Original Investigation | Psychiatry

Association Between Ghrelin and Body Weight Trajectory in Individuals

With Anorexia Nervosa
Youngjung R. Kim, MD, PhD; Meghan S. Lauze, BS; Meghan Slattery, MSN; Roy H. Perlis, MD, MSc; Laura M. Holsen, PhD; Lauren Breithaupt, PhD;
Casey M. Stern, BA; Maurizio Fava, MD; Jennifer J. Thomas, PhD; Elizabeth A. Lawson, MD; Madhusmita Misra, MD, MPH; Kamryn T. Eddy, PhD

Abstract Key Points

Question Is there an association
IMPORTANCE Individuals with anorexia nervosa maintain extremely low body weights despite
between circulating levels of the
elevations in the circulating orexigenic hormone ghrelin. Whether circulating levels of endogenous
orexigenic hormone ghrelin and body
ghrelin are associated with weight gain in anorexia nervosa is unknown.
weight trajectories in individuals with
anorexia nervosa?
OBJECTIVE To examine the association between baseline ghrelin and future weight change in
individuals with anorexia nervosa. Findings In this 18-month cohort study
of 68 girls and young women, elevated
DESIGN, SETTING, AND PARTICIPANTS This prospective cohort study was conducted between baseline ghrelin levels were associated
April 1, 2014, and March 31, 2020, in the US. Girls and women aged 10 to 22 years were recruited from with prospective weight gain in
the greater Boston area from community and area treatment centers, enrolled, and followed up for anorexia nervosa.
18 months. Statistical analyses were performed between January and August 2022.
Meaning This study offers evidence of
association between ghrelin and
EXPOSURES Presence or absence of anorexia nervosa and elevations in endogenous ghrelin.
longitudinal weight outcomes in
individuals with anorexia nervosa;
MAIN OUTCOMES AND MEASURES Changes in age- and sex-standardized body mass index
further studies are warranted to confirm
percentiles from baseline to 9- and 18-month follow-up were the main outcomes of interest.
this association and evaluate the
potential clinical utility of ghrelin in
RESULTS A total of 68 girls and young women (11 [16%] Asian, 4 [6%] Hispanic or Latina, 51 [75%]
anorexia nervosa.
White [non–Hispanic or Latina], and 2 [3%] other race or ethnicity), including 35 with anorexia
nervosa and 33 healthy controls of similar Tanner stage, were included in this study. Anorexia nervosa
and healthy control groups were not statistically different by race and ethnicity, Tanner stage, + Supplemental content
number completing follow-up visits, and the duration between baseline and follow-up visits. At Author affiliations and article information are
baseline, individuals with anorexia nervosa were slightly older (median [IQR], 20.1 [18.5-21.0] vs 18.7 listed at the end of this article.

[14.7-19.4] years; P = .005), had lower body mass index percentiles (median [IQR], 2.4 [0.3-4.7] vs
52.9 [40.4-68.3]; P < .001), and had elevated circulating ghrelin area under the curve composite
index (median [IQR], 1389.4 [1082.5-1646.4] vs 958.5 [743.0-1234.5] pg/mL; P = .003) compared
with healthy individuals. In linear mixed-effects regression analyses, baseline ghrelin was associated
with prospective weight gain after adjusting for diagnosis, age, race, and duration of follow-up (odds
ratio, 2.35; 95% CI, 1.43-3.73; P = .004).

CONCLUSIONS AND RELEVANCE In this cohort study, endogenous ghrelin was associated with
longitudinal weight gain in individuals with anorexia nervosa. Further studies are warranted to
confirm this result and examine its potential clinical utility in treatment development.

JAMA Network Open. 2023;6(3):e234625. doi:10.1001/jamanetworkopen.2023.4625

Open Access. This is an open access article distributed under the terms of the CC-BY License.

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Introduction
Anorexia nervosa (AN) is a debilitating chronic illness with a premature mortality rate 6 times that of
the general population—one of the highest among psychiatric illnesses.1,2 The mainstay of treatment
includes interventions aimed at restoring body weight, which are challenged by illness-driven
motivations to remain thin. No medications are currently approved by the US Food and Drug
Administration for AN; innovative therapies modulating endocrine signals involved in energy
homeostasis, such as ghrelin,3 are being investigated.
Ghrelin is a peptide hormone secreted primarily from the stomach that acts on the growth
hormone secretagogue receptor 1a in the hypothalamus to promote food intake and weight gain in
animal models.4-6 Surprisingly, even though intravenous ghrelin administration increases food intake
in the short term,7 elevations in endogenous ghrelin have not been associated with prospective
weight gain in humans,8-10 except in infants born small for gestational age, whose rates of weight
gain in the first year of life were positively associated with persistent elevations in ghrelin following
glucose administration.11 In AN, intravenous ghrelin administration increases food intake,12 and
endogenous ghrelin levels are significantly elevated,13 related to increased secretory bursts.14
Whether such high levels of circulating ghrelin have a role in long-term weight homeostasis is
unknown. Understanding this association is critical because there is an urgent need for novel
therapeutics to assist with weight restoration in this population. In this study, we tested endogenous
ghrelin as a baseline factor associated with longitudinal weight trajectory in individuals with AN
compared with healthy controls (HCs).

