis. Results: A total of ~85million 251bp reads were obtained. After de novo assembly and searching the assembled transcripts against the Arabidopsis thaliana and Nr databases, 19,441 top-hit transcripts were clustered as unigenes with an average length of 2,133bp. These unigenes were classified according to their putative functional categories. Cluster analysis of total unigenes with similar expression patterns and differentially expressed unigenes among different tissues,as well as transcription factor analysis were performed. We identified 25 putative glucosinolate metabolismgenes sharing 62.04-89.72% nucleotide sequence identity with the Arabidopsis orthologs. This established a broccoli glucosinolate metabolic pathway with high colinearity to Arabidopsis. Many of the biosynthetic and degradation genes showed higher expression after germination than in seeds; especially the expression of the myrosinaseTGG2 was 20-130 times higher.These results along with the previous reports that glucosinolate concentration decreased exponentially once after germination indicate the breakdown products of glucosinolates may play important roles in broccoli seed germination and sprout development. Conclusion: Our study provides the largest genetic resource of broccoli to date. These data will pave the way for further studies and genetic engineering of broccoli sprouts to develop functional vegetables containing high levels of the anticarcinogenic glucosinolates. They will also provide new insight into the genomic research of this species and its relatives.

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Suppression of LPS-induced transcription and cytokine secretion by the dietary isothiocyanate sulforaphane

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