Send Orders for Reprints to reprints@benthamscience.

net
The Open Ophthalmology Journal, 2014, 8, 39-47

39

Open Access

Glycolysis in Patients with Age-Related Macular Degeneration

Kanako Yokosako1, Tatsuya Mimura*,1, Hideharu Funatsu2, Hidetaka Noma3, Mari Goto1,
Yuko Kamei1, Aki Kondo1 and Masao Matsubara1
1

Department of Ophthalmology, Tokyo Women's Medical University Medical Center East, Tokyo, Japan

Department of Ophthalmology, Yachiyo Medical Center, Tokyo Women's Medical University, Chiba, Japan

Department of Ophthalmology, Hachioji Medical Center, Tokyo Medical University, Tokyo, Japan
Abstract: Purpose: Retinal adenosine triphosphate is mainly produced via glycolysis, so inhibition of glycolysis may
promote the onset and progression of age-related macular degeneration (AMD). When glycolysis is inhibited, pyruvate is
metabolized by lactic acid fermentation instead of entering the mitochondrial tricarboxylic acid (TCA) cycle. We
measured urinary pyruvate and lactate levels in patients with AMD.
Methods: Eight patients with typical AMD (tAMD group) and 9 patients with polypoidal choroidal vasculopathy (PCV
group) were enrolled. Urinary levels of pyruvate, lactate, -hydroxybutyrate, and -hydroxybutyrate were measured in all
patients.
Results: The mean urinary levels of pyruvate and lactate were 8.0 2.8 and 7.5 8.3 g/mg creatinine (reference values:
0.5-6.6 and 0.0-1.6), respectively, with the mean increase over the reference value being 83.6 51.1% and 426.5
527.8%, respectively. In 12 patients (70.6%), the lactate/pyruvate ratio was above the reference range. Urinary levels of hydroxybutyrate and -hydroxybutyrate were decreased by -31.9 15.2% and -33.1 17.5% compared with the mean
reference values. There were no significant differences of any of these glycolysis metabolites between the tAMD and
PCV groups. Multivariate analysis revealed that none of the variables tested, including patient background factors (age,
hypertension, diabetes, hyperlipidemia, cerebrovascular disease, alcohol, smoking, visual acuity, and AMD phenotype),
were significantly associated with the lactate/pyruvate ratio.
Conclusion: A high lactate/pyruvate ratio is a well-known marker of mitochondrial impairment, and it indicates poor
oxidative function in AMD. Our results suggest that increased lactate levels may be implicated in the pathogenesis of
AMD.

Keywords: Age-related macular degeneration, glycolysis, ketone body, lactate, pyruvate.

INTRODUCTION
Glycolysis is a critical pathway for the generation of
adenosine triphosphate (ATP), and involves the metabolism
of both glucose and lactate, an indicator of oxidative
capacity [1]. The glycolytic pathway produces pyruvate and
lactate from glucose in all mammals [1]. Under aerobic
conditions, pyruvate is converted to acetyl-CoA and CO2 in
the mitochondria and oxidized via the tricarboxylic acid
(TCA) cycle. However, pyruvate is converted to lactate by
lactate dehydrogenase (LDH) under anaerobic conditions
[2-4]. Thus, glycolysis leads to an increase of lactate
production under hypoxic conditions, and higher lactate
levels reflect tissue hypoxia. Hypoxic glycolysis is
consistently associated with increased tissue levels of lactate
and leads to various functional disorders [5-13].
Age-related macular degeneration (AMD) is a leading
cause of visual impairment and is also the major cause of
*Address correspondence to this author at the Department of Ophthalmology,
Tokyo Women's Medical University Medical Center East, 2-1-10 Nishiogu,
Arakawa-ku, Tokyo 116-8567, Japan; Tel: +81-3-3810-1111, Ext. 7765;
Fax: +81-3-3894-0282; E-mail: mimurat-tky@umin.ac.jp
1874-3641/14

blindness in the elderly. However, there have been few

reported investigation into the influence of the glycolytic
pathway on the progression of AMD. In the normal retina of
pigmented guinea pigs, lactate produced by glycolysis is
transferred from retinal glial (Mller) cells to the
photoreceptors, which then consume both lactate and glucose
for oxidative metabolism [14]. Cultured human Mller cells
mainly obtain ATP mainly from glycolysis in the presence of
both glucose and oxygen, and show low oxygen
consumption [15]. Inhibition of glycolysis is likely to be
detrimental to retinal cells because more than 50% of retinal
ATP is produced by the glycolytic pathway [15]. Although
only a few in vitro studies on the role of the glycolytic
pathway in the pathogenesis of AMD have been performed
so far [16], the findings suggested to us that impaired
glycolysis may lead to a reduction in energy metabolism of
retinal cells in AMD.
Accordingly, we hypothesized that decreased pyruvate
kinase activity in glycolysis would promote the development
and progression of AMD. To investigate this possibility, we
measured urinary levels of pyruvate and lactate in patients
with AMD. We also compared the urinary pyruvate/lactate
2014 Bentham Open