Methods
This cohort study used data collected as part of a larger parent study examining multidimensional
aspects of food motivation pathways, including hormones, neural circuitry, and psychological
symptoms, as mediators of eating disorder trajectories for 18 months in female adolescents and
young adults with low-weight eating disorders, including AN. A summary of the parent study
protocol, including eligibility criteria and study procedures, appears in the eMethods in Supplement 1.
This study followed the Strengthening the Reporting of Observational Studies in Epidemiology
(STROBE) reporting guidelines.15

Data Collection
Data collection took place between April 1, 2014, and March 31, 2020, at Massachusetts General
Hospital (MGH) with institutional review board approval. All participants provided written informed
consent or parental consent with assent of minors younger than 18 years. Participants were recruited
from clinical programs at MGH and beyond, including the MGH Eating Disorders Clinical and Research
Program, specialized treatment centers in the greater Boston area, student health centers at local
universities, and community advertisements.
Female participants aged 10 to 22 years were eligible if they met the Diagnostic and Statistical
Manual of Mental Disorders (Fifth Edition) (DSM-5)16 criteria for AN with insufficient energy intake
relative to requirements due to restrictive eating behaviors characteristic of AN, resulting in a
significantly low body weight, or were HCs without a lifetime history of eating disorders with similar
Tanner stages. DSM-5 defines low body weight for adults with AN as body mass index (BMI) less than
18.5 (calculated as weight in kilograms divided by height in meters squared) but does not define a
cutoff BMI percentile for children and adolescents with AN, and low body weight in this study was
defined as no greater than the 10th BMI percentile for age and sex.17,18 Screening for AN included
evaluation to confirm the lack of any organic conditions that could account for the low body weight.
Study participants had no history of diabetes, gastrointestinal conditions or procedures, recent
systemic hormone use, or pregnancy, which could impact ghrelin levels. Participants were medically
and psychiatrically stable with sufficiently mitigated imminent risk of harm, without dangerous and

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active medical issues (eg, severe or rapidly progressing electrolyte imbalances or anemia) or acutely
elevated psychiatric risk (eg, active suicidality). See the eMethods in Supplement 1 for details of
selection criteria.
Data were collected at baseline, 9-month, and 18-month study visits. Each visit started with
anthropometric measurements and a fasting blood draw, after which participants were asked to eat
an approximately 400-kcal mixed meal standardized for macronutrient content, with approximately
20% fat, 60% carbohydrates, and 20% protein. At 0.5, 1.0, and 2.0 hours after meal initiation, blood
samples were collected. Blood was fractionated to obtain plasma, and plasma total ghrelin was
measured using an enzyme-linked immunosorbent assay (interassay and intra-assay coefficients of
variation, 6.6%, 1.3%; EZGRT-89K, Millipore Sigma).

Statistical Analysis
To test the association between baseline ghrelin and longitudinal changes in body weight, we filtered
the data set for the availability of (1) body weight measurements at baseline and one or both
follow-up visits and (2) baseline ghrelin measurements. We tested our a priori hypothesis with a
2-sided significance level of P < .05 using linear mixed models (LMMs) estimating the weight change
from baseline to either follow-up visit as a repeated-measures outcome of interest with a random
intercept for each individual with ghrelin as the main independent variable of interest.
To compare body weight changes in a developmentally diverse cohort, we derived standardized
BMI percentiles and z scores using the lambda-mu-sigma (LMS) method to account for the changing
skewness of population growth distributions during development, with the participant’s age, sex,
measured weight in kilograms, and measured height in meters, together with the US population
data.19 For individuals older than 20 years (26 of 68 [38%] at baseline, 30 of 63 [48%] at 9 months,
and 40 of 61 [66%] at 18 months), population data were extrapolated to derive LMS-standardized
BMI percentiles.
The primary outcome of interest, the weight change index, was set a priori as the fold change in
BMI percentile from baseline to follow-up (ie, ratio) to account for the significance of weight gain in
the at-risk AN population with extremely low BMI percentiles, together with clinical considerations
taking into account the challenges in gaining weight for participants with AN vs HCs. As an example,
BMI percentile changes from 1 to 5 in participants with AN and from 51 to 55 in HCs both result in an
increase of 4, but the fold-change method factors in the clinical significance of weight gain in
individuals with low body weights (ie, [5 / 1] × 100 or 500% in AN) vs those with healthy weight (ie,
[55 / 51] × 100 or 108% in HCs), respectively. The BMI percentiles (which are always positive) rather
than equivalent z scores (may be positive, zero, or negative) were used after LMS standardization to
compute fold changes using nonzero and nonnegative values.
Dynamic ghrelin measurements across fasting and postprandial time points of the baseline visit
were integrated to derive a single composite index (ie, total area under the curve [AUC]) using the
trapezoid rule.20 Ghrelin AUC was used as the main independent variable of interest for the
univariate LMMs. Age, self-reported race, and diagnostic group at baseline visit and duration of
follow-up specific to the visit for the weight change index were used as covariates in the primary
multivariable LMM. We included race as a variable in this study (and, later in the sensitivity analysis,
ethnicity as well) to account for racial (and ethnic) differences in body weight regulation. Included
racial groups were American Indian or Alaska Native, Asian, Black or African American, White, and
other. Ethnicity groups included HIspanic or Latina and not Hispanic or Latina. Diagnostic group was
included as an interactor given clinical considerations of how chronic starvation in AN may result in
associated elevations in ghrelin. With ghrelin AUC as the main independent variable, no covariates
had significant effects on the model fit, including the interaction term, in a stepwise backward
deletion of variables. Covariate terms were scaled to mitigate the effects of any 1 term having an
undue influence on the model. To minimize bias, no outliers were removed in the main LMM, and
joint assessments of nonindependent covariates (ie, diagnosis and baseline BMI percentiles) were
reserved for sensitivity analyses.

