Alterations of the Gut Microbiota in Patients with Diabetic Nephropathy

Abstract

Diabetic nephropathy (DN) is the primary cause of end-stage renal disease. Accumulating studies have implied a critical role for the gut microbiota in diabetes mellitus (DM) and DN. However, the precise roles and regulatory mechanisms of the gut microbiota in the pathogenesis of DN remain largely unclear. In this study, metagenomics sequencing was performed using fecal samples from healthy controls (CON) and type 2 diabetes mellitus (T2DM) patients with or without DN. Fresh fecal samples from 15 T2DM patients without DN, 15 DN patients, and 15 age-, gender-, and body mass index (BMI)-matched healthy controls were collected. The compositions and potential functions of the gut microbiota were estimated. Although no difference of gut microbiota α and β diversity was observed between the CON, T2DM, and DN groups, the relative abundances of butyrate-producing bacteria (Clostridium, Eubacterium, and Roseburia intestinalis) and potential probiotics (Lachnospira and Intestinibacter) were significantly reduced in T2DM and DN patients. Besides, Bacteroides stercoris was significantly enriched in fecal samples from patients with DN. Moreover, Clostridium sp. 26_22 was negatively associated with serum creatinine (P < 0.05). DN patients could be accurately distinguished from CON by Clostridium sp. CAG_768 (area under the curve [AUC] = 0.941), Bacteroides propionicifaciens (AUC = 0.905), and Clostridium sp. CAG_715 (AUC = 0.908). DN patients could be accurately distinguished from T2DM patients by Pseudomonadales, Fusobacterium varium, and Prevotella sp. MSX73 (AUC = 0.889). Regarding the potential bacterial functions of the gut microbiota, the citrate cycle, base excision repair, histidine metabolism, lipoic acid metabolism, and bile acid biosynthesis were enriched in DN patients, while selenium metabolism and branched-chain amino acid biosynthesis were decreased in DN patients. IMPORTANCE Gut microbiota imbalance is found in fecal samples from DN patients, in which Roseburia intestinalis is significantly decreased, while Bacteroides stercoris is increased. There is a significant correlation between gut microbiota imbalance and clinical indexes related to lipid metabolism, glucose metabolism, and renal function. The gut microbiota may be predictive factors for the development and progression of DN, although further studies are warranted to illustrate their regulatory mechanisms.

Keywords: composition; diabetic nephropathy; function; gut microbiota; metagenomics.

FIG 1

Gut microbiota compositions of the CON, T2DM, and DN groups. (A) Venn diagram of gene numbers in the CON, T2DM, and DN groups. (B) Wilcoxon rank sum test of gene numbers in CON, T2DM, and DN patients. (C) Gut microbiota composition at the phylum level of each sample. (D) Gut microbiota composition at the phylum level. (E) Gut microbiota composition at the genus level. (F) Gut microbiota composition at the species level.

FIG 2

Diversity of the gut microbiota in the CON, T2DM, and DN groups. (A to C) Chao, Shannon, and ACE indexes of gut microbiota α diversity in the CON, T2DM, and DN groups. (D to F) PCoA analysis of gut microbiota β diversity in the CON, T2DM, and DN groups at the phylum, genus, and species levels.

FIG 3

Differential species of gut microbiota between the CON, T2DM, and DN groups. (A) Annotated branch diagram of the different species of the gut microbiota in CON, T2DM, and DN patients. (B) LEfSe analysis showing the most differentially abundant taxa between the CON, T2DM, and DN groups. Red, blue, and green bars represented species with relatively higher abundance in CON, T2DM, and DN patients, respectively. Only species with an LDA  of >2 are shown. (C) Increased or decreased strains in the CON, T2DM, and DN groups. *, P < 0.05, compared with CON group; #, P < 0.05, compared with T2DM group.

FIG 4

Spearman correlation analysis regarding the association between the differential species and clinical indexes. Red and blue indicate positive and negative correlations, respectively. *, P < 0.1; **, P < 0.05; ***, P < 0.005).

FIG 5

Gut microbiota biomarkers distinguishing the DN group from the CON and T2DM groups. (A) Random forest analysis of distinguishing species between the DN and CON groups, showing the 15 species with the highest contributions. (B) ROC curve differentiating the DN group from the CON group based on species with excellent effect. (C) Random forest analysis of distinguishing species between the DN and T2DM groups, showing the 15 species with the highest contributions. (D) ROC curve classifying the DN group from the T2DM group, based on species with excellent effect.

FIG 6

Function of the gut microbiota. (A) Gene numbers of KEGG classification in gut microbiota. (B) PCoA of gut microbiota function between the CON, T2DM, and DN groups. (C) ANOSIM analysis of gut microbiota function between the CON, T2DM, and DN groups. (D) LEfSe analysis of gut microbiota functions between the CON and DN groups. (E) LEfSe analysis of the gut microbiota functions between the T2DM and DN groups. (F to I) Gut microbiota function with a decreasing trend between the CON, T2DM, and DN groups.