Rationale: Worldwide, the aging population is increasing. Aging is associated with loss of independence and increased risk of co-morbidity. Sufficient protein intake is imperative to maintain skeletal muscle mass and overall health, and thereby stimulates active aging. It has been suggested that age-dependent changes in gastrointestinal (GI) tract physiology affect the amount of protein digested and absorbed, as well as the intestinal microbiota. However, it is not clear to what extent actual protein digestion and absorption are altered in older individuals. So far, no human data on the expression and activity of brush border enzymes and protein-related transporters in small intestinal epithelial cells are available. Understanding the age-dependent digestion and absorption kinetics of protein is important for creating specific diets in older individuals to improve protein intake and thereby stimulate healthy aging.

Objective: To assess the effects of aging on protein digestion and amino acid absorption in healthy humans in vivo.

Study design: Cross-sectional Study population: 12 healthy, non-obese (BMI 18.5-30kg/m2) young adults (age: 18-35 y inclusive) and 12 community dwelling older adults (age: ≥67 y).

Intervention: Subjects will undergo a standardized heavy water dosing protocol for 2 days (1 "loading day", 1 "maintenance day"). Blood and saliva sampling will be conducted throughout the heavy water dosing protocol to determine body water and amino acid enrichment levels. On the experimental test day, a gastroscopy with collection of duodenal mucosa tissue samples will be performed to allow assessment of the expression and activity of brush border enzymes and protein-related transporters, and to determine fractional duodenal mucosal protein synthesis rate. Fecal samples will be collected at baseline and after the gastroscopy.

Main study parameters/endpoints: The primary study parameter is the mRNA expression of brush border enzymes and small peptide and amino acid transporters in duodenal mucosal cells. Secondary study parameters include protein expression of brush border enzymes and small peptide and amino acid transporters in duodenal mucosal cells, fecal microbial fermentation metabolites and duodenal mucosal protein synthesis rate.

Worldwide, the proportion of older individuals in the population is increasing. The progressive loss of skeletal muscle mass with aging, or sarcopenia, has a major impact on our health care system and costs due to increased morbidity and a greater need for hospitalization and/or institutionalization. A sufficient protein intake is imperative to maintain skeletal muscle mass and overall health, and thereby stimulates active aging. Whereas the overall protein intake is generally lower in older as compared to young adults, findings indicate that older adults need more protein to increase muscle protein synthesis, known as anabolic resistance. The exact mechanisms underlying anabolic resistance are not fully discovered yet. Age-dependent changes in gastrointestinal tract physiology have been suggested as a potential cause.

Following protein ingestion, protein is cleaved into small peptides and amino acids by digestive enzymes secreted in the stomach (pepsin) and small intestinal lumen (pancreatic trypsin, chymotrypsin, elastase), as well as brush border enzymes (aminopeptidase, carboxypeptidase, dipeptidase). Brush border enzymes are responsible for the final digestion of small peptides into dipeptides, tripeptides and amino acids. Subsequently, these small peptides and amino acids are absorbed across the intestinal mucosa by various membrane-bound transporters. The majority of dietary-protein derived amino acids is released into the systemic circulation and transported and taken up by various peripheral tissues in the postprandial phase.

It has been shown that postprandial plasma amino acid availability is lower in older compared to young adults. This indicates that digestion and absorption of proteins are likely attenuated in older individuals, thereby reducing the systemic availability of protein-derived amino acids for muscle protein synthesis and other relevant physiological processes.

Limited studies have focused on changes in gastrointestinal tract physiology with aging. Human studies on the expression and activity of brush border enzymes and protein transporters (i.e. for dipeptides, tripeptides and amino acids) being crucial for the final steps of digestion and absorption, respectively, are lacking. Therefore, the aim of the present study is to assess the expression and activity of brush border enzymes and expression of protein transporters in small intestinal epithelial cells in older as compared to young adults.

Inclusion Criteria:

• Aged 18-35 years or 67+ years
• Body mass index (BMI) between 18.5 and 30 kg/m2
• Regular stool frequency ranging from 1-3 times/day

Exclusion Criteria:

• History of cardiovascular, respiratory, gastrointestinal, urogenital, neurological, psychiatric, dermatologic, musculoskeletal, metabolic, endocrine, haematological, immunologic disorders, allergy, major surgery and/or laboratory assessments which might limit participation in or completion of the study protocol, interfere with the execution of the experiment, or potential influence the study outcomes (to be decided by the principal investigator and responsible physician)
• Major abdominal surgery interfering with gastrointestinal function (upon judgement of the principal investigator and responsible physician)
• Use of medication which limit participation in or completion of the study protocol, interferes with the execution of the experiment, or potential influences the study outcomes (to be decided by the principal investigator and responsible physician)
• Use of supplementation (i.e. protein, vitamin, pre- and probiotic supplementation) within 14 days prior to testing
• Administration of investigational drugs or participation in any scientific intervention study prior or during the study, which may interfere with this study (upon judgement of the principal investigator and responsible physician) Specific diet (e.g. vegetarian, vegan, gluten free, no diary) within the study period
• Planning to lose weight during the study period
• Excessive alcohol consumption (defined as > 14 alcoholic consumptions per week)
• Smoking
• Drug use
• Pregnancy
• Lactation
• Donated blood three months prior to the test day
• No given permission to register participation in electronic patient file at MUMC+ and to add records of gastroduodenoscopy