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Table 1. Study Participant Characteristics

Characteristic AN (n = 35) HC (n = 33) Difference (95% CI)a P valueb

Age, median (IQR), y 20.1 (18.5 to 21.0) 18.7 (14.7 to 19.4) 1.6 (0.5 to 2.8) .005
Tanner stage, median (IQR)
Breast development 5 (5 to 5) 5 (4 to 5) 0 (0 to 0) .17
Genital development 5 (4 to 5) 5 (4 to 5) 0 (0 to 0) .81
Race and ethnicityc
Other than White
Any 7 (20) 6 (18) NA
American Indian or Alaska Native 0 0 NA
Asian 6 (17) 5 (15) NA
Black or African American 0 0 NA
.79
Other 1 (3) 1 (3) NA
White 28 (80) 27 (82) NA
Hispanic or Latina 3 (9) 1 (3) NA
Not Hispanic or Latina 25 (71) 26 (79) NA
Completed visitsd
Baseline 35 (100) 33 (100) NA
9 mo 31 (89) 32 (97) NA
Without 9-mo follow-up 4 (11) 1 (3) NA
.38
18 mo 31 (89) 30 (91) NA
Without 18-mo follow-up 4 (11) 3 (9) NA
Both follow-ups 27 (77) 29 (88) NA
Duration of follow-up from baseline, median (IQR), mod
9 mo 9.1 (8.6 to 9.5) 9.1 (8.8 to 9.4) 0.1 (−0.3 to 0.4) .65
18 mo 18.5 (17.8 to 19.4) 18.0 (17.7 to 18.7) 0.5 (−0.1 to 1.2) .14
BMI percentiled
Baseline 2.4 (0.3 to 4.7) 52.9 (40.4 to 68.3) −51.8 (−58.8 to −46.1) <.001
9 mo 4.7 (1.8 to 13.9) 52.0 (40.4 to 71.1) −45.0 (−53.1 to −36.7) <.001
18 mo 11.3 (3.5 to 26.5) 54.0 (43.4 to 69.7) −40.7 (−49.1 to −30.3) <.001
Weight change index, fold change in BMI percentile
from baseline visit
9 mo 1.9 (0.6 to 22.1) 1.0 (0.9 to 1.1) 0.9 (0.0 to 4.4) .06
18 mo 5.8 (2.7 to 36.1) 1.1 (0.9 to 1.2) 4.6 (2.5 to 13.1) <.001
Baseline visit ghrelin measurements, pg/mL
Fastinge 821.5 (668.4 to 1013.4) 583.5 (374.3 to 852.6) 216.6 (75.7 to 352.9) .002
Postprandiale
0.5 h 667.0 (563.3 to 814.2) (n = 33) 467.4 (373.9 to 641.6) (n = 32) 167.0 (40.7 to 274.2) .008
1h 611.5 (505.1 to 705.8) (n = 30) 475.8 (349.2 to 607.4) (n = 31) 119.9 (19.8 to 224.3) .02
2h 608.9 (464.5 to 727.1) (n = 35) 413.4 (325.1 to 604.0) (n = 33) 164.3 (65.4 to 263.9) .003
Composite AUC, pg/mL × 2 hf 1389.4 (1082.5 to 1646.4) 958.5 (743.0 to 1234.5) 327.9 (119.6 to 543.9) .003
c
Abbreviations: AN, anorexia nervosa; AUC, area under the curve; BMI, body mass index Race and ethnicity data were self-reported using fixed categories. For non-White
(calculated as weight in kilograms divided by height in meters squared); HC, healthy individuals, ethnicity breakdown is not shown because all individuals identified as
control; NA, not applicable. non–Hispanic or Latina.
a d
Groups were compared using the Wilcoxon rank sum test. Group differences in Numbers of participants with visit interval and BMI percentile data at respective study
medians computed with the Hodges-Lehmann estimation are shown with visits correspond to the numbers of participants completing each visit.
nonparametric 95% CIs, computed as the median of the set of differences between e
Total ghrelin measures at time points surrounding the test meal at baseline visit.
each value in the AN subgroup and each value in the HC subgroup. Two-tailed P < .05 Fasting and 2-hour postprandial ghrelin measurements were available for all
was considered significant. participants, and sample sizes of available data for 0.5 and 1 hour after the test meal are
b
To compare race and ethnicity, nonzero and nonoverlapping categories of Asian, White indicated.
Hispanic or Latina, White Not Hispanic or Latina, and other were used with the χ2 test. f
Composite AUC index for all available ghrelin measurements spanning 2 hours were
Similarly, to compare number of follow-up completers per visit, nonoverlapping calculated using the trapezoid method.
categories of individuals who completed follow-up at month 9 but not at month 18,
completed follow-up at month 18 but not at month 9, and completed both follow-up
visits were used with the χ2 test.

