Proteomic analysis of milk fat globule membranes from small-sized milk fat globules and their function in promoting lipid droplet fusion in bovine mammary epithelial cells

In mammary epithelial cells, milk fat is synthesized as lipid droplets and secreted in the form of globules. Milk fat globules (MFGs) are covered by a lipid-protein membrane known as the milk fat globule membrane (MFGM). We randomly divided 12 Holstein cows into control and conjugated linoleic acid (CLA) groups. The control group was fed a basal diet, while the CLA group was fed the basal diet + CLA (15 g per kg DM) for 10 days. Cow performance, milk composition, and MFG size were measured daily. On day 10, we extracted MFGM proteins (n = 3) and identified them via quantitative proteomic analysis. We investigated the effects of the MFGM proteins from control and CLA-treated milk on the lipid droplet formation in MAC-T cells. Compared with the control group, the CLA group had reduced milk fat content (3.39 g/100 mL vs. 2.45 g/100 mL) and MFG size parameters (D-[4,D-3] of 3.85 mu m vs. 3.37 mu m; D-[3,D-2] of 3.24 mu m vs. 2.83 mu m). The specific surface area (SSA) increased in the CLA group. A total of 361 differentially expressed proteins were identified in the CLA group by iTRAQ quantitative proteomic analysis. Among these proteins, 100 were upregulated and 251 were downregulated (p < 0.05). In MAC-T cells, CLA-MFGM proteins increased the diameter of the lipid droplets to 1.32 mu m. CLA-MFGM proteins decreased the proportion of the small lipid droplets (15.33% vs. 47.78%) and increased the proportion of the large lipid droplets (25.04% vs. 11.65%). CLA-MFGM proteins promoted lipid droplet fusion. Therefore, MFGM proteins play an important role in the regulation of the lipid droplet size.

Automated summary

Milk fat in mammary epithelial cells is synthesized and secreted as lipid droplets in the form of milk fat globules (MFGs) covered by a lipid-protein membrane called the milk fat globule membrane (MFGM). This study investigated the effects of MFGM proteins from control and conjugated linoleic acid (CLA)-treated milk on lipid droplet formation in MAC-T cells. Results showed that CLA reduced milk fat content and MFG size parameters, while increasing the specific surface area (SSA). Proteomic analysis identified differentially expressed proteins in the CLA group, and CLA-MFGM proteins promoted lipid droplet fusion and influenced droplet size distribution in MAC-T cells.