NSA: Serum Proteins (2009)

Citation:

Afolabi PR, Jahoor F, Jackson AA, Stubbs J, Johnstone AM, Faber P, Lobley G, Gibney E, Elia M. The effect of total starvation and very low energy diet in lean men on kinetics of whole body protein and five hepatic secretory proteins. Am J Physiol Endocrinol Metab. 2007 Dec; 293(6): E1,580-E1,89. Epub 2007 Sep 18.

PubMed ID: 17878226
 
Study Design:
Non-randomized trial.
Class:
C - Click here for explanation of classification scheme.
Quality Rating:
Neutral NEUTRAL: See Quality Criteria Checklist below.
Research Purpose:

The purpose of this study was to determine whether the rate of weight loss, independent of magnitude, affects whole body protein metabolism and the synthesis and plasma concentrations of specific hepatic secretory proteins and whether plasma concentrations and synthetic rates of these proteins are related.

Inclusion Criteria:

Although inclusion criteria were not explicitly provided, subjects met these criteria:

  • Lean men
  • Good health
  • Stable weight within previous two months
  • Nonsmokers
  • Not on medication.
Exclusion Criteria:
Description of Study Protocol:

Recruitment

Not specified.

Design

  • Admitted to residential metabolic facility 
  • Followed a weight maintenance diet for seven days
  • Participants divided into two groups: One to follow a starvation diet for five days, the second to follow a very low energy diet (VLED) for about nine days, with the goal to lose approximately 5% of initial body weight
  • Participants on the VLED continued diet until day 21 to achieve a weight loss of not more than 10% of initial body weight
  • Protein kinetic measurements were taken twice in the starvation group and three times in the VLED group. 

Intervention

  • For first seven days, all 11 participants followed:
    • Maintenance diet:
      • 40% fat, 45% carbohydrate, and 15% protein, given in three isoenergetic meals daily
      • Energy intake based on 1.4 times the predicted basal metabolic rate and altered occasionally to maintain body weight
  • Second stage:
    • Six participants followed a starvation diet for five days, with water only
    • Five participants followed a VLED diet for about nine days (until 5% body weight lost):
      • 40% fat, 45% carbohydrate, 15% protein
      • 2.5MJ per day, given daily as three isoenergetic meals
  • The VLED diet subjects continued the diet until day 21 to achieve a weight loss of not more than 10% of initial body weight.

Statistical Analysis

  • Repeated measures ANOVA to assess differences between baseline and after weight loss
  • Split level repeated-measures ANOVA (SPANOVA) (with subject as blocking factor) assessed the effect of rate of 5% weight loss between groups
  • Analysis of covariance used to control for baseline values to assess the effect of rate of 5% weight loss (but data was not described because the effect of the baseline values was not significant).
Data Collection Summary:

Timing of Measurements

  • Starvation diet: Baseline and day five
  • VLED diet: Baseline, day nine and day 21.

Dependent Variables

  • Fractional and absolute rates of synthesis of five hepatic secreted proteins, measured with [13C] glycine infusion:
    • Retinol binding protein (RBP)
    • Prealbumin [transthyretin (TTR)]
    • High-density lipoprotein (HDL)-apoA1
    • Thyroxine (TR)
    • α1-antitrypsin (α1-AT)
  • Whole body protein turnover, measured with [1-13C] leucine infusion
    • Synthesis
    • Breakdown
    • Oxidation. 

Independent Variables

Rate of weight loss, measured by type and duration of diet (starvation for five days or VLED for nine days).

Other Variables

  • Urinary nitrogen
  • Glucose
  • Nonesterified fatty acids (NEFA)
  • β-hydroxybutyrate
  • Triacylglycerol (TAG)
  • Cholesterol
  • Retinol
  • Iron.
Description of Actual Data Sample:

Final N

11 subjects (six in starvation group, five in VLED group).