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Odds ratios (ORs) are easier to interpret than LMM estimates for clinical outcomes such as
changes in body weight, which can be dichotomized to derive clinically informative outcome
categories (ie, 10% weight gain); however, dichotomization of a continuous outcome can result in
loss of information. We derived OR estimates for longitudinal weight gain positively associated with
ghrelin AUC directly from LMMs.21
To test the robustness of the primary analysis, we performed a complementary set of sensitivity
analyses using LMMs with (1) outlier removal, (2) different sets of covariates with ghrelin AUC, (3)
same set of covariates with individual ghrelin measurements, (4) alternatively defined outcome
variable for longitudinal change in body weight, (5) alternate assumptions of variance estimators, and
(6) alternatively defined random-effects terms. Because ORs were derived for fixed-effect terms
only, 3 of the 17 LMMs from sensitivity analyses that tested how different random-effect model terms
affect the model fit are not shown in the summary table for clarity of presentation of other sensitivity
analyses. Each random-effect term was reduced, and likelihood ratio tests of model reductions were
assessed with sequential analysis of variance decomposition of random effects. In LMMs with
alternate random effect terms, we observed associations consistent with the main LMM (P < .05 for
all 3 LMMs). Moreover, multivariable linear regression models (LMs) for 9- and 18-month visits were
tested to assess the sensitivity of repeated-measures approach in LMMs. In addition, we tested the
sensitivity of our approach to derive the ORs directly from LMMs by dichotomizing the continuous
outcome with a range of cutoff values for percentage of weight gain in a logistic regression approach
with univariate generalized linear mixed-effects models (GLMMs) with baseline ghrelin AUC as the
positively associated independent variable.21 Lastly, to explore the possibility of large effects of
ghrelin AUC diluting effects of associated diagnostic groups in the LMM, we performed subgroup
analyses with subsets of data limited to participants with AN or HCs. We limited subgroup analysis to
LMMs with 3 terms, with ghrelin AUC and sets of 2 covariate terms from the main model (not
including diagnosis).
All multivariable LMMs and LMs were assessed with stepwise backward deletion of single terms
to assess how dropping each term would affect model fit. Outliers beyond 2 SDs of the mean weight
change index were assessed but not removed except when indicated (ie, sensitivity analysis with
outlier removal and models using subsets of data as in simple LMs for each follow-up and LMMs for
subgroup analyses) to mitigate the potential for amplified outlier effects in models with fewer data
points as well as to prevent uneven outlier effects in models with data subsets, which could introduce
undue bias (ie, outliers of weight outcome only in the AN group or greater proportion of outliers at 9
months than 18 months). Similar to the a priori hypothesis tested with the primary analysis, post hoc
hypothesis testing for sensitivity and subgroup analyses used a 2-sided significance level of P < .05

Figure 1. Association Between Baseline Ghrelin and Prospective Change in Body Weight

4
Weight change index

2
AN

1 Ghrelin area under the curve (AUC) is plotted against

the weight change index, defined as body mass index
HC percentile (surrogate of body weight) at follow-up
0
relative to baseline. Weight change index values shown
are model adjusted for age, race, and interval duration
between baseline and follow-up. Data from 68
500 1000 1500 2000 2500 3000 individuals (124 data points) are shown. AN indicates
Ghrelin AUC, pg/mL × 2 h anorexia nervosa; HC, healthy control.

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together with corrections for multiple comparisons using the Benjamini-Hochberg method to
mitigate the risk of chance discoveries (type I errors). Even so, sensitivity and subgroup analyses
should be interpreted as exploratory given the post hoc nature.
Statistical analyses were performed between January and August 2022 with R and RStudio,
versions 4.0.0 and 0.99.902 (R Foundation for Statistical Computing).22 Additional details are
reported in the eMethods in Supplement 1.

