Investigative Ophthalmology & Visual Science, Vol. 33, No. 9, August 1992
Copyright © Association for Research in Vision and Ophthalmology
Plasma Docosahexaenoic Acid Levels in Various
Genetic Forms of Retinitis Pigmenfosa
Junxian Gong,* Bernard Rosner.f Deirdre G. Wees* Elior L. Derson.f
Carol A. Weigel-DiFranco,f and Ernst J. Schaefer*
In 188 patients from separate families with various forms of retinitis pigmentosa (RP) and 91 normal
subjects, plasma fatty acids were measured as a percentage of total plasma fatty acids, and their
concentrations were determined using capillary-column gas-liquid chromatography. After controlling
for the effects of age and gender, those with RP had significantly lower (P < 0.01) mean plasma
percentages and concentrations of the omega-3 fatty acids: 18:3u>3 (alpha-linolenic acid), 22:3u;3
(13,16,19 docosatriaenoic acid), and 22:6a>3 (docosahexaenoic acid, DHA) compared with the group of
normal subjects. The mean percentages were reduced 15%, 14%, and 10%, respectively, below the mean
percentages in normal subjects. Analysis by genetic type revealed that the X-linked and isolate forms of
RP had significantly lower (P < 0.01) mean percentage values for DHA (18% and 17%, respectively).
Dominant and recessive forms of RP had DHA levels close to normal. Mean absolute plasma DHA
concentrations in X-linked RP were not significantly different from the concentrations in the control
subjects, although these levels were significantly lower in patients with isolate RP. These data identify
the possibility that some forms of RP may have alterations in plasma omega-3 fatty acid metabolism
resulting in decreased plasma DHA content. These observations await additional confirmation using an
analysis of the fatty acid content of specific erythrocyte phospholipid classes. Invest Ophthalmol Vis
Sci 33:2596-2602,1992
Several investigators have observed abnormalities
of plasma docosahexaenoic acid (DHA) in various
common forms of retinitis pigmentosa (RP). In one
study,1 the DHA (22:6co3) content of plasma phospholipids was 36% lower (P < 0.05) in affected patients in
three kindreds with Usher's syndrome, an inherited
disorder involving congenital deafness and retinitis
pigmentosa, compared with that in control subjects.
Others2 found decreased plasma DHA content in autosomal dominant RP and isolate (simplex) RP. In
other studies,3"5 affected patients in four of five Xlinked kindreds had a reduced plasma content of
DHA compared with their unaffected relatives.1
These reductions were significantly different (P
From the *Lipid Metabolism Laboratory, Tufts University
School of Medicine, and the |Berman-Gund Laboratory for the
Study of Retinal Degenerations, Harvard Medical School, Massachusetts Eye and Ear Infirmary, Boston, Massachusetts.
Supported by the George Gund Foundation (Cleveland, OH), the
National Retinitis Pigmentosa Foundation, Inc. (Baltimore, MD),
and specialized research center grant P50EY02014 from the National Eye Institute (Bethesda, MD). JG was supported in part by a
fellowship from the United Nations University Fund (Tokyo, Japan).
Submitted for publication: June 1, 1990; accepted February 28,
1992.
Reprint requests: Ernst J. Schaefer, Lipid Metabolism Laboratory, Tufts University School of Medicine, 711 Washington Street,
Boston, MA 02111.
< 0.05) from control levels in two of the four families.1"4 Hyperlipidemia also was seen in some of these
kindreds.1"5 It was suggested that RP is associated
with abnormal plasma lipoproteins6 and an increased
frequency of the rare apoE isoform of apolipoprotein
E2.7 However, apoE is not essential for normal fatty
acid metabolism. In its absence, plasma fatty acid levels are normal, and no RP is found.8'9 In another
study, the DHA (22:6OJ3) content of plasma phospholipids was elevated in 26 affected relatives in a large
family with RP from the same geographic area.10
Fatty acid abnormalities recently were discovered in a
strain of miniature poodles with progressive rod-cone
degeneration; in affected dogs, the plasma DHA levels
were significantly reduced (P < 0.005).'1>12 These studies suggested a genetic defect in the delta-4 desaturase
step that converts 22:5oo3 to 22:6co3 and 22:4co6 to
22:5o;6 in poodles.
