Original Article

Longitudinal Growth Differentiation Factor 15 (GDF15)
and Long-term Intraocular Pressure Fluctuation in

Glaucoma: A Pilot Study

Jonathan B. Lin, MD, PhD1; Arsham Sheybani, MD1; Andrea Santeford, MS1; Rajendra S. Apte, MD, PhD1,2,3

1Departments of Ophthalmology and Vision Science, Washington University, USA
2Departments of Developmental Biology, Washington University, USA

3Departments of Medicine, Washington University, USA

Abstract
Purpose: Growth Differentiation Factor 15 (GDF15) was previously identified as a molecular
marker of retinal ganglion cell stress in rodent models of glaucoma and was elevated in the
aqueous humor (AH) of patients with primary open-angle glaucoma as a possible risk factor
for glaucoma progression. The purpose of this study was to determine whether changes
in the AH GDF15 levels were associated with intraocular pressure (IOP) changes in eyes
undergoing glaucoma surgery.
Methods: Here, we performed a prospective, longitudinal pilot study in nine patients to
determine whether changes in AH GDF15 levels from surgery to post-surgery follow-up were
associated with IOP fluctuation. An initial AH sample was taken from the peripheral corneal
paracentesis during planned glaucoma surgery, and a second sample was taken during an
outpatient follow-up visit, approximately six months later.
Results: There was a statistically significant correlation between GDF15 fold change and IOP
standard deviation (r = 0.87, P = 0.003), IOP range (r = 0.87, P = 0.003), and maximum IOP (r
= 0.86, P = 0.003). There was no correlation between the GDF15 fold change and baseline
IOP (r = 0.50, P = 0.17), final IOP (r = 0.038, P = 0.92), or mean IOP (r = 0.40, P = 0.28).
Conclusion: Our findings in this pilot study suggest that longitudinal changes in AH GDF15
may be associated with IOP fluctuation during the postoperative period. Further studies are
necessary to corroborate these findings in a larger patient population and to explore the
possibility that AH GDF15 may be used not only to improve treatment algorithms but also as
a surrogate endpoint in clinical trials.

Keywords: GDF15; Glaucoma; Neurodegeneration; Molecular Markers

J Ophthalmic Vis Res 2021; 16 (1): 21–27

Correspondence to:

Rajendra S. Apte, MD, PhD. Washington University, 660
South Euclid Ave., Campus Box 8096, St. Louis, MO
63110, USA.
E-mail: apte@wustl.edu
Received: 17-07-2020 Accepted: 16-11-2020

Access this article online

Website:
https://knepublishing.com/index.php/JOVR

DOI:
10.18502/jovr.v16i1.8245

INTRODUCTION

Glaucoma is a neurodegenerative disease
characterized by progressive death of retinal

This is an open access journal, and articles are distributed under the terms of
the Creative Commons Attribution-NonCommercial-ShareAlike 4.0 License, which
allows others to remix, tweak, and build upon the work non-commercially, as
long as appropriate credit is given and the new creations are licensed under the
identical terms.

How to cite this article: Lin JB, Sheybani A, Santeford A, Apte RS.
Longitudinal Growth Differentiation Factor 15 (GDF15) and Long-term
Intraocular Pressure Fluctuation in Glaucoma: A Pilot Study. J Ophthalmic
Vis Res 2021;16:21–27.

© 2021 LIN ET AL. THIS IS AN OPEN ACCESS ARTICLE DISTRIBUTED UNDER THE CREATIVE COMMONS ATTRIBUTION LICENSE | PUBLISHED BY

KNOWLEDGE E 21

http://crossmark.crossref.org/dialog/?doi=10.18502/jovr.v16i1.8245&domain=pdf&date_stamp=2019-07-17
https://knepublishing.com/index.php/JOVR