Results
A total of 198 individuals consented or assented to participate and 76 (38.4%) did not complete the
baseline study visit: 47 failed the screen (see reasons in the eMethods in Supplement 1), 11 were lost
to follow-up, and 18 withdrew consent or assent before completing the baseline visit. Of the 122 who

Table 2. Sensitivity Analyses for the Final Modela

Regression model OR (95% CI) P value

Reference
Main model
Univariate 1.81 (1.29-2.49) .005
Multivariableb 2.35 (1.43-3.73) .004
Outlier removal in the data set
Outliers − SD methodc 4.49 (2.61-7.34) <.001
Ghrelin measurements instead of ghrelin AUC
Fasting 2.17 (1.31-3.50) .01
0.5 h 2.81 (1.70-4.48) <.001
1h 4.27 (2.47-7.00) <.001 Abbreviations: AUC, area under the curve; BMI, body
2h 2.17 (1.32-3.47) .01 mass index (calculated as weight in kilograms divided
by height in meters squared); OR, odds ratio.
Alternatively defined weight change indexes as outcome of interest
a
P values are from the Wald χ2 test for the fixed
BMI
effects of respective ghrelin term in the linear mixed-
z Score delta change 2.63 (1.60-4.20) <.001
effects regression model (LMM) and are not adjusted
Percentile delta fold change 2.35 (1.43-3.73) .004 for multiple comparisons. Multiple comparisons
Percentile log fold change 2.96 (1.79-4.74) <.001 adjustment of the false discovery rate did not change
Alternative sets of covariates results. For all LMMs in sensitivity analyses, except
Tanner stage when testing for effects of removing the outliers, no
By breast development instead of age 2.43 (1.49-3.85) .002 outliers were removed to avoid introducing bias.
b
By genital development instead of age 2.42 (1.49-3.82) .002 All parameters were kept the same as the reference
Ethnicity main multivariable model of the study with single
change as described for each sensitivity analysis.
Instead of race 2.50 (1.53-3.97) .002
c
From the data set with 124 data points, 6 outlier
With race as nonoverlapping categories 2.37 (1.45-3.77) .004
weight change indexes (<5%) from individuals with
BMI percentiles at baseline instead of diagnostic group 2.31 (1.44-3.61) .003
anorexia nervosa (AN) were removed before
Alternative assumptions of variance estimators modeling. For 1 individual, data from both visits were
Restricted maximum likelihood instead of maximum likelihood 2.35 (1.43-3.73) .006 identified as outliers; for 3 individuals, data from only
Linear models using data from specified follow-up visit onlyd 1 follow-up visit were identified as outliers; and for 1
Ghrelin at 9 mo individual who only had single follow-up data, this
Fasting 5.28 (2.31-10.67) <.001 was identified as an outlier, resulting in the final
sample size of 33 for the AN subgroup. No healthy
0.5 h 10.77 (4.35-22.21) <.001
control participants had weight change values in the
1h 8.42 (3.43-17.37) <.001
extremes (n = 33).
2h 11.52 (4.65-23.80) <.001 d
Linear regression models (LMs) were used to confirm
AUC 8.32 (3.51-16.84) <.001
the findings from the LMMs using weight change
Ghrelin at 18 mo index at a single follow-up visit only, adjusted for
Fasting 2.42 (1.14-4.81) .02 diagnostic group, age, race, and follow-up duration.
0.5 h 3.29 (1.55-6.36) .001 The LMs were built with a data set after removal of
1h 2.60 (1.22-5.11) .01 the 6 outlier data points before performing the
analyses. There were 4 and 2 outlier data points in
2h 3.83 (1.74-7.62) <.001
the 9- and 18-month follow-up data with the final
AUC 3.02 (1.43-5.90) .003
data set from 59 and 61 individuals, respectively.

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completed baseline visit procedures, 9 were lost to follow-up and 8 withdrew consent or assent
before completing the 9-month study visit, resulting in data from 105 individuals (86.1%) who were
followed up longitudinally for at least 9 months. Of those followed up longitudinally, 70 met the
criteria for the AN or HC group. Weight or ghrelin data were not available for 2 individuals with AN,
resulting in available data from a total of 68 individuals.

Baseline Characteristics
A total of 68 girls and young women (11 [16%] Asian, 4 [6%] Hispanic or Latina, 51 [75%] White [non–
Hispanic or Latina], and 2 [3%] other race or ethnicity), including 35 with AN (median [IQR] age, 20.1
[18.5-21.0] years) and 33 HCs (median [IQR] age, 18.7 [14.7-19.4] years) of similar Tanner stage, were
included in this study. The AN group was significantly older than the HC group by 1.6 years (95% CI,
0.5-2.8 years; P = .005). The AN and HC groups were comparable in race and ethnicity, Tanner
stages, number completing follow-up, and duration of follow-up. For individuals with AN, we
observed lower BMI percentiles at all visits (between-group difference for baseline visit, −51.8; 95%
CI, −58.8 to −46.1; P < .001) (Table 1) and higher baseline visit ghrelin AUC (between-group
difference in medians, 327.9 pg/mL; 95% CI, 119.6-543.9 pg/mL × 2 hours; P = .003). The AN group
gained weight with median BMI percentile fold changes of 1.9 (IQR, 0.6-22.1) at approximately 9
months and 5.8 (IQR, 2.7-36.1) at approximately 18 months, whereas the HC group had relatively
stable weights over time with median BMI percentile fold changes of 1.0 (IQR, 0.9-1.1) at
approximately 9 months and 1.1 (IQR, 0.9-1.2) at approximately 18 months. See Table 1 for clinical and
demographic characteristics of the study participants.