40

The Open Ophthalmology Journal, 2014, Volume 8

ratio between typical AMD (tAMD) and polypoidal

choroidal vasculopathy (PCV).
MATERIALS AND METHODS
Study Design
This was a prospective, nonrandomized, cross-sectional,
consecutive case series study conducted at the Tokyo
Women's Medical University Medical Center East hospital
and affiliated hospitals. This study was performed in
accordance with the Helsinki Declaration. Our institutional
review board approved the study and informed consent was
obtained from each subject.
Subjects
The study population consisted of seventeen patients with
AMD aged 75.5 4.8 years (mean SD), with an age range
of 66-84 years (Table 1). Seventeen patients with a clinical
diagnosis of typical wet-type AMD, who presented to
hospitals associated with Tokyo Women's Medical
University Medical Center East, were enrolled and were
divided into two groups. Eight patients with tAMD were
assigned to the tAMD group and 9 patients with PCV formed
the PCV group. There were no patients with retinal
angiomatous proliferation (RAP) in this study.
Exclusion criteria were (1) age 60 years, (2) high
myopia greater than 5 diopters, (3) best-corrected visual
acuity (BCVA) better than 0.20 logMAR (20/30) in the eye
affected by AMD, (4) other vitreoretinal diseases (i.e.,
vitreomacular traction syndrome, epiretinal membrane,
Table 1.

Yokosako et al.

macular hole, diabetic retinopathy, and retinal vein

occlusion), (5) amblyopia, (6) cataract > grade 3 according
to the Emery-Little classification of nuclear hardness [17],
and (7) Alzheimer disease or other dementias.
In each patient, the age, gender, medical history, and
ocular history were assessed at the initial visit. A diagnosis
of hypertension, coronary artery disease, or diabetes was
based on data from the medical records. Hypertension was
defined as current antihypertensive therapy or a blood
pressure > 140/90 mmHg. The diagnosis of diabetes was
based on information from the subjects or on use of
antidiabetic medications.
The background characteristics of the study population
are presented in Table 1. Among the 17 patients, 7 patients
(41.2%) had hypertension, 3 patients (17.6%) had diabetes, 1
patient (5.8%) had hyperlipidemia, and 2 patients (11.8%)
had a history of cerebrovascular disease. None of the patients
had coronary heart disease, renal disease, or liver disease.
With respect to ocular diseases, there was none of the
patients had glaucoma or diabetic retinopathy. There were no
significant differences of patient demographic or disease
characteristics between the tAMD and PCV groups
(Table 1).
Analysis of Urinary Metabolites
Measurement of urinary metabolites was performed with
gas chromatography/mass spectrometry (GC/MS) by US
BioTek Laboratories (Seattle, WA) according to the
company's published method [18]. The subjects were not

Clinical profile of the subjects.

Total

tAMD

PCV

P Value (tAMD vs PCV)

17

75.5 4.8

75.3 3.2

75.7 5.9

*0.4328

6/11

1/7

5/4

**0.0882

Refraction (D)

0.2 1.6

0.9 1.1

-0.4 1.8

*0.0564

LogMAR visual acuity

0.6 0.5

0.5 0.2

0.7 0.7

*0.1479

Hypertension

7 (41.2%)

3 (37.5%)

4 (44.4%)

**0.5806

Diabetes

3 (17.6%)

1 (12.5%)

2 (22.2%)

**0.5471

Hyperlipidemia

1 (5.8%)

0 (0.0%)

1 (11.1%)

**0.5294

Coronary heart disease

0 (0.0%)

0 (0.0%)

0 (0.0%)

**NA

Renal disease

0 (0.0%)

0 (0.0%)

0 (0.0%)

**NA

Liver disease

0 (0.0%)

0 (0.0%)

0 (0.0%)

**NA

Cerebrovascular disease

2(11.8%)

1(11.8%)

1(11.1%)

**0.7352

Alcohol intake

1 (5.8%)

1 (12.5%)

2 (22.2%)

**0.5471

Smoking

2(11.8%)

2 (25.0%)

2 (22.2%)

**0.6647

Cataract/IOL/Clear lens

11 /4 / 2

6/2/0

5/2/2

**0.3607

Glaucoma

0 (0.0%)

0 (0.0%)

0 (0.0%)

**NA

Diabetic retinopathy

0 (0.0%)

0 (0.0%)

0 (0.0%)

**NA

Number of Patients
Age (years)
Gender (Female/ Male)

Systemic Diseases

Ocular Diseases

IOL= Intraocular lens. NA=Not applicable. *Unpaired Students t-test. **Chi-square test of independence or Fishers exact probability test.