Longitudinal GDF15 and IOP Fluctuation in Glaucoma; Lin et al

ganglion cells (RGCs). Although it is currently the
second leading cause of blindness worldwide,[1]
the molecular pathogenesis of RGC death remains
elusive. Therefore, interventions are currently
centered on lowering intraocular pressure (IOP), a
risk factor for disease progression.[2] Most clinical
decision-making is based upon measuring IOP and
surrogates of glaucomatous neurodegeneration,
such as Humphrey visual field, cup-to-disc ratio,
and nerve fiber layer thickness. Unfortunately,
these surrogate metrics are imprecise in their
ability to quantify disease severity and, in some
cases, are subjective and unreliable. Therefore,
there is a clinical need for molecular markers that
measure RGC health and stress prior to cell death
to guide optimal medical and surgical management
of glaucoma patients.

It was previously reported that Growth
Differentiation Factor 15 (GDF15), a member of
the Transforming Growth Factor beta (TGF-β)
superfamily, is a molecular marker of RGC stress
in rodent models of glaucoma.[3] Validation studies
in well-characterized human patients showed
that GDF15 levels not only were elevated in
the aqueous humor (AH) of primary open-angle
glaucoma (POAG) patients compared to control
patients without glaucoma but also increased
stepwise with increasing visual field loss by
Hodapp-Parrish-Anderson staging.[3] However,
because of the cross-sectional study design, they
were unable to determine whether changes in AH
GDF15 levels were associated with IOP changes,
which have been reported as possible risk factors
for progression. To explore this possibility, we
performed a prospective, longitudinal pilot study
to determine whether changes in AH GDF15
levels over a follow-up period of approximately six
months are associated with IOP changes in eyes
undergoing glaucoma surgery.

METHODS

We recruited nine participants from one large
academic institution. All patients gave written
informed consent. This study was approved by
the Institutional Review Board (IRB) of the Human
Research Protection Office (HRPO) of the local
Ethics Committee. All procedures adhered to the
tenets of the Declaration of Helsinki. Patients
were included if they had any form of glaucoma,
including POAG or secondary glaucoma, and
were determined to be candidates for Molteno®

glaucoma implant (Molteno Ophthalmic Limited,
Dunedin, New Zealand) or Ahmed® glaucoma valve
(New World Medical, Rancho Cucamonga, CA)
surgery. Eyes were excluded if there was active
inflammatory eye disease, any retinopathy, or any
optic nerve degeneration from non-glaucomatous
causes. To determine appropriate sample size,
we performed a power analysis using G*Power
3.1.9.2.[15] Estimating an effect size of r = 0.75 based
on previous data, we calculated a sample size of N
= 9 to achieve 80% power at a two-tailed alpha of
0.05.

Two AH samples were obtained from each
patient. The first AH sample was obtained in the
operating room during planned glaucoma surgery.
Briefly, a blunt cannula attached to a tuberculin
syringe was inserted into the initial peripheral
corneal paracentesis and used to remove 50–
100 µl of AH. The second AH sample was
obtained during a clinic visit, approximately six
months after the initial surgery. Briefly, using sterile
technique, a needle on a syringe was used to
enter the anterior chamber temporally, anterior
to the limbus, to gently aspirate AH, with care
taken to not deform the anterior chamber. In both
cases, AH samples were immediately placed on
dry ice and then stored at –80ºC until further
analysis. We measured GDF15 levels of all AH
samples at the same time using the commercially
available human GDF15 Quantikine enzyme-linked
immunosorbent assay (ELISA) kit (R&D Systems), as
described previously.[3] The individual performing
GDF15 measurements (XXX) was masked to
demographic and clinical information to minimize
bias.

Demographic information, clinical information,
and IOP measurements were obtained by
retrospective chart review. All IOP values were
measured by Goldmann Applanation Tonometry,
performed by ophthalmologists or optometrists
who were masked to the study data. Participants
had IOP measurements taken as a part of routine
clinical care at postoperative day 1, postoperative
week 1, postoperative month 1, and additional
follow-up visits as clinically indicated. The primary
variables of interest were measures of IOP
fluctuation that have been previously reported
as risk factors for glaucoma progression, such as
IOP standard deviation, IOP range, and maximum
IOP. We also analyzed baseline IOP (measured at
the clinic visit prior to glaucoma surgery), final IOP
(measured at the same clinic visit during which the

22 JOURNAL OF OPHTHALMIC AND VISION RESEARCH VOLUME 16, ISSUE 1, JANUARY-MARCH 2021



Longitudinal GDF15 and IOP Fluctuation in Glaucoma; Lin et al

second AH sample was collected), and mean IOP
over the follow-up period.