Associations Between Baseline Ghrelin and Longitudinal Weight Trajectory

We applied LMMs to estimate the longitudinal weight gain associated with ghrelin AUC. Univariate
LMM indicated a statistically significant association (OR, 1.81; 95% CI, 1.29-2.49; P = .005). Adjusting
for covariation with diagnosis, age, race, and follow-up duration in the main multivariable LMM, the
OR of the association was 2.35 (95% CI, 1.43-3.73; P = .004). We observed no statistically significant
associations for the covariate terms in the main LMM. Results of the main LMM are in Figure 1; raw
data not adjusted for covariation appear in eFigure 1 in Supplement 1.

Figure 2. Subgroup Analyses of Baseline Ghrelin and Prospective Change in Body Weight

OR Favors Favors
Covariates (95% CI) weight loss weight gain P value
BMI percentile and follow-up duration
AN 2.57 (1.46-4.20) .003
HC 1.09 (0.67-1.75) .92
BMI percentile and age
In models stratified by subgroups, ghrelin area under
AN 2.73 (1.53-4.52) .001
the curve (AUC) was positively associated with future
HC 0.98 (0.60-1.61) .79
weight gain only in the anorexia nervosa (AN)
BMI percentile and race
subgroup, unadjusted and after adjusting for any 2
AN 2.52 (1.45-4.11) .003
combinations of the covariates from the final model.
HC 1.05 (0.64-1.72) .97
Outliers were removed for subgroup analyses resulting
Age and follow-up duration
in data points from 33 participants with AN and 33
AN 2.97 (1.67-4.88) <.001
healthy controls (HCs). Subgroup models could not be
HC 1.15 (0.72-1.81) .78
adjusted for the same set of covariates in the main
Age and race
5-term model given the sample sizes. Given the
AN 2.86 (1.61-4.71) <.001
exploratory nature of subgroup analysis, analysis
HC 0.96 (0.58-1.60) .75
expanded the hypothesis testing to include linear
Race and follow-up duration
mixed models (LMMs) that replace ghrelin AUC with
AN 2.86 (1.65-4.60) <.001
individual ghrelin measurements, which are shown in
HC 1.16 (0.72-1.85) .78
eFigure 3 in Supplement 1. Findings from the 60 LMMs
0.5 1 10 were consistent with the models with ghrelin AUC
OR (95% CI) term as shown here. BMI indicates body mass index.

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We tested the validity and robustness of the multivariable LMM assessing our a priori
hypothesis through an exploratory series of complementary sensitivity analyses. We found
statistically significant positive associations between baseline ghrelin and longitudinal weight gain in
all 27 regression models (Table 2). Validity of deriving ORs from LMMs was assessed with GLMMs
with dichotomization of the outcome with 21 different weight cutoffs (eFigure 2 in Supplement 1).
Lastly, we conducted subgroup analyses to test the association between diagnostic group and
weight trajectory that may have been overshadowed by ghrelin AUC in the main model. We found
statistically significant positive associations between ghrelin and longitudinal weight gain only in the
AN subgroup and not for HCs, which was a consistent finding across 60 LMMs tested for subgroups
(12 LMMs with ghrelin AUC in Figure 2; 48 other LMMs in eFigure 3 in Supplement 1), with no
changes after correction for multiple comparisons (false discovery rate P < .025).

Discussion
This prospective cohort study found that baseline ghrelin was positively associated with longitudinal
weight gain in girls and young women with AN in community settings during approximately 18
months of follow-up. Of note, robust associations in our extensive series of sensitivity analyses
support the validity of results regardless of modeling with ghrelin measurements from fasting or
postprandial time points or the composite index and regardless of how longitudinal change in body
weight is defined. Although the primary analysis did not find statistically significant results of the
diagnostic grouping jointly with ghrelin, exploratory subgroup analyses support the specificity of
association only in AN.
Our finding that ghrelin is associated with longitudinal weight gain in AN is in stark contrast to
prior longitudinal investigations in other human populations, and this is the first, to our knowledge,
to report this association in those older than infants. Prior studies8-10 have consistently
demonstrated the lack of association between ghrelin and weight trajectory despite the short-term
effects of ghrelin in stimulating food intake in humans7 and preclinical evidence for a weight
regulatory role of ghrelin. The only prior human study11 that observed a positive association between
ghrelin and prospective weight gain was in infants born small for gestational age, with the association
observed only with ghrelin measurements following glucose administration but not for fasting
ghrelin. Although we observed robust associations across fasting and postprandial time points, we
note that AN and small for gestational age cohorts have low body weights relative to expected for
developmental stage based on population norms. It is possible that having very high levels of
circulating ghrelin in AN (an observation in agreement with prior studies13) together with
vulnerability from extremes of low body weights may exert an effect on energy homeostasis and
modulate weight-regulatory mechanisms in this vulnerable population.
Large-scale genomic investigations have identified heritable risk factors for AN, which implicate
genetic loci associated with metabolic regulation.23 Ghrelin is known to have multifaceted roles in
regulating glucose metabolism and is being investigated as a potential new avenue of therapeutics
development for metabolic diseases such as type 2 diabetes.24 In addition, genes encoding
components of the ghrelin signaling pathway have been studied as risk factors for AN, with 1 study25
reporting that specific single-nucleotide polymorphisms of the ghrelin gene were associated with
longitudinal weight recovery in AN. As such, effects of ghrelin signaling may be associated with the
course of illness in AN, but this should be interpreted with caution because evidence regarding
effects of ghrelin signaling on longitudinal weight gain remains limited.
For instance, a prior randomized clinical trial3 evaluated the effects of a ghrelin receptor agonist,
relamorelin, on body weight outcome for 22 women with AN (mean age, 28.9 years). Ghrelin agonist
treatment resulted in greater weight increases after 4 weeks compared with placebo, with means
(SEMs) of 0.86 (0.40) and 0.04 (0.28) kg, but between-group difference in means was not
statistically significant (P = .07). This finding is in line with our findings regarding the positive
association between endogenous levels of ghrelin and longitudinal weight gain. Even so, it remains

