219 EFFECTS OF ADMINISTRATION OF MILK FROM TRANSGENIC COWS CONTAINING RECOMBINANT HUMAN LACTOFERRIN IN A PIG MODEL OF MALNUTRITION

Abstract

Infant mortality is still a major problem, with the interaction between malnutrition and diarrhoea among the leading causes of death. One option to fight both diarrhoea and malnutrition is breastfeeding. Benefits of breast milk are attributed to the actions of antimicrobial proteins in human milk, such as lactoferrin (LF), which increase intestinal and systemic immune functions. One way to convey the benefits of LF to children is the use of transgenic animals that express human proteins in the mammary gland. In this sense, the availability of animal milk with properties of human milk can be a potential source to increase and prolong the protective benefits of human milk in reducing disease and stimulating growth. Transgenic cows expressing rhLF were produced by pronuclear microinjection with the goal of using the milk to improve human health. To test this hypothesis, we have created a model of malnutrition in pigs by reducing the intake (50%) of calories and protein. The animals (n = 26) were randomly divided as follows: after weaning at 3 weeks of age, 18 animals were fed the protein and calorie-restricted diet (mal) for 3 weeks and 8 animals served as a control group and were fed standard feed (full-fed). After 3 weeks, 4 animals in each group were necropsied and the remaining animals (n = 18) were placed into the following experimental groups: 4 animals remained in the control group (full-fed-no milk), and the 14 malnourished animals were divided as follows: 4 animals were maintained on food restriction but received no milk (mal-no milk) and 10 animals were maintained on food restriction with 5 receiving 500 mL of control milk/day (con milk) and 5 receiving 500 mL of rhLF milk/day (rhLF milk) for a total of 15 days. Intestinal permeability and morphology, mRNA expression of tight junction proteins (ZO1, claudin, occludin), and cytokines (TGF-β, TLR-4, IL-10, TNF-α, IL-6 IL-8, CCL-11) in the intestine, and hematological parameters were assessed. Data were analysed by ANOVA with P-values <0.05 considered statistically significant. The restricted diet was capable of inducing a state of malnutrition after 3 weeks as demonstrated by multiple changes in blood chemistry, a significant decrease in gut surface area, and an increase in electrical conductance indicative of compromised intestinal barrier function. Supplementation of the diet with either control milk or rhLF milk promoted the recovery of the intestine as indicated by significantly improved intestinal morphology and permeability. Levels of TNF-α were increased in the mal-no milk group; however, rhLF-fed animals were capable of regulating the expression of TNF-α, which did not significantly differ from full-fed controls. Tight junction proteins were also significantly up-regulated in the rhLF group. Overall, a model of malnutrition was established and the administration of both control and rhLF milk was beneficial in the recovery of the gastrointestinal tract. Our intention is that such milk from transgenic animals can benefit malnourished children around the world.