Mechanistic Approach to Preventing Atrophy and Restoring Function in Older Adults

The investigative team has completed recent studies showing that essential amino acid (EAA)
supplementation attenuates quadriceps atrophy and accelerates the return of functional
mobility following TKA. For patients on EAA, quadriceps atrophy was only -6% and -3% in the
operative and non-operative quadriceps, respectively, 6 wks after TKA, but -18% and -10%,
respectively, in patients on Placebo, a threefold difference. Of clinical relevance, the
patients on EAA were able to maintain strength and demonstrated an accelerated return of
functional mobility vs. Placebo 6 wks post-TKA. Positive changes at the cellular level are
likely responsible for the reduction in muscle loss and preservation of strength, and to
explain the acceleration of the return of functional mobility. As such, we hypothesize that
twice-daily ingestion of 20 g of EAA from 1 wk before to 6 wks after TKA will increase basal
rates of muscle protein synthesis via inactivation of catabolic signaling (FoxO3a), and
up-regulation of anabolic and cyto-protective proteins. We also propose that short-term
atrophy prevention and accelerated return of functional mobility will translate into
long-term (6 mo and 1 yr post-TKA) structural and functional adaptations, leading to improved
quality of life in TKA patients with EAA supplementation vs. Placebo.

The study will use a two-arm parallel design to determine the effect of EAA supplementation
on post-TKA muscle cell structure and function, and quality of life in the shorter term (6
months post-TKA) and long term (1 yr post-TKA) vs. Placebo. Subjects will be 80 older male
and female adults having primary TKA at the Slocum Center for Orthopedics and Sports
Medicine, Eugene, Oregon. Slocum study staff will pre-screen/identify potential subjects for
recruitment based on inclusion/exclusion criteria. Eligible patients will be invited to join
the study. After the completion of informed consent procedures, participants will be assigned
a unique patient identification number, and randomly assigned to either EAA or Placebo on a
1:1 allocation ratio, with blinding of treatment condition to subject and research
staff/statisticians collecting data. Twice-daily ingestion of 20 g of supplement will begin 7
days prior to TKA and end at 6 wks post. EAA subjects will receive EAA and Placebo subjects
will receive the non-essential amino acid Alinine. Subjects will document compliance with the
supplement protocol in a log book and will return empty vials. Subjects will be followed for
1 yr.

Key research questions are: (a) Does EAA prevent shorter-term (6 wks post-TKA) bilateral
muscle atrophy, preserve quadriceps strength, and accelerate the return of functional
mobility vs. Placebo? (b) Are there shorter-term sex differences on outcome measures? (c)
Will EAA increase longer-term (6 mo post-TKA) quadriceps strength and functional mobility vs.
Placebo? (d) Does EAA improve long-term (1 yr post-TKA) functional mobility and measures of
quality of life vs. Placebo? (e) Are there long-term sex differences in functional mobility
or measures of quality of life? Assessment points will be at 6 wks, 4 wks, and 1 wk prior to
surgery, and 1 wk, 2 wks, 6 wks, 6 mos, and 1 yr post-TKA. At different assessment points, we
will collect demographic, medical (e.g., length of hospital stay, tourniquet use),
physiological (e.g., muscle biopsy, MRI, DEXA, strength; blood tests), pharmacologic,
functional mobility, physical activity (accelerometer), psychometric (e.g., quality of life;
Veterans RAND 12-item Health Survey [VR-12]), food intake (3-day diary), physical therapy,
and perceived pain data. The following schedule of assessments will be followed: no more than
6 mo pre-TKA (enrollment and screening), 6 wks pre-TKA (screening, surveys, DEXA scan, MRI
test, physical activity and food recording), 4 weeks pre-TKA (blood draw, strength and
functional mobility testing, muscle and fat biopsy); 1 wk pre-TKA and in hospital (physical
activity and food recording); 1 wk post-TKA (physical activity and food recording); 2 wks
post-TKA (blood draw, physical activity and food recording); 6 wks post-TKA (blood draw, MRI
test, strength and functional mobility testing, physical activity and food recording, muscle
and fat biopsy); 3 mos post-TKA (surveys, strength and functional mobility testing, physical
activity and food recording); 6 mos post-TKA (surveys, DEXA scan, MRI test, strength and
functional mobility testing, physical activity and food recording, muscle and fat biopsy),
and 1 yr (surveys, DEXA scan, MRI test, strength and functional mobility testing, physical
activity and food recording, muscle and fat biopsy).