We performed statistical analysis and data
visualization with R Version 3.6.2 and RStudio
Version 1.2.5003. To compare means between
two groups, we used the Mann–Whitney U test
due to small sample size. To compare pre- and
post-surgery number of medications, we used
the Wilcoxon signed-rank test. To determine
associations between continuous variables, we
calculated Pearson product-moment correlation
coefficients. Because of relatively small sample
sizes, we also calculated Kendall rank correlation
coefficients to confirm our results. We considered
P < 0.05 to be statistically significant.

RESULTS

Demographic and clinical characteristics of the
participants are shown in Table 1. There were four
male and five female participants. The mean age
was 71.0 years (standard deviation: 9.6 years).
Eight patients had POAG, while one patient had
glaucoma secondary to presumed herpes simplex
uveitis/trabeculitis, which had been inactive
for greater than three months. Three patients
underwent placement of a Molteno® glaucoma
implant; six patients underwent placement of
an Ahmed® glaucoma valve. Three patients
underwent surgery in their left eye; six in the
right eye. The mean follow-up duration was 183.4
days (standard deviation: 28.0 days, minimum:
131 days, maximum: 215 days). Patients were on
significantly fewer classes of medications after
surgery compared to before surgery (P = 0.013).

There was no significant correlation between
baseline AH GDF15 and baseline IOP, mean IOP,
or final IOP (P > 0.05). Similarly, there was
no significant correlation between follow-up AH
GDF15 and baseline IOP, mean IOP, or final IOP (P
> 0.05). Of the nine participants, six had increased
AH GDF15 levels over the follow-up interval,
while three had decreased AH GDF15 levels at
follow-up approximately six months later [Figure
1]. All participants had between four to nine IOP
measurements during the follow-up period [Figure
2]. When dichotomizing participants into those
who had increased (“GDF15 Up”; N = 6) versus
decreased (“GDF15 Down”; N = 3) AH GDF15 levels,
there were no statistically significant differences in
the baseline IOP, final IOP, mean IOP, IOP standard

deviation, IOP range, or maximum IOP (P > 0.05
by Mann–Whitney U tests). GDF15 fold change
from baseline to follow-up was not correlated with
baseline IOP, final IOP, or mean IOP [Figures 3A–
C]. In contrast, GDF15 fold change was strongly
correlated with IOP standard deviation, IOP range,
and maximum IOP [Figures 3D–F] with statistical
significance achieved with both parametric and
non-parametric tests.

DISCUSSION

In this prospective, observational pilot study, we
analyzed whether changes in AH GDF15 levels
from baseline to follow-up at approximately six
months were associated with IOP fluctuation.
Our findings suggest that AH GDF15 fold change
is indeed associated with IOP fluctuation. IOP
fluctuation that occurs over months to years
has been reported as a risk factor for visual
field progression in glaucoma in the Advanced
Glaucoma Intervention Study (AGIS),[4, 5] the
Collaborative Initial Glaucoma Treatment Study
(CIGTS),[6] and the Japanese Archive of Multicentral
Databases in Glaucoma (JAMDIG).[7] Thus, our
findings suggest that increases in AH GDF15
measurements may be associated with increased
risk of glaucoma progression. Although not all
studies have corroborated IOP fluctuation as a risk
factor for glaucoma progression, Kim and Caprioli
previously hypothesized that this discrepancy
may be due to higher mean IOP in some study
populations that could potentially mask the effect
of IOP fluctuation.[8]

