Dietary Supplementation with Naringin Improves Systemic Metabolic Status and Alleviates Oxidative Stress in Transition Cows via Modulating Adipose Tissue Function: A Lipid Perspective

Abstract

Dairy cows face metabolic challenges around the time of calving, leading to a negative energy balance and various postpartum health issues. Adipose tissue is crucial for cows during this period, as it regulates energy metabolism and supports immune function. Naringin, one of the main flavonoids in citrus fruit and their byproducts, is a potent antioxidant and anti-inflammatory phytoconstituent. The study aimed to evaluate the effects of supplemental naringin on performance, systemic inflammation, oxidative status, and adipose tissue metabolic status. A total of 36 multiparous Holstein cows (from ~21 d prepartum through 35 d postpartum) were provided a basal control (CON) diet or a CON diet containing naringin (NAR) at 30 g/d per cow. Supplemental NAR increased the yield of raw milk and milk protein, without affecting dry matter intake. Cows fed NAR showed significantly lower levels (p < 0.05) of serum non-esterified fatty acid (NEFA), C-reactive protein, IL-1β, IL-6, malonaldehyde, lipopolysaccharide (LPS), aspartate aminotransferase, and alanine aminotransferase, but increased (p < 0.05) glutathione peroxidase activity relative to those fed CON. Supplemental NAR increased (p < 0.05) adipose tissue adiponectin abundance, decreased inflammatory responses, and reduced oxidative stress. Lipidomic analysis showed that cows fed NAR had lower concentrations of ceramide species (p < 0.05) in the serum and adipose tissue than did the CON-fed cows. Adipose tissue proteomics showed that proteins related to lipolysis, ceramide biosynthesis, inflammation, and heat stress were downregulated (p < 0.05), while those related to glycerophospholipid biosynthesis and the extracellular matrix were upregulated (p < 0.05). Feeding NAR to cows may reduce the accumulation of ceramide by lowering serum levels of NEFA and LPS and increasing adiponectin expression, thereby decreasing inflammation and oxidative stress in adipose tissue, ultimately improving their systemic metabolic status. Including NAR in periparturient cows' diets improves lactational performance, reduces excessive lipolysis in adipose tissue, and decreases systemic and adipose tissue inflammation and oxidative stress. Integrating lipidomic and proteomic data revealed that reduced ceramide and increased glycerophospholipids may alleviate metabolic dysregulations in adipose tissue, which in turn benefits systemic metabolic status.

Keywords: adipose tissue; naringin; oxidative stress; systemic inflammation; transition dairy cow.

Figure 1

Serum and adipose tissue lipidomes. Principal component analysis (PCA) of (a) serum and (b) adipose tissue lipidomics. Orthogonal partial least squares discriminant analysis (OPLS-DA) of (c) serum and (d) adipose tissue lipidomics. Volcano plot of identified lipid species in (e) serum and (f) adipose tissue. Yellow symbols: significantly decreased lipid species (variable importance in projection (VIP) > 1, p < 0.05). Blue symbols: significantly increased lipid species (VIP > 1, p < 0.05). CON, control; NAR; naringin; FC, fold change.

Figure 2

Lipidomic alteration in serum and adipose tissue samples. Differential lipid species in (a) serum and (b) adipose tissue are presented in the fold-change of NAR relative to CON. KEGG pathway analysis of differential lipid species in (c) serum and (d) adipose tissue samples. BisMePA, bismethyl phosphatidic acid; Cer, ceramide; CL, cardiolipin; Co, coenzyme; DG, diglyceride; Hex2Cer, dihexosylceramide; LPC, lysophosphatidylcholine; MePC, methylphosphatidylcholine; MGDG, monogalactosyldiacylglycerol; OHAFA, (O-acyl)-1-hydroxy fatty acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PI, phosphatidylinositol; PIP2, phosphatidylinositol 4,5-bisphosphate; PS, phosphatidylserine; SiE, sitosteryl ester; SM, sphingomyelin; TG, triglyceride; ZyE, zymosteryl.

Figure 3

Association analysis of the serum and adipose tissue lipidome. (a) The correlation between differential lipid species in the serum and adipose tissue samples by Procrustes analysis; (b) the correlation between differential sphingolipid (SP) species in serum and adipose tissue samples by Procrustes analysis; (c) Pearson’s correlation between serum differential sphingolipids and serum biochemical parameters; (d) Pearson’s correlation between adipose tissue differential sphingolipids and adipose tissue biochemical parameters; (e) Pearson’s correlation between adipose tissue differential glycerophospholipids (top 30 of VIP value) and adipose tissue biochemical parameters. ALT, alanine aminotransferase; ASC, apoptosis-associated speck like protein containing a CARD; AST, aspartate aminotransferase; Cer, ceramide; CL, cardiolipin; CON, control; CRP, C-reactive protein; dMePE, dimethylphosphatidylethanolamine; GSH-Px, glutathione peroxidase; Hex2Cer, dihexosylceramide; IL, interleukin; LPS, lipopolysaccharide; MDA, malonaldehyde; NAR, naringin; NEFA, non-esterified fatty acid; NLRP3, NOD-like receptor thermal protein domain-associated protein 3; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PI, phosphatidylinositol; PS, phosphatidylserine; SM, sphingomyelin; SOD, superoxide dismutase; SP, sphingolipid; TP, total protein.

Figure 4

Adipose tissue proteome analysis. (a) Venn diagram of the identified proteins; (b) PCA score plots of proteome data; (c) volcano plot of the proteome; (d) heatmap of differentially expressed proteins. CON, control; NAR, naringin; FC, fold change.

Figure 5

Proteomic alteration in adipose tissue samples. GO enrichment analysis of (a) upregulated differentially expressed proteins (DEP) and (b) downregulated DEP; (c) KEGG enrichment analysis of DEP; (d) selected DEP related to sphingolipid metabolism and glycerophospholipid metabolism; (e) selected DEP related to lipolysis and inflammation; (f) selected DEP related to heat stress and extracellular matrix. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. CON, control; NAR, naringin.

Figure 6

Integrated pathway and network analysis of the adipose tissue lipidome and proteome. (a) Diagram of lipid metabolism, including glycerolipid metabolism, glycerophospholipid metabolism, and sphingolipid metabolism; (b) protein–protein interaction (PPI) network of selected differentially expressed proteins (DEP); (c) Pearson’s correlation network between differential lipids (glycerophospholipid and sphingolipid species) and selected DEP; red lines represent positive, while blue lines indicate negative correlations (p < 0.05 and |r| > 0.50); (d) Pearson’s correlation analysis between selected DEP and serum biochemical parameters. * p < 0.05, ** p < 0.01. ALT, alanine aminotransferase; AST, aspartate aminotransferase; Cer, ceramide; CL, cardiolipin; CRP, C-reactive protein; dMePE, dimethylphosphatidylethanolamine; GSH-Px, glutathione peroxidase; IL, interleukin; LPS, lipopolysaccharide; MDA, malonaldehyde; NEFA, non-esterified fatty acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PI, phosphatidylinositol; PS, phosphatidylserine; SM, sphingomyelin; SOD, superoxide dismutase; TP, total protein.