Glycolysis in Age-Related Macular Degeneration

The Open Ophthalmology Journal, 2014, Volume 8

allowed to eat or drink on the morning of the test day.

Subjects were also not allowed to take any medications or
supplements before urine collection. Urine samples were
collected in disposable paper cups, after voiding the first
urine of the day and a special urine collection strip (Dip 'N
Dry; US BioTek, Seattle, WA, USA) was dipped into each
urine sample. The strips were packed and sealed in an
aluminum foil bag containing desiccant gel, stored under dry
conditions at room temperature, and then shipped to US
BioTek Laboratories. According to US BioTek Protocol, 4
organic acids in the urine samples (benzoate, lactate, hydroxybutyrate, and -hydroxybutyrate) were measured by
GC/MS. Creatinine was also measured and urinary acid
levels were normalized by the creatinine level. Persons
involved in these analyses were blinded to patient data.
Statistical Analysis
The unpaired Students t-test was used to compare mean
values between the two groups and the chi-square test was
employed to compare percentages. Correlation coefficients were
calculated by Pearsons correlation analysis. Factors associated
with the lactate/pyruvate ratio were investigated by multivariate
logistic regression analysis, with the explanatory variables
including various patient characteristics. Variables were
selected in a stepwise fashion according to their significance for
discrimination between groups. The level of significance was
set at p < 0.05 for all analyses. Statistical analysis was
performed with SAS System 9.1 software (SAS Institute Inc.,
Cary, North Carolina, USA).
Table 2.

41

RESULTS
The mean urinary pyruvate concentration and the mean
percent increase of pyruvate (calculated relative to the mean +
standard deviation) were 8.0 2.8 (g/mg creatinine) and
83.6 51.1 (%), respectively. In 11 patients (64.7%), the
urinary pyruvate level was above the reference value (Table 2,
Fig. 1). The mean urinary concentration and mean percent
increase of lactate were 7.5 8.3 and 426.5 527.8%,
respectively, and 12 patients (70.6%) showed an increase of
lactate above the reference value (Table 2, Fig. 2).
The mean urinary concentration and percent increase of
-hydroxybutyrate were 0.6 0.4 and -31.9 15.2%,
respectively (Table 2, Fig. 3). Finally, the mean urinary
concentration and percent increase of -hydroxybutyrate
were 0.3 0.3 and -33.1 17.5%, respectively (Table 2,
Fig. 4). Both -hydroxybutyrate and -hydroxybutyrate were
within their reference ranges.
The mean lactate/pyruvate ratio was 0.9 0.9, and
12 patients (70.6%) had a ratio above the reference range
(Table 2, Fig. 5). There were no significant differences of
these glycolysis metabolites and parameters between the
tAMD and PCV groups (Table 2).
Table 3 shows the correlations between the
various urinary glycolysis metabolites, while Fig. (6) shows
the correlations of the urinary levels of lactate,
-hydroxybutyrate, and -hydroxybutyrate with pyruvate.
There was no significant correlation between pyruvate and

Comparison of urinary metabolites between tAMD and PCV groups.

*Reference Range Female/Male (g/mg Cr)

Total (N=17)

tAMD (N=8)

PCV (N=9)

P Value tAMD vs PCV

0.6-6.6/0.5-5.9

8.0 2.8

8.3 3.1

7.7 2.4

**0.3502

Pyruvate
Concentration (g/mg Cr)
% Increase

83.6 51.1

93.4 59.0

74.9 41.0

**0.2500

Abnormal level

11/17 (64.7%)

5/8 (62.5%)

6/9 (66.6%)

0.6267

Lactate
Concentration (g/mg Cr)

0.0-1.6/0.0-1.6

7.5 8.3

4.6 5.4

10.1 9.6

**0.0960

% Increase

426.5 527.8

244.2 343.4

588.5 604.7

**0.0960

Abnormal level

12/17 (70.6%)

5/8 (62.5%)

7/9 (77.7%)

0.4367

0.2-2.8/0.2-2.8

0.6 0.4

0.5 0.4

0.7 0.4

**0.1233

% Increase

-31.9 15.2

-36.7 15.6

-27.6 13.5

**0.1233

Abnormal level

0/17 (0.0%)

0/8 (0.0%)

0/9 (0.0%)

NA

0.0-1.9/0.0-1.9

0.3 0.3

0.3 0.3

0.4 0.3

**0.3406

% Increase

-33.1 17.5

-35.1 17.6

-31.3 17.2

**0.3406

Abnormal level

0/17 (0.0%)

0/8 (0.0%)

0/9 (0.0%)