Given this possible role of IOP fluctuation in
glaucoma progression, especially for patients who
show progression despite having IOPs near goal,
the ability to use a molecular marker such as GDF15
as a marker of long-term IOP fluctuation is highly
desirable. Routine IOP measurements to assess
for fluctuation is time-consuming given the need
for repeated clinic visits and is rarely performed
outside of clinical trials due to demands not only for
the clinician but also for patients and their families.
Home tonometer devices such as the Icare® HOME
tonometer (Icare USA, Raleigh, NC) are available
but have uncertain reliability. Additionally, devices
such as the Triggerfish® Contact Lens Sensor
(SENSIMED, Lausanne, Switzerland) can measure
changes in ocular dimensions thought to be related
to IOP but is typically used for only a 24-hr period
and is still experimental, as studies investigating

JOURNAL OF OPHTHALMIC AND VISION RESEARCH VOLUME 16, ISSUE 1, JANUARY-MARCH 2021 23



Longitudinal GDF15 and IOP Fluctuation in Glaucoma; Lin et al

Table 1. Demographic and clinical characteristics of study participants

Characteristic Value

Age, Mean ± SD𝑎 71.0 ± 9.6
Sex, N𝑏 (%)

Male 4 (44.4)

Female 5 (55.6)

Type of Glaucoma, N (%)

Primary open-angle glaucoma 8 (88.9)

Glaucoma secondary to inflammation 1 (11.1)

Type of Procedure, N (%)

Molteno® glaucoma implant 3 (33.3)

Ahmed® glaucoma valve 6 (66.7)

Study Eye, N (%)

OS 3 (33.3)

OD 6 (66.7)

Pre-surgery Medication Classes, Median (Range) 4 (2 – 4)

Post-surgery Medication Classes, Median (Range) 2 (0 – 3)𝑐

𝑎SD: standard deviation; 𝑏N: number of participants; 𝑐There is a significant difference between pre- and post-surgery number of
medication classes by the Wilcoxon signed rank test: P = 0.013

�

�

�

�

�

�

�

�

�

�

�

�

�

�

�

�

�

�

0

500

1000

1500

Baseline Follow−Up

A
H

 G
D

F
1

5
 (

p
g

/m
L

)

Figure 1. Of the nine participants, six had increased aqueous humor (AH) Growth Differentiation Factor 15 (GDF15) from baseline
to follow-up at approximately six months (shades of red), while three had decreased AH GDF15 (shades of blue). Circles denote
patients who received the Ahmed® glaucoma valve; squares denote patients who received the Molteno® glaucoma implant.

24 JOURNAL OF OPHTHALMIC AND VISION RESEARCH VOLUME 16, ISSUE 1, JANUARY-MARCH 2021



Longitudinal GDF15 and IOP Fluctuation in Glaucoma; Lin et al

�

�
� �

�

�

�

�
�

�

�

� �

�

�

�

�

�

�

�

�

�

�

�

�

�
�

�

�

�

��

�

�

�

�

�

�

�

�

�
� � �

�

�

�

��

�

�

�
�

�

�

�

�

�

�

�

�

�

�

�

�

�

GDF15 Down GDF15 Up

0 100 200 0 100 200

10

20

30

40

Time (days)

IO
P

 (
m

m
 H

g
)

Figure 2. Serial intraocular pressure measurements by Goldmann Applanation Tonometry for participants whose aqueous humor
(AH) Growth Differentiation Factor 15 (GDF15) increased (“GDF15 Up”; N = 6; shades of red) and those whose AH GDF15 decreased
(“GDF15 Down”; N = 3; shades of blue). Day 0 was set as the day of glaucoma surgery when the initial AH sample was collected.
Circles denote patients who received the Ahmed® glaucoma valve; squares denote patients who received the Molteno® glaucoma
implant.

their correlation with IOP measurements obtained
through other validated methods have yielded
mixed results.[9]

Although glaucoma is one of the leading
causes of blindness worldwide, identifying reliable
molecular markers has been challenging.[10]
This lack of molecular markers has led to
reliance on surrogate markers of glaucomatous
neurodegeneration for clinical decision-making,
even though these surrogate markers are
imprecise and sometimes unreliable. Additionally,
there is a great need for novel molecular markers of
RGC health that can be used as reliable surrogate
endpoints for clinical trials.[11] Although further
validation is necessary to demonstrate a direct
link to glaucoma progression, we propose that AH
GDF15 may be a molecular marker of long-term IOP
fluctuation that may be used in future therapeutic
trials.

