Proline plays important roles in protein synthesis and structure, metabolism (particularly the synthesis of arginine, polyamines, and glutamate via pyrroline-5-carboxylate), and nutrition, as well as wound healing, antioxidative reactions, and immune responses. On a pergram basis, proline plus hydroxyproline are most abundant in collagen and milk proteins, and requirements of proline for whole-body protein synthesis are the greatest among all amino acids. Therefore, physiological needs for proline are particularly high during the life cycle. While most mammals (including humans and pigs) can synthesize proline from arginine and glutamine/glutamate, rates of endogenous synthesis are inadequate for neonates, birds, and fish. Thus, work with young pigs (a widely used animal model for studying infant nutrition) has shown that supplementing 0.0, 0.35, 0.7, 1.05, 1.4, and 2.1% proline to a proline-free chemically defined diet containing 0.48% arginine and 2% glutamate dose dependently improved daily growth rate and feed efficiency while reducing concentrations of urea in plasma. Additionally, maximal growth performance of chickens depended on at least 0.8% proline in the diet. Likewise, dietary supplementation with 0.07, 0.14, and 0.28% hydroxyproline (a metabolite of proline) to a plant protein-based diet enhanced weight gains of salmon. Based on its regulatory roles in cellular biochemistry, proline can be considered as a functional amino acid for mammalian, avian, and aquatic species. Further research is warranted to develop effective strategies of dietary supplementation with proline or hydroxyproline to benefit health, growth, and development of animals and humans.

We report on how a dimer of the cell-penetrating peptide TAT, dfTAT, penetrates live cells by escaping from endosomes with a particularly high efficiency. By mediating endosomal leakage, dfTAT also delivers proteins into cultured cells after a simple co-incubation procedure. Cytosolic delivery is achieved in most cells in a culture and only a relatively small amount of material remains trapped inside endosomes. Delivery does not require binding interactions between dfTAT and a protein, multiple molecules can be delivered at once, and delivery can be repeated. Remarkably, dfTAT-mediated delivery does not noticeably impact cell viability, proliferation, or gene expression. This new delivery strategy should be extremely useful for cell-based assays, cellular imaging applications, and the ex vivo manipulation of cells.

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Embryonic loss and intrauterine growth restriction (IUGR) are significant problems in humans and other animals. Results from studies involving pigs and sheep have indicated that limited uterine capacity and placental insufficiency are major factors contributing to suboptimal reproduction in mammals. Our discovery of the unusual abundance of the arginine family of amino acids in porcine and ovine allantoic fluids during early gestation led to the novel hypothesis that arginine plays an important role in conceptus (embryo and extra-embryonic membranes) development. Arginine is metabolized to ornithine, proline, and nitric oxide, with each having important physiological functions. Nitric oxide is a vasodilator and angiogenic factor, whereas ornithine and proline are substrates for uterine and placental synthesis of polyamines that are key regulators of gene expression, protein synthesis, and angiogenesis. Additionally, arginine activates the mechanistic (mammalian) target of rapamycin cell signaling pathway to stimulate protein synthesis in the placenta, uterus, and fetus. Thus, dietary supplementation with 0.83 % L-arginine to gilts consuming 2 kg of a typical gestation diet between either days 14 and 28 or between days 30 and 114 of pregnancy increases the number of live-born piglets and litter birth weight. Similar results have been reported for gestating rats and ewes. In sheep, arginine also stimulates development of fetal brown adipose tissue. Furthermore, oral administration of arginine to women with IUGR has been reported to enhance fetal growth. Collectively, enhancement of uterine as well as placental growth and function through dietary arginine supplementation provides an effective solution to improving embryonic and fetal survival and growth.

Although there are published studies of proline biochemistry and nutrition in cultured cells and postnatal animals, little is known about proline metabolism and function in the conceptus (embryo/fetus, associated placental membranes, and fetal fluids). Because of the invasive nature of biochemical research on placental and fetal growth, animal models are often used to test hypotheses of biological importance. Recent evidence from studies with pigs and sheep shows that proline is a major substrate for polyamine synthesis via proline oxidase, ornithine aminotransferase, and ornithine decarboxylase in placentae. Both porcine and ovine placentae have a high capacity for proline catabolism and polyamine production. In addition, allantoic and amniotic fluids contain enzymes to convert proline into ornithine, which is delivered through the circulation to placental tissues. There is exquisite metabolic coordination among integrated pathways that support highest rates of polyamine synthesis and concentrations in placentae during early gestation when placental growth is most rapid. Interestingly, reduced placental and fetal growth are associated with reductions in placental proline transport, proline oxidase activity, and concentrations of polyamines in gestating dams with either naturally occurring or malnutrition-induced growth retardation. Conversely, increasing proline availability in maternal plasma through nutritional or pharmacological modulation in pigs and sheep enhances concentrations of proline and polyamines in placentae and fetal fluids, as well as fetal growth. These novel findings suggest an important role for proline in conceptus metabolism, growth and development, as well as a potential treatment for intrauterine growth restriction, which is a significant problem in both human medicine and animal agriculture.