THE EFFECTS OF DIET INDUCED OBESITY ON SECRETION, MILK COMPOSITION, AND MAMMARY GLAND MICROENVIRONMENT IN A MOUSE MODEL

Open Access
Author:
Velasquez, Vanessa Renee
Graduate Program:
Nutrition
Degree:
Master of Science
Document Type:
Master Thesis
Date of Defense:
June 07, 2011
Committee Members:
  • Shannon Leanne Kelleher, Thesis Advisor
Keywords:
  • obesity
  • DIO
  • mouse mammary gland
  • lactation
  • milk secretion
Abstract:
Breast milk is the ideal food for infants, offering essential nutrients and energy for growth and proteins from the mother to develop immunity. Exclusive breastfeeding is therefore recommended for the infant during the first 6 months of life. Obese women however are less likely to initiate breastfeeding, breastfeed for shorter durations, and often suffer a delay in the onset of lactogenesis. 34% of reproductive age women are obese, thus obesity-impaired lactation is a significant problem. Recent research is focused on the impaired onset of lactogenesis with little information on how obesity affects milk secretion after the onset of lactogenesis and limited information on milk composition. DBA/2J mice were used to characterize how diet induced obesity (DIO) affects early, established lactation in a series of 3 studies. Female mice were fed either a high fat diet (HFD) with 45% kcal from fat or a control diet (CD) with 10% kcal from fat throughout the study. Mice were bred ~5 weeks after start of experimental diet and the body weights of the HFD group were significantly heavier than the control group. Experiments were performed during early lactation at LD 4-6. In study 1, we explored effects of DIO on secretion during established lactation. Milk secretion was measured by weigh-suckle-weigh, mammary gland morphology and the number of alveoli were visually assessed by hemoatoxylin and eosin (H and E) staining. The expression of major milk proteins was analyzed by real-time PCR and immunoblot. There was no significant effect of a high fat diet on milk secretion, mammary gland morphology, number of alveoli, or mRNA expression of secretory proteins &#946;-casein and &#945;-lactalbumin between mice fed HFD vs. mice fed CD. The mRNA expression of whey acidic protein (WAP) in the mammary gland was significantly lower (P<0.01) in mice fed HFD compared with mice fed CD, yet protein expression of WAP in the mammary gland was not different between groups. The protein expression of &#946;-casein was significantly (P<0.05) greater in the mammary glands of mice fed HFD compared with mice fed CD. Study 1 determined that DIO does not impair secretion during established lactation as evidenced by no impairment in volume of milk secreted, no abnormalities in mammary gland morphology and no impairment in the expression of secretory proteins. These findings emphasize the importance of an obese mother receiving adequate lactational support in order to establish the onset of lactogenesis as obesity should otherwise cause no secretory impairment. In study 2, we explored effects of DIO on milk composition. Fat content, lactose and total protein concentration, protein composition and zinc concentration were analyzed. There was no significant effect of a high fat diet on % fat, lactose, or zinc concentration. Mice fed HFD had significantly lower (P<0.001) total protein concentration in milk. When the protein composition of milk was analyzed, serum albumin in milk was lower (P<0.05) but expression of caseins was greater (&#945;-casein, P = 0.065; &#946;-casein, P < 0.01; &#947;-casein, P < 0.01; &#949;-casein, P < 0.05) in milk from mice fed HFD. Study 2 determined that DIO results in deceased total protein in milk but increased casein secretion, which could impact infant health. This study emphasizes the necessity for additional research on the effects of milk from an obese mother on infant growth and health. In study 3, we explored effects of DIO on the mammary gland microenvironment, as the cells and molecules that surround mammary epithelial cells may influence their function. Mammary gland adiposity was compared between groups using H and E staining. Macrophage infiltration was analyzed by immunohistochemistry (IHC) using the macrophage marker F4/80 and the gene expression of pro-inflammatory cytokines and Receptor Activator of Nuclear Factor &#954; B (RANK) was analyzed by real time-PCR. We observed larger adipocytes and more macrophages present in mammary glands from mice fed HFD compared with mice fed CD. The mRNA expression of pro-inflammatory cytokines (TNF-&#945;, P<0.05; MCSF, P<0.01; and IL-6, P=0.07) in the mammary glands of mice fed HFD was greater compared with mice fed CD. The mRNA expression of RANK was significantly higher (P<0.05) in mice fed HFD compared with mice fed CD. Study 3 determined that DIO alters the mammary gland microenvironment by increasing adiposity, macrophage infiltration and the expression of pro-inflammatory cytokines and RANK. The altered cytokine milieu of an inflammatory lactating mammary gland microenvironment could offer insight into the mechanisms that delay lactogenesis. In conclusion, DIO produces an inflammatory microenvironment in the mammary gland, which may impact the expression of proteins in the mammary gland and in milk. These studies offer preliminary data for many new avenues in research including additional regulatory mechanisms of secretary proteins, the safety of milk from an obese mother, and RANK as a target for obesity-impaired lactation. Our studies also conclude that if an obese mother decides to breastfeed, she should be offered lactational support to allow her to initiate lactogenesis, as obesity should not cause any additional impairment on lactation.