The importance of non-essential amino acids for maintaining skeletal muscle protein synthesis rates in healthy young men

Muscle tissue consists of proteins. These proteins are built up of small
building blocks: amino acids. Some amino acids must be consumed through the
diet because they cannot be made within the body (essential amino acids), while
some amino acids can be made within the body (non-essential amino acids).
Regardless of the source, all amino acids are required for complete muscle
protein building. Therefore, if non-essential amino acids are not consumed
through foods, they must be made adequately in the body, which is a process
that requires energy. However, it is not yet known how a diet lacking
non-essential amino acids changes muscle building, energy metabolism, and
muscle metabolism. The goal of this study is to investigate if non-essential
amino acids are an important component of a human diet.

The purpose of this study is to determine if a diet lacking NEAA, with or
without being replaced by additional EAA, influences skeletal muscle protein
synthesis rates and whole-body metabolism in healthy young men.

Randomized, parallel design, double-blind, dietary intervention study in which
males will consume a standardized low-protein diet supplemented with various
amounts of essential amino acids and non-essential amino acids (in the form of
free amino acid drinks) for 10-days.

All subjects will consume a standardized low-protein diet (<2% energy as
protein) supplemented with various amounts of EAA and NEAA in the form of free
amino acid drinks. Subjects will be randomized to either 1) a conventional diet
group (standardized diet supplemented with 7% energy as EAA + 10% as NEAA), 2)
a diet lacking NEAA (supplemented with 7% EAA + 0% NEAA), or 3) a diet lacking
NEAA and replaced by additional EAA (supplemented with 17% EAA + 0% NEAA,
isonitrogenous with conventional diet). The dietary intervention will be 10
days in duration and deuterium oxide will be consumed throughout the
intervention period.

The risks involved in participating in this experiment are minimal. The blood
sample will be comparable to a normal blood draw and the only risk is a small
local hematoma. This is also true for muscle biopsies. Muscle biopsies will be
taken through a small (5 mm) incision, following local anaesthetic of the skin
and muscle fascia, and will heal completely. Muscle biopsies will only be
obtained by an experienced physician. Skin biopsies will be taken from the
buttocks using a small (4 mm) punch. Skin biopsies are frequently used method
for diagnostic and therapeutic purposes and will heal completely. The
ingestion of deuterium oxide has been applied in numerous published studies and
is entirely safe and non-toxic in the amounts provided in the present study. We
will take 4 blood samples (80 mL total) during the experimental trial. For the
experimental trials (D2O dosing day, Test Day 1, Test Day 2, test Day 3),
participants have to be fasted, so they are not allowed to eat and drink
(except for water) from 22:00 the evening before. Also, 3 days prior to the
experimental trial participants should keep their diet as constant as possible,
should not perform any type of intense physical exercise, and should not
consume alcohol. Furthermore, we will ask the participants to fill out a
dietary and activity record for 3 days prior to the experimental trial. The
time investment is 2 hours (screening), 4 hours (D2O Dosing Day), 2 hours (Test
Day 1), 0.5 hours (Test Day 2), 2.5 hours (Test Day 3) and filling in the food
and activity dairies diaries (30 minutes per day). There are some risks of
discovering an incidental finding when performing an MRI, however if the
participant does not want to receive such information, they cannot participant
in the research. If there are unexpected findings in the form of abnormal
values/structures identified during the MRI (and any other research tests
performed), the participant will be informed.

The diet lacking NEAA is expected to pose no risk to participants, as it has
been shown that a prolonged diet very low in protein does not lower fat-free
mass (diet composed of 5% energy from protein) or lower muscle protein
synthesis rates (diet composed of 9% energy from protein), suggesting that a
very low protein intake does not compromise muscle mass maintenance. The
experimental diets contain normal commercially available nutritional
ingredients that are safe for human consumption and for this reason do not form
any health risks. Furthermore, the diets are tailored to the participant*s
energy and macronutrient needs, and composed according to the dietary
guidelines of the Dutch National Health Council.

There is no direct benefit for the participants, only their contribution to
scientific knowledge of the role of nutritional components in a diet. The
current study will provide valuable information about the role of essential and
non-essential amino acids in the human diet. A small benefit for the
participants is that they will get insight into their body composition (MRI).