NA

-Hydroxybutyrate
Concentration (g/mg Cr)

-Hydroxybutyrate
Concentration (g/mg Cr)

Lactate/Pyruvate ratio
Value

0.1-0.2/0.1-0.2

0.9 0.9

0.6 0.7

1.2 1.0

**0.0955

% Increase

315.0 369.9

250.8 173.6

440.7 411.1

**0.0734

Abnormal value

12/17 (70.6%)

5/8 (62.5%)

7/9 (77.7%)

0.4367

Cr = Creatinine. NA=Not applicable. % Increase means the percent increase relative to the mean + standard deviation. *Reference ranges are gender-specific. NA=not applicable.
**Unpaired Students t-test. Chi-square test of independence or Fishers exact probability test.

Figure 1
42

The Open Ophthalmology Journal, 2014, Volume 8

Yokosako et al.

% Increase
200

Pyruvate g/mg creatinine

Pyruvate g/mg creatinine

14
12
10
8
6
4
Reference'Range

150
100
50
0
-50

60

70

80

90

-100
-150

0
60

70

80

Age (Years)

Figure 2

90

-200

Age (Years)

Fig. (1). Urinary pyruvate concentration and percent increase of pyruvate relative to the mean reference value.

% Increase
2000

Lactate g/mg creatinine

Lactate g/mg creatinine

35
30
25
20
15
10
5
Reference'Range

0
60

70

Figure 3

80

Age (Years)

90

1500
1000
500
0
-500

60

70

80

90

-1000
-1500
-2000

Age (Years)

Fig. (2). Urinary lactate concentration and percent increase of lactate relative to the mean reference value.

-Hydroxybutyrate
g/mg creatinine

3
Reference'Range

2.5
2
1.5
1
0.5

-Hydroxybutyrate
g/mg creatinine

% Increase
100
80
60
40
20
0
-20 60

70

80

90

-40
-60
-80

0
60

70

80

Age (Years)

90

-100

Age (Years)

Fig. (3). Urinary -hydroxybutyrate concentration and percent increase of -hydroxybutyrate relative to the mean reference value.

Figure 4
The Open Ophthalmology Journal, 2014, Volume 8

43

% Increase
100

2
1.8
1.6
1.4
1.2
1
0.8
0.6
0.4
0.2
0

Reference'Range

80

-hydroxybutyrate
g/mg creatinine

-hydroxybutyrate
g/mg creatinine

Glycolysis in Age-Related Macular Degeneration

60
40
20
0
-20 60

70

80

90

-40
-60
-80

60
Figure 5

70

80

Age (Years)

90

-100

Age (Years)

Fig. (4). Urinary -hydroxybutyrate concentration and percent increase of -hydroxybutyrate relative to the mean reference value.

% Increase
1250

Pyruvate/ Lactate ratio

Pyruvate/ Lactate ratio

5
4
3
2
1

1000
750
500
250
0
-250 60

70

80

90

-500
-750

Reference'Range -1000

0
60

70

80

Age (Years)

90

-1250

Age (Years)

Fig. (5). The lactate/pyruvate ratio (a marker of mitochondrial impairment) and the percent increase of this ratio relative to the mean
reference value.

any of the other glycolysis metabolites (Table 3). On the

other hand, the urinary lactate level was significantly
correlated with the level of -hydroxybutyrate (r=0.642,
p=0.0027) and also with that of -hydroxybutyrate (r=0.627,
p=0.0035) according to Pearsons correlation analysis
(Table 3).
When the influence of patient characteristics on the
lactate/pyruvate ratio was investigated, the strongest
correlation was found with the type of AMD (r=0.328),
although it was not significant (p=0.0995, Table 4).
Multivariate logistic regression analysis demonstrated that
none of the clinical parameters tested were associated with
the lactate/pyruvate ratio (Table 4).
DISCUSSION
This prospective study showed that urinary pyruvate and
lactate levels were increased in the majority of patients with
AMD, and the urinary lactate/pyruvate ratio was also
elevated. These findings suggest that decreased pyruvate
kinase activity in the process of glycolysis, which may

inhibit ATP production, may be associated with the

pathogenesis of AMD.
Urinary metabolic profiling using GC/MS has the ability
to provide considerable information on the variations of
metabolites in human body fluids [19]. The blood metabolic
profile may not be specific to a particular disease because it
is affected by metabolism in various organ systems [19].
Additionally, measurement of metabolites by GC/MS using
the US BioTek system is only for urine samples, so we
investigated urinary metabolites in the present study.
We found that pyruvate and lactate levels were increased
by 83.6% and 426.5% compared with their mean reference
values in our 17 patients with AMD. The increase of urinary
pyruvate was probably associated with increased glucose
uptake in these patients. On the other hand, urinary
-hydroxybutyrate and -hydroxybutyrate levels were
decreased by 31.9% and 33.1%, respectively. LDH catalyzes
the conversion of lactate to pyruvate with simultaneous
reduction and oxidation of nicotinamide adenine
dinucleotide (NAD+) to form reduced NAD (NADH) [20,

Figure 6
44

The Open Ophthalmology Journal, 2014, Volume 8

Yokosako et al.