Anthropometrics

No significant differences between groups at baseline in:

  • Body weight, mean in kg ± standard deviation
    • Starvation group: 71.04±9.15
    • VLED group: 67.04±8.15
  • Fat, percentage of body weight, mean ± standard deviation
    • Starvation group: 13.47±4.31
    • VLED group: 13.68±6.46
  • Urine nitrogen, grams per day, mean ± standard deviation
    • Starvation group: 12.35±1.18
    • VLED group: 12.23±1.94.

Location

Cambridge, United Kingdom.

 

Summary of Results:
  • Mean whole body protein synthesis decreased in both groups at 5% weight loss, with no difference between groups
  • Protein breakdown was significantly different between groups, with an increase of breakdown in the starvation group compared to a decrease in breakdown in the VLED group. However, these rates were not significantly different from baseline at 5% weight loss in either group.
  • Protein breakdown was significantly lower in the VLED group after 7% weight loss
  • The rate of protein oxidation was significantly higher in the starvation group (but not significantly different in the VLED group) after 5% weight loss
  • Oxidation was significantly different between groups.

Leucine Kinetic, μmol•kg-1•h-1

Starvation Group (Rapid Weight Loss)

VLED Group (Slower Weight Loss)

SPANOVA

 

Baseline

(Mean ± SD)

5% Weight Loss

(Mean ± SD)

 Baseline

(Mean ± SD)

5% Weight Loss

(Mean ± SD)

7% Weight Loss

(Mean ± SD)

 P Value

(Effect of Weight Loss)

P Value

(Interaction)

Synthesis

93.96±6.95

91.36±8.96

97.54±12.69

88.66±13.47 85.67±10.72* 0.036 0.205

Breakdown

104.48±7.20

108.58±10.89

108.61±13.52

100.37±15.64 94.67±12.16*

0.313

0.012
Oxidation 10.52±2.11 17.22±3.01* 11.07±2.41 11.71±5.63 8.99±2.07 0.006 0.015

 *P<0.05 vs. baseline.

  • At 5% weight loss, plasma concentrations of RBP and TTR decreased in both groups, and were not significantly different between groups
  • α1-AT concentrations were significantly different between groups, with an increase in the starvation group but no change in the VLED group.

Plasma Protein Concentration, g per L

Starvation Group (Rapid Weight Loss)

VLED Group (Slower Weight Loss)

SPANOVA

 

Baseline

(Mean ± SD)

5% Weight Loss

(Mean ± SD)

 Baseline

(Mean ± SD)

5% Weight Loss

(Mean ± SD)

7% Weight Loss

(Mean ± SD)

P Value

(Effect of Weight Loss)

P Value

(Interaction)

HDL-apoA1

1.24±0.19

1.26±0.21

1.48±0.25

1.23±0.27* 1.06±0.14* 0.107  0.063

RBP

0.031±0.005

0.018±0.007*

0.039±0.005

0.025±0.008* 0.026±0.004*

0.001

1.000
TTR 0.20±0.03 0.13±0.02* 0.22±0.02 0.15±0.01* 0.21±0.09 0.0005 0.464
α1-AT 0.78±0.11 1.04±0.18*  0.87±0.13 0.86±0.09 0.89±0.22 0.038 0.025
TR 1.64±0.33 1.55±0.20 1.65±0.42 1.55±0.23 1.50±0.30 0.400 0.992

 *P<0.05 vs. baseline. 

  • The fractional synthetic rate (FSR) of RBP was significantly higher at 5% weight loss in both groups. However, the difference between the groups was also significant, with the rate being higher in the starvation group
  • The FSRs of HDL-apoA1 and α1-AT were significantly higher at 5% weight loss in the starvation group only, but the difference between groups was significant for HDL-apoA1 only.