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 JAMA Network Open | Psychiatry Association Between Ghrelin and Body Weight Trajectory in Anorexia Nervosa

unknown whether ghrelin signaling has a direct effect on weight gain in AN, and additional studies to
understand ghrelin biology specific to AN are warranted given the urgent need for new treatments
because of the lack of US Food and Drug Administration–approved medications for this illness, which
has one of the highest mortality rates in psychiatry.

Limitations
This study has some limitations. Although the cohort age range spans the full developmental
spectrum when AN typically presents, the generalizability of our results is limited by the relatively
modest sample size and lack of racial, ethnic, and gender diversity. Larger longitudinal studies are
needed to confirm the association of elevated endogenous ghrelin with weight trajectory in AN. We
also acknowledge the limitations in deriving ORs from LMMs, because the method was developed
without accounting for the variance of random effects, which could result in an overestimation in
applications to LMMs despite the benefits over GLMMs. This limitation is mitigated by rigorous
hypothesis testing directly with LMMs together with extensive and complementary sensitivity
analyses, including a series with GLMMs that support the validity and robustness of our findings.
Nevertheless, the use of ORs was intended to enhance the interpretability of the results for an
outcome variable critical to AN in clinical settings.

Conclusions
In this cohort study, circulating levels of ghrelin in adolescent girls and young women with AN were
positively associated with prospective weight gain during 18 months of follow-up. This study
provides evidence supporting further research into the role of ghrelin biology in weight regulation
in AN.

ARTICLE INFORMATION
Accepted for Publication: February 7, 2023.
Published: March 24, 2023. doi:10.1001/jamanetworkopen.2023.4625
Open Access: This is an open access article distributed under the terms of the CC-BY License. © 2023 Kim YR
et al. JAMA Network Open.
Corresponding Author: Youngjung R. Kim, MD, PhD, Center for Quantitative Health, Department of Psychiatry,
Massachusetts General Hospital and Harvard Medical School, 185 Cambridge St CPZN 6300, Boston, MA 02114
(ykim82@mgh.harvard.edu).
Author Affiliations: Center for Quantitative Health, Department of Psychiatry, Massachusetts General Hospital
and Harvard Medical School, Boston (Kim, Perlis); Neuroendocrine Unit, Department of Medicine, Massachusetts
General Hospital and Harvard Medical School, Boston (Lauze, Slattery, Lawson); Division of Women’s Health,
Department of Medicine, Brigham and Women’s Hospital, Boston, Massachusetts (Holsen); Department of
Psychiatry, Brigham and Women’s Hospital and Harvard Medical School, Boston, Massachusetts (Holsen); Eating
Disorders Clinical and Research Program, Department of Psychiatry, Massachusetts General Hospital and Harvard
Medical School, Boston (Breithaupt, Stern, Thomas, Eddy); Department of Psychiatry, Massachusetts General
Hospital and Harvard Medical School, Boston (Fava); Department of Pediatrics, Division of Pediatric
Endocrinology, Massachusetts General Hospital and Harvard Medical School, Boston (Misra).
Author Contributions: Dr Kim had full access to all of the data in the study and takes responsibility for the integrity
of the data and the accuracy of the data analysis. Drs Lawson, Misra, and Eddy contributed equally to the
manuscript.
Concept and design: Thomas, Lawson, Misra, Eddy.
Acquisition, analysis, or interpretation of data: All authors.
Drafting of the manuscript: Kim, Eddy.
Critical revision of the manuscript for important intellectual content: All authors.
Statistical analysis: Kim.

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 JAMA Network Open | Psychiatry Association Between Ghrelin and Body Weight Trajectory in Anorexia Nervosa

Obtained funding: Lawson, Misra, Eddy.