One limitation of the present study is the
relatively small sample size. Although we achieved
the necessary sample size for adequate statistical
power, our small sample size does not permit us
to control for possible covariates, such as age

and gender. Another limitation of the study is
the heterogeneity of glaucoma subtype and the
type of surgery that these patients underwent.
We cannot rule out the possibility that differences
in the underlying disease pathophysiology or
underlying differences of the post-operative
IOP profiles of the Ahmed® glaucoma valve
versus the Molteno® glaucoma implant may
have influenced our findings. Future longitudinal
studies in larger populations are necessary to
address these limitations and may also incorporate
functional testing to directly measure glaucoma
progression.

One strength of our study is that we have a
well-characterized patient population for whom
we have longitudinal GDF15 measurements. This
within-subjects design allowed us to account
for inter-individual variability since GDF15
has shown to be elevated in other contexts,
such as neurodegenerative and cardiovascular
disease.[12, 13]

Although many groups have explored AH
biomarkers for numerous ocular diseases,[14]
many of these studies have analyzed samples

JOURNAL OF OPHTHALMIC AND VISION RESEARCH VOLUME 16, ISSUE 1, JANUARY-MARCH 2021 25



Longitudinal GDF15 and IOP Fluctuation in Glaucoma; Lin et al

� �GDF15 Down GDF15 Up

�

�

�

�

�

�

�

�

�

�
�

�

�

�

�

r=0.50, P=0.17

0

10

20

30

40

0 2 4 6

GDF15 Fold Change

B
a

s
e

li
n

e
 I
O

P
 (

m
m

 H
g

)

A

�

�

�

�

�

�

�

�

��

�

�

�

�

�

�

�

r=0.038, P=0.92

0

10

20

30

40

0 2 4 6

GDF15 Fold Change
F

in
a

l 
IO

P
 (

m
m

 H
g

)

B

�

�
��

�
�

�

�

�
��

�
� �

�

r=0.40, P=0.28

0

10

20

30

40

0 2 4 6

GDF15 Fold Change

M
e

a
n

 I
O

P
 (

m
m

 H
g

)

C

�

�

�

�

��
�

�
�

�

�

��
�

�
�

r=0.87, P=0.003

0

5

10

15

20

0 2 4 6

GDF15 Fold Change

IO
P

 S
tn

d
 D

e
v

 (
m

m
 H

g
)

D

�

�

�

�

��
�

�

�

�

�

��
�

�

�

r=0.87, P=0.003

0

10

20

30

40

50

0 2 4 6

GDF15 Fold Change

IO
P

 R
a

n
g

e
 (

m
m

 H
g

)

E

�

�

�

�

��
�

��

�

�

��
�

��

r=0.86, P=0.003

0

20

40

60

0 2 4 6

GDF15 Fold Change

M
a

x
 I
O

P
 (

m
m

 H
g

)

F

Figure 3. Aqueous humor Growth Differentiation Factor 15 (GDF15) fold change from baseline to six-month follow-up was not
correlated with baseline intraocular pressure (IOP; A), final IOP (B), or mean IOP (C). In contrast, there was a strong correlation
between GDF15 fold change and IOP standard deviation (stnd dev; D), IOP range (E), and maximum (max) IOP (F). r = Pearson
correlation coefficients. Similar significance levels were found with non-parametric Kendall rank correlation coefficients. Shaded
regions indicate 95% confidence interval bands. Circles denote patients who received the Ahmed® glaucoma valve; squares
denote patients who received the Molteno® glaucoma implant.

obtained during cataract or glaucoma surgery.
It is important to note that it is possible to
collect AH in the outpatient setting. This
procedure has minimal risks when performed
by an experienced practitioner and was well
tolerated by participants in this study. Although
it is somewhat invasive, we propose that the
ability to quantitatively assess RGC health may
outweigh any risks associated with such as a
procedure.