 

Fractional Synthetic Rate of Plasma Protein, Percentage per Day

Starvation Group (Rapid Weight Loss)

VLED Group (Slower Weight Loss)

SPANOVA

 

Baseline

(Mean ± SD)

5% Weight Loss

(Mean ± SD)

 Baseline

(Mean ± SD)

5% Weight Loss

(Mean ± SD)

7% Weight Loss

(Mean ± SD)

P Value

(Effect of Weight Loss)

P Value

(Interaction)

HDL-apoA1

31.33±7.45

56.54±11.39*

30.03±4.39

28.51±3.38 31.52±11.14 0.008 0.003

RBP

37.98±8.74

68.62±18.17*

 33.65±5.84

40.77±9.77* 36.29±17.32*

0.0005

0.007
TTR 41.06±5.76 52.69±17.2 37.41±2.9  40.61±9.3 41.76±1.54 0.146 0.376
α1-AT 26.12±4.53 54.48±21.45* 22.52±8.05 29.63±9.06 34.78±19.35 0.010 0.088
TR 21.07±7 29.1±9.40 16.02±2.98 20.64±3.19 22.05±9.79 0.049 0.584

 *P<0.05 vs. baseline.

  • After 5% weight loss, absolute synthetic rate (ASR) of HDL-apoA1 and α1-AT was significantly higher in the starvation group but not the VLED group compared to baseline; however, the difference between groups was statistically significant
  • The ASR of TTR was significantly lower in the VLED group after 5% weight loss, but not in the starvation group, and the difference between groups was not significant.

Absolute Synthetic Rate of Plasma Protein, mg•kg-1•day-1

Starvation Group (Rapid Weight Loss)

VLED Group (Slower Weight Loss)

SPANOVA

 

Baseline

(Mean ± SD)

5% Weight Loss

(Mean ± SD)

 Baseline

(Mean ± SD)

5% Weight Loss

(Mean ± SD)

7% Weight Loss

(Mean ± SD)

P Value

(Effect of Weight Loss)

P Value

(Interaction)

HDL-apoA1

18.25±6.07

35.35±13.39*

21.53±5.73

17.66±5.91 16.79±5.73 0.032 0.003

RBP

0.56±0.21

0.64±0.36

0.61±0.07

0.50±0.13 0.49±0.30

0.753

0.079
TTR 3.75±0.918 3.28±1.23 4.08±0.93 2.95±0.842* 4.18±1.525 0.069 0.413
α1-AT 9.38±1.38 27.22±11.20* 9.44±3.57 13.02±5.03 16.92±10.70 0.002 0.02
TR 16.83±10.39 22.23±9.58 13.13±6.03 16.19±4.50 17.44±9.98 0.638 0.138

Other Findings

  •  Changes in plasma protein concentrations did not always mirror the changes in fractional or absolute synthetic rates
    • During starvation, the concentration of HDL-apoA1 did not change significantly despite a major increase in both FSR and ASR
    • During VLED, the concentration of HDL-apoA1 declined, but without alteration in FSR or ASR
    • There was a similar responses for RBP
    • For α1-AT, the changes in concentration, FSR and ASR were parallel (all increased) during starvation but not during VLED
  • With the exception of cholesterol, circulating concentrations of nutrients and metabolites were significantly different between groups after 5% weight loss
    • Glucose and retinol concentrations were significantly lower than baseline in the starvation group and significantly different from the VLED group
    • NEFA, β-hydroxybutyrate, and TAG concentrations were significantly higher than baseline in the starvation group and significantly different from the the VLED group
    • Iron concentrations were significantly lower than baseline in the VLED group and significantly different from the starvation group.
Author Conclusion:

This study suggests that the concentration and synthetic rates of several hepatic proteins respond to weight loss, but these changes are probably due more to the degree of dietary restriction (starvation vs. VLED) than the magnitude of weight loss. The study also challenges the concept of tight coupling between synthesis and circulating concentration of specific proteins, including those suggested as markers of visceral protein status (e.g., RBP, TTR).

Funding Source:
Government: Scottish Executive Environment and Rural Affairs Department
Industry:
Slimming World
Other:
Reviewer Comments:

The authors described the characteristics of the study participants, but did not explicitly provide inclusion or exclusion criteria. They also gave the number of participants examined, but did not explicitly state whether any participants had dropped out. The authors did not describe how participants were assigned to either the starvation group or VLED group. The number of participants was small, and a power analysis was not provided; therefore, any small effects may not have been identified due to small sample size. The participants were all lean, healthy men and were not selected at random, so findings cannot be generalized. 