Fig. (6). Correlations between the urinary level of pyruvate and other glycolysis metabolites.

21]. In the presence of oxygen, cells convert glucose to

pyruvate and then generate ATP from its oxidation after
pyruvate is transported into the mitochondria and converted
to acetyl coenzyme A (acetyl CoA). Subsequently, pyruvate
is metabolized via the tricarboxylic acid (TCA) cycle.
Mitochondrial NADH and flavine adenine dinucleotide
(FADH2) provide energy for ATP production through the
process of oxidative phosphorylation via the electron
transport chain [22, 23]. Under anaerobic conditions,
however, pyruvate is converted to lactate by LDH [22, 23],
and it is not utilized in the TCA cycle after conversion. Our
finding that the percent increase of lactate (426.5%) was 5fold greater than that the percent increase of pyruvate
(83.6%) suggests that glycolysis was shifted toward the

production of lactate. Additionally, the lactate/pyruvate ratio

was increase by 315.0% compared with the mean reference
value. These results suggest that the transition from
glycolytic to mitochondrial metabolism is impaired in
patients with AMD. The lower urinary levels of hydroxybutyrate and -hydroxybutyrate also support the
interpretation that the TCA cycle was inhibited by
impairment of the last energy-generating step in the process
of glycolysis.
The pyruvate level in the urine showed no correlation
with the urinary levels of lactate, -hydroxybutyrate, or hydroxybutyrate. On the other hand, significant positive
correlations were found among these three metabolites,
because all of them are downstream from pyruvate in the

Glycolysis in Age-Related Macular Degeneration

Table 3.

The Open Ophthalmology Journal, 2014, Volume 8

45

Correlations between glycolysis metabolites.

Pyruvate
r

Lactate
P Value

Pyruvate

-HB

-HB

P Value

P Value

P Value

0.300

0.1208

-0.025

0.5380

0.312

0.1118

0.642

0.0027

0.627

0.0035

0.7175

0.0006

Lactate
-HB
Correlation coefficients were calculated by Pearsons product moment formula.
HB = hydroxybutyrate, r = correlation coefficient.

Table 4.

Correlations between the lactate/pyruvate ratio and patient characteristics with multivariate odds ratios and 95%
confidence intervals.

Variable

Correlation Coefficient

Multivariate Analysis

P Value

OR

(95% CI)

P Value

Age (years)

0.099

0.3532

0.995

(-0.26 0.22)

0.9613

Hypertension

-0.192

0.7704

0.427

(-2.57 0.68)

0.2790

Diabetes

-0.03

0.5408

0.597

(-2.46 1.22)

0.5512

Hyperlipidemia

-0.013

0.5204

0.814

(-3.86 3.06)

0.8977

Cerebrovascular disease

0.150

0.2831

2.465

(-2.02 3.51)

0.4885

Alcohol

-0.291

0.8711

0.558

(-2.88 1.47)

0.5669

Smoking

-0.062

0.5928

0.743

(-2.84 1.97)

0.7905

LogMAR visual acuity

0.087

0.3705

0.741

(-2.59 1.75)

0.7662

AMD type (tAMD=0/PCV=1)

0.328

0.0995

2.320

(-0.86 2.36)

0.2794

R = Two-tailed Pearsons correlation coefficients were calculated to assess associations between the Lactate/Pyruvate ratio and patient characteristics (N=17). OR = odds ratio. CI =
confidence interval.

glycolytic process. However, urinary levels of hydroxybutyrate (0.6 g/mg creatinine) and hydroxybutyrate (0.3 g/mg creatinine) were less than one
tenth of the lactate level (7.5 g/mg creatinine), being
decreased by 31.9% and 33.1% relative to the mean
reference value, respectively. Although statistically
significant, the correlations among these metabolites are
probably of little clinical importance.
The mean urinary lactate level and the lactate/pyruvate
ratio were respectively increased by 244.2% and 250.8% in
the tAMD group and by 588.5% and 440.7% in the PCV
group. These findings indicate that the amount of pyruvate
entering the TCA cycle from the glycolytic pathway was
insufficient in both types of AMD, suggesting that tAMD
and PCV may share a common mechanism which involves
activation of LDH and a shift from aerobic to anaerobic
conditions in the mitochondrial-glycolytic interaction. When
the lactate level and the lactate/pyruvate ratio were compared
between the tAMD and PCV groups, both were higher in the
PCV group than the tAMD group (0.6 vs 1.2 g/mg and
250.8% vs 440.7% increase, respectively), although the
differences were not significant. Multivariate analysis did
not extract any of the variables as independent predictors of
the lactate/pyruvate ratio, but showed a weak positive
association with the type of AMD (OR=2.320), and PCV
was more closely associated with the lactate/pyruvate ratio
than tAMD. The reason why the lactate level was higher in
the PCV group than in the tAMD group is unclear but may