Resource Availability

The data analyzed in this study are available from
the corresponding author on reasonable request.

Financial Support and Sponsorship

None.

Conflicts of Interest

There are no conflicts of interest.

REFERENCES

1. Quigley HA, Broman AT. The number of people with
glaucoma worldwide in 2010 and 2020. Br J Ophthalmol
2006;90:262–267.

2. Ban N, Siegfried CJ, Apte RS. Monitoring
Neurodegeneration in glaucoma: therapeutic implications.
Trends Mol Med 2018;24:7–17.

3. Ban N, Siegfried CJ, Lin JB, Shui Y-B, Sein J, Pita-Thomas
W, et al. GDF15 is elevated in mice following retinal
ganglion cell death and in glaucoma patients. JCI Insight
2017;2: e91455.

4. Caprioli J, Coleman AL. Intraocular pressure fluctuation a
risk factor for visual field progression at low intraocular
pressures in the advanced glaucoma intervention study.
Ophthalmology 2008;115:1123–1129.e3.

5. Nouri-Mahdavi K, et al. Predictive factors for glaucomatous
visual field progression in the Advanced Glaucoma
Intervention Study. Ophthalmology 2004;111:1627–1635.

26 JOURNAL OF OPHTHALMIC AND VISION RESEARCH VOLUME 16, ISSUE 1, JANUARY-MARCH 2021



Longitudinal GDF15 and IOP Fluctuation in Glaucoma; Lin et al

6. Musch DC, et al. Intraocular pressure control and long-
term visual field loss in the Collaborative Initial Glaucoma
Treatment Study. Ophthalmology 2011;118:1766–
1773.

7. Fujino Y, et al. Evaluation of glaucoma progression
in large-scale clinical data: the Japanese Archive of
Multicentral Databases in Glaucoma (JAMDIG). Invest
Ophthalmol Vis Sci 2016;57:2012–2020.

8. Kim JH, Caprioli J. Intraocular pressure fluctuation: is it
important? J Ophthalmic Vis Res 2018;13:170–174.

9. Dunbar GE, Shen BY, Aref AA. The Sensimed Triggerfish
contact lens sensor: efficacy, safety, and patient
perspectives. Clin Ophthalmol 2017;11:875–882.

10. Bhattacharya SK, Lee RK, Grus FH, Group, the Seventh
ARVO/Pfizer Ophthalmics Research Institute Conference
Working Group. Molecular biomarkers in glaucoma. Invest
Ophthalmol Vis Sci 2013;54:121.

11. Medeiros FA. Biomarkers and surrogate endpoints:
lessons learned from glaucoma. Invest Ophthalmol Vis
Sci 2017;58:BIO20–BIO26.

12. Lindholm D, et al. Association of multiple biomarkers with
risk of all-cause and cause-specific mortality after acute
coronary syndromes: a secondary analysis of the PLATO
biomarker study. JAMA Cardiol 2018;3:1160–1166.

13. Maetzler W, et al. GDF15/MIC1 and MMP9 cerebrospinal
fluid levels in Parkinson’s disease and lewy body
dementia. PLOS ONE 2016;11:e0149349.

14. Tamhane M, Cabrera-Ghayouri S, Abelian G, Viswanath V.
Review of biomarkers in ocular matrices: challenges and
opportunities. Pharm Res 2019;36:40.

15. Faul F, Erdfelder E, Buchner A, Lang A-G. Statistical power
analyses using G*Power 3.1: tests for correlation and
regression analyses. Behav Res Methods 2009;41:1149–
1160.

JOURNAL OF OPHTHALMIC AND VISION RESEARCH VOLUME 16, ISSUE 1, JANUARY-MARCH 2021 27