Elsevier

Gene

Volume 521, Issue 1, 25 May 2013, Pages 15-23
Gene

miR-27a suppresses triglyceride accumulation and affects gene mRNA expression associated with fat metabolism in dairy goat mammary gland epithelial cells

https://doi.org/10.1016/j.gene.2013.03.050Get rights and content

Highlights

  • A correlation exists between miR-27a and milk triglyceride accumulation in goat.

  • miR-27a expression correlates with lactation cycle and prolactin concentration.

  • Over-expression of miR-27a decreases triglyceride and fatty acid contents.

  • Over-expressed miR-27a suppresses the ratio of unsaturated/saturated fatty acid.

  • miR-27a significantly affects gene mRNA expression related to milk fat metabolism.

Abstract

MicroRNAs (miRNAs), a well-defined group of small RNAs containing about 22 nucleotides, participate in various biological metabolic processes. miR-27a is a miRNA that is known to regulate fat synthesis and differentiation in preadipocyte cells. However, little is known regarding the role that miR-27a plays in regulating goat milk fat synthesis. In this study, we determined the miR-27a expression profile in goat mammary gland and found that miR-27a expression was correlated with the lactation cycle. Additionally, prolactin promoted miR-27a expression in goat mammary gland epithelial cells. Further functional analysis showed that over-expression of miR-27a down-regulated triglyceride accumulation and decreased the ratio of unsaturated/saturated fatty acid in mammary gland epithelial cells. miR-27a also significantly affected mRNA expression related to milk fat metabolism. Specifically, over-expression of miR-27a reduced gene mRNA expression associated with triglyceride synthesis by suppressing PPARγ protein levels. This study provides the first experimental evidence that miR-27a regulates triglyceride synthesis in goat mammary gland epithelial cells and improves our understanding about the importance of miRNAs in milk fat synthesis.

Introduction

Goat milk has unique biochemical qualities. Goat milk has higher protein, carbohydrate, and calcium content than cow milk (Hansen et al., 1984, Juarez and Ramos, 1987). Goat milk also contains greater amounts of short-chain fatty acids, medium-chain fatty acids, and unsaturated fatty acids than cow milk (Hansen et al., 1984, Juarez and Ramos, 1987). Some of unsaturated fatty acids (e.g., c9-C18:1, C18:3n-3 and c9,t11-C18:2) are considered good for human health (Haenlein, 2004). And goat milk is sometimes used to treat human dyspepsia and gastrointestinal dysfunction (Haenlein, 2004). Milk triglycerides, which account for 99% of milk fat, are synthesized from diverse fatty acids in the epithelial cells of mammary gland. After their synthesis, milk triglycerides coalesce to form fat droplets (Bionaz and Loor, 2008, Hansen et al., 1984). The synthesis and transfer of triglycerides involve many processes (e.g., de novo synthesis of fatty acids, triglyceride synthesis, fat droplet formation, and fatty acid uptake and transport), indicating that a unique network of genes participates in milk fat synthesis.

MicroRNAs (miRNAs) are endogenous, small, non-coding RNAs, around 22 nucleotides in length (Barte, 2004). By binding to target mRNA, miRNA can reduce mRNA translation in animals. The miRNA–mRNA base pairing is only 6–8 nucleotides long (Lewis et al., 2005). Therefore, it has been hypothesized that miRNAs have numerous target genes. MiRNAs regulate many metabolic processes, including tissue development (Tanaka et al., 2009), cell differentiation (Sun et al., 2011), and lipid metabolism (Sacco et al., 2012). miRNAs are key regulators of lipid synthesis, oxidation, and homeostasis in various cell lines (Bommer and MacDougald, 2011, Esau et al., 2006) and tissues (Lynn, 2009, Nakanishia et al., 2009). For example, miR-27a suppresses fat synthesis in preadipocyte cell lines (Qun et al., 2008), and miR-107 regulates triglyceride concentrations in fat tissue (Trajkovski et al., 2011). Recent studies indicate that miRNAs may be involved in milk fat metabolism in mammary gland during lactation. miRNAs (e.g., miR-27a and miR-107) in mammary gland of mouse and cow are expressed differently during different stages of lactation (Avril-Sassen et al., 2009, Chen et al., 2010). These miRNAs might be related to milk fat synthesis. Little is known about the biological role of miRNAs in goat mammary gland or the effect of miRNAs on milk fat metabolism. Furthermore no goat miRNA has been deposited in miRBase 17.0. Only a few miRNAs have accession numbers in GenBank. The sequence of goat miR-27a is published in GenBank. Therefore, in this experiment, we evaluated the potential ability of miR-27a to affect milk fat synthesis in goat mammary epithelial cells.

To our knowledge, this is the first study to verify the ability of miR-27a to suppress milk triglyceride accumulation and to alter the fatty acid composition in goat mammary gland epithelial cells. We also determined the expression of goat miR-27a during different lactation stages and analyzed the sensitivity of miR-27a to prolactin. These results significantly advance our understanding about the role of miRNAs during lactation.

Section snippets

Animals, tissue sampling, and RNA extraction

Xinong Saanen Dairy Goat was bred in the experimental farm of Northwest Agricultural University since 1940 and recognized as a specialized dairy goat breed in 1958. Goats from the herd of the farm will be referred to as Xinong Saanen Dairy Goat in this paper.

Three healthy, three-year-old goats of similar weight were selected from the herd for this study. The goats were in the second lactation. Mammary gland tissues were surgically collected from the same goats at mid-lactation (120 days after

miR-27a expression correlates with lactation

The mammary gland undergoes extensive changes in tissue structure and milk production between the dry period and lactation. To investigate whether miR-27a expression correlates with lactation, we assessed miR-27a expression in goat mammary gland. We sampled the mammary gland tissues of three random dairy goats during mid-lactation (120 days after parturition) and during the dry period (60 days before parturition). The total RNA was extracted from the mammary gland tissue and then pooled within

miR-27a decreased triglyceride accumulation

Mammary gland epithelial cells undergo periodic cycles of growth and development, differentiation, and apoptosis corresponding to the physiological states of pregnancy, lactation, and involution (Flint and Gardner, 1994, Folley, 1949). These processes are controlled by a variety of hormones, including insulin and prolactin (Topper and Freeman, 1980). Prolactin plays important role in mammary gland development during the pregnancy (Hennighausen et al., 1997). During lactation, prolactin can

Conclusion

In conclusion, the data suggest a role for miR-27a as a regulator of milk triglyceride accumulation and composition in epithelial cells of goat mammary gland. Additional study is needed about the specific regulation of miR-27a on milk triglyceride synthesis of lactating goats. Such information could potentially be used to improve milk nutrient composition of dairy animals.

Acknowledgments

We are grateful to Dr. William Gale for his critical comments and language editing of the manuscript and to Prof. Li Changan for biopsy sampling. This work was co-supported by the Special Fund for Agro-scientific Research in the Public Interest of China (no. 201103038) and the National Natural Science Foundation of China (31072013).

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