Quality Criteria Checklist: Primary Research
Relevance Questions
  1. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (Not Applicable for some epidemiological studies) N/A
  2. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? Yes
  3. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dieteticspractice? Yes
  4. Is the intervention or procedure feasible? (NA for some epidemiological studies) N/A
 
Validity Questions
1. Was the research question clearly stated? Yes
  1.1. Was (were) the specific intervention(s) or procedure(s) [independent variable(s)] identified? Yes
  1.2. Was (were) the outcome(s) [dependent variable(s)] clearly indicated? Yes
  1.3. Were the target population and setting specified? Yes
2. Was the selection of study subjects/patients free from bias? No
  2.1. Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? Yes
  2.2. Were criteria applied equally to all study groups? ???
  2.3. Were health, demographics, and other characteristics of subjects described? No
  2.4. Were the subjects/patients a representative sample of the relevant population? No
3. Were study groups comparable? Yes
  3.1. Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) No
  3.2. Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? Yes
  3.3. Were concurrent controls or comparisons used? (Concurrent preferred over historical control or comparison groups.) No
  3.4. If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? Yes
  3.5. If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable.) N/A
  3.6. If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., "gold standard")? N/A
4. Was method of handling withdrawals described? No
  4.1. Were follow-up methods described and the same for all groups? No
  4.2. Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) No
  4.3. Were all enrolled subjects/patients (in the original sample) accounted for? ???
  4.4. Were reasons for withdrawals similar across groups? N/A
  4.5. If diagnostic test, was decision to perform reference test not dependent on results of test under study? N/A
5. Was blinding used to prevent introduction of bias? No
  5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? No
  5.2. Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) Yes
  5.3. In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? N/A
  5.4. In case control study, was case definition explicit and case ascertainment not influenced by exposure status? N/A
  5.5. In diagnostic study, were test results blinded to patient history and other test results? N/A
6. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were interveningfactors described? Yes
  6.1. In RCT or other intervention trial, were protocols described for all regimens studied? Yes
  6.2. In observational study, were interventions, study settings, and clinicians/provider described? N/A
  6.3. Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Yes
  6.4. Was the amount of exposure and, if relevant, subject/patient compliance measured? Yes
  6.5. Were co-interventions (e.g., ancillary treatments, other therapies) described? No
  6.6. Were extra or unplanned treatments described? No
  6.7. Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? Yes
  6.8. In diagnostic study, were details of test administration and replication sufficient? N/A
7. Were outcomes clearly defined and the measurements valid and reliable? Yes
  7.1. Were primary and secondary endpoints described and relevant to the question? Yes
  7.2. Were nutrition measures appropriate to question and outcomes of concern? Yes
  7.3. Was the period of follow-up long enough for important outcome(s) to occur? Yes
  7.4. Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Yes
  7.5. Was the measurement of effect at an appropriate level of precision? Yes
  7.6. Were other factors accounted for (measured) that could affect outcomes? ???
  7.7. Were the measurements conducted consistently across groups? Yes
8. Was the statistical analysis appropriate for the study design and type of outcome indicators? Yes
  8.1. Were statistical analyses adequately described and the results reported appropriately? Yes
  8.2. Were correct statistical tests used and assumptions of test not violated? Yes
  8.3. Were statistics reported with levels of significance and/or confidence intervals? Yes
  8.4. Was "intent to treat" analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? N/A
  8.5. Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? Yes
  8.6. Was clinical significance as well as statistical significance reported? No
  8.7. If negative findings, was a power calculation reported to address type 2 error? No
9. Are conclusions supported by results with biases and limitations taken into consideration? Yes
  9.1. Is there a discussion of findings? Yes
  9.2. Are biases and study limitations identified and discussed? Yes
10. Is bias due to study's funding or sponsorship unlikely? ???
  10.1. Were sources of funding and investigators' affiliations described? Yes
  10.2. Was the study free from apparent conflict of interest? ???