Abstract
Background and Aims
Dysbiosis, or changes in the gut microbiota composition, had been related to the developed of several pathologies, such as chronic kidney disease. Until now, multiple studies have focused on the influence of diet on outcomes of patients with CKD. These patients with advanced disease are recommended a restricted intake of vegetable fiber due to the phosphorus and potassium levels, and low proteins to avoid the generation of uremic toxins. It is known that dietary changes lead to alterations in gut microbiota, but also in microbial metabolites production, some of which could be beneficial for the host. A recent and exciting area of research has begun to explore the role of microbiota-derived metabolites in the renal physiology. Short-chain fatty acids (SCFA, acetate, propionate and butyrate) are a type metabolite produced from dietary fiber by gut microbiota that enter in the bloodstream leading to distal effects, such as modulation of the immune cells. SCFAs are essential to maintain the permeability of the intestinal epithelial barrier, the metabolic functions and have potent anti-inflammatory effects. The aim of this study was to identify the SCFAs levels during the progression of CKD and determinate the functional role of these metabolites in the renal inflammation.
Method
SFCAs (acetate, propionate and butyrate) levels were determined using gas chromatography-mass spectrometry in fecal samples collected from patients with different stages of CKD (n=60) and age-matched healthy control (n=20). Moreover, common bacterial families were determined by quantitative PCR. Additionally, the in-vitro effect of the three SCFAs was evaluated in the human tubular epithelial cell line HK2 using RNA-seq, specific silencing with siRNAs and histone deacetylases (HDAC) inhibitors. To evaluate the effect in immune cells, monocyte and macrophages were treated with LPS and ATP /Nigericin to induce inflammasome activation.
Results
The SCFAs levels were significantly lower in patients with CKD than in healthy controls, mainly propionate and butyrate. Moreover, these levels progressively decreased with the developed of the disease, showing the patients with stage 5 (CKD5) have the lowest levels that correlates with a lesser abundance of Clostridium IV family. According to the renal function, butyrate levels were positively correlated with the glomerular filtration rate and negatively with the blood urea nitrogen and creatinine levels. Surprisingly, high propionate levels correlate with the most elevated serum calcidiol concentrations. Functionally, propionate and butyrate show a similar pattern in the modulation of inflammatory genes in HK2 cells. Most regulated pathways are associated with Inflammatory response (GO:0006954: IL6, TNF, CCL2, RELB, IRAK2, NFKB1,CCL20) and immune response (GO:0006955: CSF2, CXCL3, CD40, IL7R, LIF). Additionally, both SCFAs regulates the expression of multiple epigenetic enzymes involves in the chromatin remodeling, mainly in histone acetylation. In monocytes/macrophages, propionate and butyrate inhibits the IL1B, CASP, and ASC gene transcription damaging the IL-1β secretion. We determined that the effect of SCFAs in these in-vitro models is mediated by inhibition of HDAC although also change other histone modifications (H3K9me3, H3K27me3) and through the GPR109A receptor.
Conclusion
Our initial results showed that patients with advanced CKD have low levels of SCFAs, and those were correlated with the renal function. Treatment of human renal and immune cells with propionate and butyrate induces profound changes in the chromatin structure, changing the whole-genome gene expression and modulating key pathways in the renal pathology. Increasing the SCFAs levels in those patients could be a potential therapeutic strategy to slow down the disease progression.
The Role of the Gut Microbiome in Diabetes and Obesity-Related Kidney Disease