Administrative, technical, or material support: Kim, Lauze, Breithaupt, Stern, Thomas, Lawson.
Supervision: Perlis, Fava, Thomas, Lawson, Misra, Eddy.
Conflict of Interest Disclosures: Dr Kim has not done any personal consulting; any consulting she has done has
been on behalf of Massachusetts General Hospital (MGH). Dr Perlis reported receiving personal fees from
Genomind, Belle Artificial Intelligence, Psy Therapeutics, Burrage Capital, and Circular Genomics outside the
submitted work. Dr Breithaupt reported receiving personal fees from Otsuka Pharmaceutical outside the
submitted work. Dr Fava reported receiving research support from Acadia Pharmaceuticals, Aditum Bio
Management Company, Allergan, Alkermes, Altimate Health Corporation, Alto Neuroscience, Ancora Bio, Angelini
S.p.A., Aptinyx, Arbor Pharmaceuticals, Avanir Pharmaceuticals, Axsome, Benckiser Pharmaceuticals, BioClinica,
Biogen, BioHaven, BioShin Ltd, Cambridge Science Corporation, Centrexion Therapeutics Corporation, Cerecor,
Cybin IRL Ltd, Eliem Therapeutics Ltd, Gate Neurosciences, GenOmind, Gentelon, Happify, Johnson & Johnson,
Lundbeck, Marinus Pharmaceuticals, Methylation Sciences, Millennium Pharmaceutics, Minerva Neurosciences,
Neuralstem, Neurocrine Biosciences, NeuroRX, Novaremed, Novartis, Otsuka, Pfizer, Premiere Research
International, Praxis Precision Medicines, Protagenic Therapeutics, Relmada Therapeutics, Reckitt, Shenox
Pharmaceuticals, Stanley Medical Research Institute, Taisho, Takeda, University of Michigan, Vistagen, WinSanTor,
Xenon Pharmaceuticals, National Institute of Drug Abuse, National Institutes of Health, National Institute of
Mental Health, and Patient-Centered Outcomes Research Institute outside the submitted work; lecturing at Global
Medical Education Mood Disorders Summit, November 2020; having equity holdings in Compellis, Neuromity, Psy
Therapeutics, and Sensorium Therapeutics; holding patents for Sequential Parallel Comparison Design, licensed
by MGH to Pharmaceutical Product Development, (US_7840419, US_7647235, US_7983936, US_8145504,
US_8145505); and having patent application for a combination of Ketamine Plus Scopolamine in Major Depressive
Disorder, licensed by MGH to Biohaven; having patents for pharmacogenomics of Depression Treatment with
Folate (US_9546401, US_9540691); and having copyrights for the MGH Cognitive & Physical Functioning
Questionnaire, Sexual Functioning Inventory, Antidepressant Treatment Response Questionnaire,
Discontinuation-Emergent Signs & Symptoms, Symptoms of Depression Questionnaire, and SAFER from Belvio,
Lippincott, Williams & Wilkins, Wolkers Kluwer, and World Scientific Publishing Co. Dr Fava has not done any
personal consulting; any consulting he has done has been on behalf of MGH, except for Sensorium Therapeutics.
The most up-to-date lifetime disclosures for Dr Fava can be viewed online (https://mghcme.org/faculty). Dr
Lawson reported receiving grants from Tonix Pharmaceuticals, serving on a scientific advisory board and receiving
stock options from OXT Therapeutics, and receiving personal fees from UpToDate (royalties) outside the
submitted work. Dr Misra reported receiving personal fees from AbbVie, Ipsen, and Sanofi outside the submitted
work. No other disclosures were reported.
Funding/Support: The parent study was supported by grant R01 MH103402 from the National Institute of Mental
Health (Drs Misra, Eddy, and Lawson). This work was supported in part by grants K99 MH127366 (Dr Kim), K24
MH120568 (Dr Lawson), R01 MH116270 (Dr Perlis), R01 MH123804 (Dr Perlis), K23 MH127465 (Dr Breithaupt),
and R03 MH126143 (Dr Breithaupt) from the National Institute of Mental Health and grants P30 DK040561 (Dr
Lawson) and R01 DK104772 (Dr Holsen) from the National Institute of Diabetes and Digestive and Kidney
Diseases, the Brain Behavior Research Foundation (Dr Breithaupt), the Phyllis & Jerome Lyle Rappaport
Foundation (Dr Kim), and the MGH Department of Psychiatry (Dr Kim).
Role of the Funder/Sponsor: The funding sources had no role in the design and conduct of the study; collection,
management, analysis, and interpretation of the data; preparation, review, or approval of the manuscript; and
decision to submit the manuscript for publication.
Disclaimer: Dr Perlis is an Associate Editor for JAMA Network Open but was not involved in the editorial review or
decision on this manuscript.
Data Sharing Statement: See Supplement 2.
Additional Contributions: We are grateful to all study participants for their involvement in the study. We thank our
colleagues at the MGH Eating Disorders Clinical and Research Program, Division of Pediatric Endocrinology,
Division of Adolescent Medicine, and Neuroendocrine Unit, for their support of participant recruitment, sample
collection, and experimental work.

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SUPPLEMENT 1.
eMethods. Supplemental Methods
eFigure 1. Association Between Baseline Ghrelin and Prospective Change in Body Weight Without Adjustment for
Covariates
eFigure 2. Sensitivity Analysis for Deriving Odds Ratios From Linear Regression Models
eFigure 3. Expanded Subgroup Analyses of Baseline Ghrelin and Prospective Change in Body Weight

SUPPLEMENT 2.
Data Sharing Statement

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