be partly due to minor differences of choroidal

microvascular changes between tAMD and PCV.
Additionally, the PCV group may have had more advanced
AMD because visual acuity was worse in this group than in
the tAMD group, although there was no significant
difference between the two groups. Choroidal microvascular
abnormalities associated with PCV are reported to include
arteriosclerotic vascular wall changes with hyalinization,
which differ from the changes of CNV associated with
tAMD [24-27]. PCV also shows histopathologic similarities
with branch retinal vein occlusion, which features hyalinelike degeneration of the vessel walls [25]. Microvascular
endothelial cells sustain their growth and proliferation via
aerobic glycolysis, which involves conversion of pyruvate to
lactate [28]. Glycolysis may contribute to age-related
alterations of the choroidal microvasculature and to the
pathogenesis of PCV. However, we cannot comment about
the influence of the glycolytic pathway on vascular changes
in AMD because no direct evidence of a relationship has yet
been obtained in vivo using animal models or in vitro using
cultured choroidal endothelial cells.
It is still unknown how glycolysis contributes to the
pathogenesis of AMD, which is a multifactorial condition
that is chiefly associated with exposure to UV light [29] and
aging [30]. There are several possible mechanisms by which
glycolysis could promote the progression of AMD [31].
Normal cells obtain ATP via the oxygen-dependent pathway
of oxidative phosphorylation and also via the oxygen-

46

The Open Ophthalmology Journal, 2014, Volume 8

independent pathway of glycolysis. Under hypoxic

conditions, pyruvate is metabolized to lactic acid instead of
entering the TCA cycle, but this process has less capacity to
produce ATP from glucose. The primary factor mediating
this hypoxic metabolic switch is hypoxia-inducible factor
(HIF), an oxygen-sensitive transcriptional activator [32-34].
The retina requires a continuous supply of oxygen and
glucose to maintain normal function and viability. Hypoxic
metabolism has been reported to occur in the RPE [31, 35]
and the retina employs both anaerobic and aerobic glycolysis
[36-40]. Approximately 80% of the glucose consumed by the
outer retina is utilized for aerobic glycolysis [40], while the
RPE has a high rate of lactate production concomitant with
high oxygen consumption [41-43]. Elevation of the blood
lactate level is associated with visual dysfunction in diabetic
patients without retinopathy [44]. Taken together, these
reports provide considerable support for our hypothesis that
inhibition of glycolysis at the final step, which is the shift
from glycolysis (pyruvate to lactate) to mitochondrial
oxidative metabolism, may contribute to the pathogenesis of
AMD.
The main limitation of this study was that there was no
control group without AMD. To overcome this problem, we
employed reference values obtained in healthy subjects.
Second limitation is that we could not measure the
intraocular levels of pyruvate and lactate. Further
investigations using animal models of AMD might solve this
problem. Third limitation is that our patients took drugs for
hypertension (n=7), diabetes (n=3), hyperlipidemia (n=1),
and cerebrovascular disease (n=2). Additionally, 11 patients
suffered from cataract and 3 patients had diabetes. Although
urinary glycolysis metabolites had no correlation with these
risk factors, the urinary levels of glycolysis may be liable to
bias from these variables.
CONCLUSION
In summary, we performed the first investigation of
glycolysis metabolites in patients with AMD. We found that
urinary pyruvate and lactate levels were increased in AMD
patients. In addition, the lactate/pyruvate ratio may be linked
to the prognosis of AMD, which would potentially make it a
useful clinical parameter. Abnormalities of glycolysis seem
to be involved in the pathogenesis of AMD.

Yokosako et al.

REFERENCES
[1]
[2]
[3]
[4]

[5]
[6]
[7]
[8]
[9]

[10]
[11]
[12]
[13]
[14]
[15]
[16]

[17]
[18]

TRIAL REGISTRATION
This trial was approved by the regional ethics committee
(H24-2611) and is registered with the UMIN Clinical Trials
Registry, number UMIN000013684.

[19]
[20]

CONFLICT OF INTEREST
The authors report no conflicts of interest. The authors
alone are responsible for the content and for writing this
paper.
ACKNOWLEDGEMENTS
This work was supported in part by a Grant-in-Aid for
Scientific Research from the Ministry of Education, Culture,
Sports, Science and Technology of Japan and Health Labour
Sciences Research Grant from The Ministry of Health
Labour and Welfare of Japan.

