Effects of Protein Versus Carbohydrate Supplementation on Markers of Immune Response in Master Triathletes: A Randomized Controlled Trial.

Objective: This study examines the long-term effects of ingesting hydrolyzed beef protein versus carbohydrate on indirect markers of immunity during 10 weeks of endurance training in master-aged triathletes (n = 16, age 35-60 years). Methods: Participants were randomly assigned to either a hydrolyzed beef protein (PRO, n = 8) or nonprotein isoenergetic carbohydrate (CHO, n = 8) condition, which consisted of ingesting 20 g of each supplement, mixed with water, once a day immediately post workout, or before breakfast on nontraining days. Salivary human neutrophil peptides (HNP1-3) were measured before and after performing an incremental endurance test to volitional exhaustion at both pre and post intervention. Additionally, baseline levels of platelets, neutrophils, eosinophil basophils, monocytes, and lymphocytes were determined at pre and post intervention. Results: No significant changes in baseline concentration and secretion rate of salivary HNP1-3 were observed for either treatment. The CHO group showed a nonsignificant decrease in resting HNP1-3 concentrations following the intervention (p = 0.052, effect size d = 0.53). Protein supplementation demonstrated a significant reduction in lymphocyte counts pre to post intervention (mean [SD]: 2.30 [0.57] vs. 1.93 [0.45] 103/mm3, p = 0.046, d = 0.77), along with a moderate but not statistically significant increase (d = 0.75, p = 0.051) of the neutrophil-to-lymphocyte ratio. Conclusions: In master-aged triathletes, postworkout ingestion of only protein, with no carbohydrate, may not be as effective as carbohydrate alone to attenuate negative long-term changes of some salivary and cellular immunological markers. Future studies should consider the co-ingestion of both macronutrients.

Prodefensins are matrix proteins of specific granules in human neutrophils.

Defensins are potent antimicrobial and proinflammatory peptides. The human neutrophil defensins human neutrophil peptide (HNP)-1-3 are synthesized as 94 amino acide (aa) preproHNPs, which are converted to 75 aa proHNPs by cotranslational removal of a 19 aa endoplasmic reticulum signal peptide. At the promyelocytic stage of myelopoiesis, proHNPs are further proteolytically modified and accumulate in azurophil granules as 29-30 aa HNPs. In contrast, proHNPs produced by more mature myeloid cells are not subjected to proteolytic cleavage and undergo a high degree of constitutive exocytosis. The proHNPs are devoid of antimicrobial potential, and the significance of their secretion is unknown. To investigate whether mature neutrophils contain proHNPs, we developed antibodies against proHNP-1 by DNA immunization of rabbits. In addition, antibodies against the 45 aa proHNP pro-piece were raised by conventional immunization procedures. These antibodies allowed detection of proHNPs in homogenates of peripheral blood neutrophils. The proHNPs were isolated by affinity chromatography, and their identity was confirmed by mass spectrometry and N-terminal aa sequence analysis. Finally, the neutrophil proHNPs were identified as novel matrix proteins of specific granules by subcellular fractionation experiments, release studies, and immunoelectron microscopy. Thus, human neutrophils not only store large amounts of mature defensins in azurophil granules but also contain a more easily mobilized reservoir of unprocessed prodefensins in specific granules.