Data will be collected from a variety of sources, including surveys, electronic medical
records, medical and functional tests, and staff reports. Data will be entered and
double-verified in password-protected spreadsheets sand databases stored behind a firewall.
Any electronic medical records data captured will use secure data transfer and
HIPAA-compliant protocols approved by the University of Oregon IRB. Staff reports will be
forwarded directly to project data managers. All personal data will be identified by numbers
rather than names. Interim reports of project results will be made to the Data and Safety
Monitoring Board.

Power analyses indicated that a sample size of 80 subjects would be sufficient to detect
anticipated effects on primary outcomes: operative leg quadriceps volume (MRI), non-operative
leg quadriceps volume (MRI), isometric strength, Get Up And Go test, stair climb up test,
stair climb down test, and six-minute walk test. The mean effect size across these outcomes,
based on baseline- to 6-week percent change in pilot data) was d = 1.01, reflecting large
effects. For a sample size of 30/group, this study can detect ESs of d > .79 with 85% power,
and the minimally detectable ES drops to 0.68 or 0.52 with pretest covariates of r = .50 or
.75. Every effort will be made to reduce attrition and obtain data on all participants at all
assessment points. However, attrition is expected. Because this study will have power to
detect anticipated physiological and functional effects with 30 subjects per condition, we
will enroll 80 total subjects (40/condition) to account for possible attrition.

The EAA intervention is designed to attenuate muscle loss, improve functional outcomes, and
enhance quality of life. Data analyses will focus on shorter-term, longer-term results, and
change over time. Preliminary analyses will employ descriptive statistics to understand the
nature of the data and ensure that data distributions are appropriate for the statistical
tests employed. Chi-square tests and analyses of variance, as appropriate, will be conducted
to evaluate the equivalence of continuing participants vs. dropouts on demographic, medical
history, and recent levels of dependent variables.

We will conduct random coefficients analysis (RCA) to model muscle and functional mobility
across time, as well as covariates that could affect outcomes, such as age, sex, physical
activity, and dietary intake. RCA models trajectories from assessments nested within
subjects; test of condition are represented by the interaction between a time factor and
treatment condition. The RCA avoids the many pitfalls associated with traditional repeated
measures ANOVA: It adjusts for within-individual dependence or autocorrelation in the data,
can model nonlinear growth, does not require fixed spacing among assessments, and
accommodates missing values over time. By using any available data across time, the RCA will
limit the effects of missing data, reducing bias and increasing power. Based on our
preliminary data, we estimate that we will be able to collect >92% of all data points.

Expected study outcomes are as follows:

1. With successful completion of this research, we expect to demonstrate that EAA prevents
muscle atrophy bilaterally, as our preliminary data suggest. To date, it is not known if
EAA can prevent atrophy.

2. We expect to show that atrophy prevention will lead to strength gains and accelerated
return of functional mobility. We further expect to show that EAA may have positive
effects on central activation deficits, as our preliminary data suggest that strength is
increased by 6 wks with EAA. To date, it is not known if preserving muscle following TKA
will increase strength and augment the return of functional mobility.

3. We expect to document that EAA will increase muscle cell size (CSA) bilaterally, in the
vastus lateralis of the operative and non-operative quadriceps. It is not known if
atrophy prevention and early return of functional mobility will stimulate positive gains
in muscle cell structure.

4. We expect to demonstrate that EAA increases mitochondrial mass bilaterally and increases
mitochondrial respiration at rest. By sampling from the non-operative (control leg) and
operative leg, we expect to show that EAA normalizes mitochondrial function over time (6
mo and/or 1 yr post-TKA). It is not known if early gains in functional mobility will
positively impact muscle cell function.

5. We expect to show that quality of life (as measured by instruments such as VR-12) will
be significantly increased with EAA. It is not known to what extent the above positive
gains will have on longer-term (6 mo and 1 yr) quality of life.

We expect each of the above to stimulate and be transformative. EAA supplements are
inexpensive ($800/patient or $16/day), are well tolerated, and can be implemented
immediately.

Inclusion Criteria:

- Age: between 50-80 years.

- Primary TKA surgery.

Exclusion Criteria:

1. Previous TKA and/or total hip arthroplasty surgery (older subjects).

2. Dementia or related mental issues that may potentially put the subject at risk as
determined by the surgeon.

3. Untreated endocrine disease (Hypo/Hyperthyroidism, Addison's or Cushing's syndrome,
etc.).

4. Significant heart, liver, kidney, blood, or respiratory disease.

5. Peripheral vascular disease.

6. Active cancer.

7. Recent (within 6 months) treatment with anabolic steroids.

8. Alcohol or drug abuse.

9. Inability to have MRI