[21]
[22]
[23]
[24]

[25]

Kreisberg RA. Lactate homeostasis and lactic acidosis. Ann Intern

Med 1980; 92(2 Pt 1): 227-37.
Cohen RD, Woods HF. Lactic acidosis revisited. Diabetes 1983;
32(2): 181-91.
Bakker J, Coffernils M, Leon M, Gris P, Vincent JL. Blood lactate
levels are superior to oxygen-derived variables in predicting
outcome in human septic shock. Chest 1991; 99(4): 956-62.
Kompanje EJ, Jansen TC, van der Hoven B, Bakker J. The first
demonstration of lactic acid in human blood in shock by Johann
Joseph Scherer (1814-1869) in January 1843. Intensive Care Med
2007; 33(11): 1967-71.
Semenza GL, Roth PH, Fang HM, Wang GL. Transcriptional
regulation of genes encoding glycolytic enzymes by hypoxiainducible factor 1. J Biol Chem 1994; 269(38): 23757-63.
Gatenby RA, Gillies RJ. Why do cancers have high aerobic
glycolysis? Nat Rev Cancer 2004; 4(11): 891-9.
Bi X, Lin Q, Foo TW, et al. Proteomic analysis of colorectal cancer
reveals alterations in metabolic pathways: mechanism of
tumorigenesis. Mol Cell Proteomics 2006; 5(6): 1119-30.
Denkert C, Budczies J, Weichert W, et al. Metabolite profiling of
human colon carcinoma--deregulation of TCA cycle and amino
acid turnover. Mol Cancer 2008; 7: 72.
Chan EC, Koh PK, Mal M, et al. Metabolic profiling of human
colorectal cancer using high-resolution magic angle spinning
nuclear magnetic resonance (HR-MAS NMR) spectroscopy and
gas chromatography mass spectrometry (GC/MS). J Proteome Res
2009; 8(1): 352-61.
Qiu Y, Cai G, Su M, et al. Serum metabolite profiling of human
colorectal cancer using GC-TOFMS and UPLC-QTOFMS. J
Proteome Res 2009; 8(10): 4844-50.
Bernal-Mizrachi C, Semenkovich CF. Fast predators or fast food,
the fit still survive. Nat Med 2006; 12(1): 46-7.
Juraschek SP, Shantha GP, Chu AY, et al. Lactate and risk of
incident diabetes in a case-cohort of the atherosclerosis risk in
communities (ARIC) study. PLoS One 2013; 8(1): e55113.
Vazquez A. Metabolic states following accumulation of
intracellular aggregates: implications for neurodegenerative
diseases. PLoS One 2013; 8(5): e63822.
Poitry-Yamate CL, Poitry S, Tsacopoulos M. Lactate released by
Mller glial cells is metabolized by photoreceptors from
mammalian retina. J Neurosci 1995; 15(7 Pt 2): 5179-91.
Winkler BS, Arnold MJ, Brassell MA, Puro DG. Energy
metabolism in human retinal Mller cells. Invest Ophthalmol Vis
Sci 2000; 41(10): 3183-90.
Chung SH, Shen W, Gillies MC. Laser capture microdissectiondirected profiling of glycolytic and mTOR pathways in areas of
selectively ablated Mller cells in the murine retina. Invest
Ophthalmol Vis Sci 2013; 54(10): 6578-85.
Thylefors B, Chylack LT Jr, Konyama K, et al. WHO Cataract
Grading Group. A simplified cataract grading system. Ophthalmic
Epidemiol 2002; 9(2): 83-95.
Salmi H, Kuitunen M, Viljanen M, Lapatto R. Cow's milk allergy
is associated with changes in urinary organic acid concentrations.
Pediatr Allergy Immunol 2010; 21(2 Pt 2): e401-6.
Chan EC, Pasikanti KK, Nicholson JK. Global urinary metabolic
profiling procedures using gas chromatography-mass spectrometry.
Nat Protoc 2011 8; 6(10): 1483-99.
Vassault A. Lactate dehydrogenase. In: Bergmeyer Methods of
Enzymatic Analysis, Vol III, 3rd ed. Deerfield Beach, FL: Verlag
Chemie 1983; p. 118.
Selwood T, Jaffe EK. Dynamic dissociating homo-oligomers and
the control of protein function. Arch Biochem Biophys 2012;
519(2): 131-43.
Lehninger AL. Principles of biochemistry. Worth: New York 1982.
Semenza GL. HIF-1 mediates the Warburg effect in clear cell renal
carcinoma. J Bioenerg Biomembr 2007; 39(3): 231-4.
Terasaki H, Miyake Y, Suzuki T, Nakamura M, Nagasaka T.
Polypoidal choroidal vasculopathy treated with macular
translocation: clinical pathological correlation. Br J Ophthalmol
2002; 86(3): 321-7.
Okubo A, Sameshima M, Uemura A, Kanda S, Ohba N.
Clinicopathological
correlation
of
polypoidal
choroidal
vasculopathy revealed by ultrastructural study. Br J Ophthalmol
2002; 86(10): 1093-8.

Glycolysis in Age-Related Macular Degeneration

[26]

[27]
[28]

[29]
[30]
[31]
[32]
[33]
[34]

The Open Ophthalmology Journal, 2014, Volume 8

Kuroiwa S, Tateiwa H, Hisatomi T, Ishibashi T, Yoshimura N.

Pathological features of surgically excised polypoidal choroidal
vasculopathy membranes. Clin Experiment Ophthalmol 2004;
32(3): 297-302.
Nakashizuka H, Mitsumata M, Okisaka S, et al. Clinicopathologic
findings in polypoidal choroidal vasculopathy. Invest Ophthalmol
Vis Sci 2008; 49(11): 4729-37.
Parra-Bonilla G, Alvarez DF, Al-Mehdi AB, Alexeyev M, Stevens
T. Critical role for lactate dehydrogenase A in aerobic glycolysis
that sustains pulmonary microvascular endothelial cell
proliferation. Am J Physiol Lung Cell Mol Physiol 2010; 299(4):
L513-22.
Chalam KV, Khetpal V, Rusovici R, Balaiya S. A review: role of
ultraviolet radiation in age-related macular degeneration. Eye
Contact Lens 2011; 37(4): 225-32.
Chakravarthy U, Wong TY, Fletcher A, et al. Clinical risk factors
for age-related macular degeneration: a systematic review and
meta-analysis. BMC Ophthalmol 2010; 10: 31.
Chiu CJ, Taylor A. Dietary hyperglycemia, glycemic index and
metabolic retinal diseases. Prog Retin Eye Res 2011; 30(1): 18-53.
Brahimi-Horn MC, Pouyssgur J. The hypoxia-inducible factor and
tumor progression along the angiogenic pathway. Int Rev Cytol
2005; 242: 157-213.
Kim JW, Dang CV. Cancer's molecular sweet tooth and the
Warburg effect. Cancer Res 2006; 66(18): 8927-30.
Ke Q, Costa M. Hypoxia-inducible factor-1 (HIF-1). Mol
Pharmacol 2006; 70(5): 1469-80.

Received: April 11, 2014

[35]
[36]
[37]
[38]
[39]
[40]
[41]

[42]
[43]
[44]

Revised: July 18, 2014

47

Winkler BS, Sauer MW, Starnes CA. Modulation of the Pasteur

effect in retinal cells: implications for understanding compensatory
metabolic mechanisms. Exp Eye Res 2003; 76(6): 715-23.
Cohen LH, Noell WK. Glucose catabolism of rabbit retina before
and after development of visual function. J Neurochem 1960; 5:
253-76.
Krebs HA. The Pasteur effect and the relations between respiration
and fermentation. Essays Biochem 1972; 8: 1-34.
Winkler BS. Glycolytic and oxidative metabolism in relation to
retinal function. J Gen Physiol 1981; 77(6): 667-92.
Ames A 3rd, Li YY, Heher EC, Kimble CR. Energy metabolism of
rabbit retina as related to function: high cost of Na+ transport. J
Neurosci 1992; 12(3): 840-53.
Wang L, Kondo M, Bill A. Glucose metabolism in cat outer retina.
Effects of light and hyperoxia. Invest Ophthalmol Vis Sci 1997;
38(1): 48-55.
Miceli MV, Newsome DA, Schriver GW. Glucose uptake, hexose
monophosphate shunt activity, and oxygen consumption in cultured
human retinal pigment epithelial cells. Invest Ophthalmol Vis Sci
1990; 31(2): 277-83.
Coffe V, Carbajal RC, Salceda R. Glucose metabolism in rat retinal
pigment epithelium. Neurochem Res 2006; 31(1): 103-8.
Kaur C, Foulds WS, Ling EA. Hypoxia-ischemia and retinal
ganglion cell damage. Clin Ophthalmol 2008; 2(4): 879-89.
Mondal LK, Baidya KP, Bhattacharya B, Giri A, Bhaduri G.
Relation between increased anaerobic glycolysis and visual acuity
in long-standing type 2 diabetes mellitus without retinopathy.
Indian J Ophthalmol 2006; 54(1): 43-4.

Accepted: July 21, 2014

Yokosako et al.; Licensee Bentham Open.

This is an open access article licensed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/)
which permits unrestricted, non-commercial use, distribution and reproduction in any medium, provided the work is properly cited.