Rod outer segment membranes contain exceptionally large amounts of polyunsaturated fatty acids, especially DHA, which comprises almost one half of the
esterified fatty acid in outer segment phospholipids.1314 Outer segment membranes are also unusual because of their high phospholipid content (8090 mol% of total lipid) and low cholesterol content
(8-9 mol% of total lipid). Moreover, the fatty acid
content of the phospholipids is exceptional because of
their high degree of polyunsaturation. These factors
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DOCO5AHEXAENOIC ACID IN RP / Gong er ol
combine to produce membranes of high fluidity. The
proportion of DHA in phosphatidylethanolamine
and phosphatidylserine in the membranes of outer
segments of photoreceptor cells is typically 45-60%
higher than that in any other tissue. It comprises 75100% of the fatty acids in the 2 position of these phospholipids, the position usually occupied by polyunsaturated fatty acids. In contrast, the plasma fatty acid
content of DHA is only 2-3% of the total plasma fatty
acids.15 In addition, 5-25% of retinal phospholipid
molecules also contain DHA in the 1 position, which
is unusual because this position usually is occupied by
a saturated fatty acid.15 The significance of the special
lipid composition of the rod photoreceptor outer segment is not understood, but it is thought to be necessary for the normal functioning of rhodopsin in the
initial stage of phototransduction. Activation by a
photon of light causes a rapid change in the conformation of rhodopsin, which undergoes lateral and rotational movement in the rod outer segment membranes. The high degree of fluidity and flexibility of
disc membranes appears to be essential for this dynamic behavior of rhodopsin.1416"18
Although DHA turnover in the retina must be
rapid, prolonged dietary deficiency of this fatty acid
and its omega-3 precursors is necessary to decrease its
retinal levels. It appears to be retained by the retina
even when animals are fed diets that are deficient in
omega-3 fatty acids.1519-20 The mechanism of conservation is not clear. It may be recycled in the photoreceptors before the tips are shed and phagocytized,
DHA may be recycled directly to the pigment epithelium by the phagosomes, or it may be released by the
phagosomes into the blood stream and recycled
through the liver and plasma.14
Current evidence indicates that alpha linolenic acid
(18:3co3), its major dietary precursor, can be converted to DHA in the liver by elongation and desaturation.21-22 Little is known about the biologic pathways
in DHA (22:6co3) synthesis and transport. In normal
dogs, their retinas have a limited capacity for elongation and desaturation of 18:3co3 and 22:6co3."-14
Through the bloodstream, DHA is distributed from
the liver to other tissues.14 Major food sources are fish
and some plant oils. The vertebrate retina may depend on blood lipids and/or other carriers to supply
essential fatty acids for normal outer-segment phospholipid synthesis. Specific carriers may be involved
in transporting DHA (1) from the liver to the eye and
(2) across the pigment epithelium and interphotoreceptor matrix to supply the photoreceptor cells.18-2123
If DHA is important for normal retinal function in
terms of maintaining outer segment membrane fluidity, then deficiencies of this fatty acid or abnormalities
in its metabolism could lead to alterations in mem-
2597
brane function (and possibly to RP). This possibility
and the reported abnormalities found in plasma DHA
in some patients with RP1"5 prompted us to investigate the plasma levels of additional patients with different genetic types of this condition.
Materials and Methods
Sample Selection
We evaluated the plasma fatty acid levels in 188
patients (age range, 18-49 yr), one per family, with
common forms of RP. The diagnosis in each patient
was based on findings on ocular examinations, including electroretinography done at the BermanGund Laboratory. These patients were classified genetically after their family histories were reviewed as
autosomal dominant, autosomal recessive, X-linked,
or isolate, using previously described criteria.2425
Plasma fatty acid levels also were evaluated in 91 normal control subjects (age range, 18-49 yr), none of
whom had any family history of RP. In patients and
control subjects, blood (20 ml) was drawn after they
had fasted overnight (12-14-hr). The sample was
placed in 0.1% ethylenediaminetetraacetic acid-containing tubes, and the plasma was separated from the
erythrocytes by centrifugation at 2500 rpm and 4°C.
The plasma was stored immediately at -70°C under
nitrogen gas before analysis.
Lipid and Fatty Acid Analysis
All analyses were done in a masked fashion. Only
numbered samples were provided to the Lipid Metabolism Laboratory at Tufts University. A modification
of Lepage's method was used for lipid extraction and
transesterification of the fatty acids.26-27 Plasma (100
/A) and 40 ^g of an internal standard (margaric acid,
17:0) were mixed in 13 100-mm glass tubes. We
added 2 ml of a 4:1 (v/v) methanol-benzene solution
and 200 n\ of acetyl chloride with slow stirring. Nitrogen gas was added to each tube before they were sealed
with Teflon tape, tightly closed with Teflon-lined caps
(Fisher Scientific, Springfield, NJ), and then methanolized at 100°C for 1 hr in a Reacti-Therm (Pierce
Chemical Co., Rockford, IL) heating-stirring dry
block. Then the tubes were cooled in water, and 1 ml
of isoctane was added to dilute the solution. We
added 5 ml of 6% potassium carbonate to stop the
reaction and neutralize the mixture. The tubes were
centrifuged at 1500 X g for 15 min. Subsequently, the
clear upper phase that contained the fatty acid methyl
esters (FAME) was transferred to a 1-ml autosampler
vial. After being overlaid with anhydrous sodium sulfate (1 mm layer), the tubes were filled with nitrogen,
and the vials were sealed with crimp caps. They were
ready for injection into the measuring device. An au-
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INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / August 1992
tomated system was used for analysis; this consisted
of a Hewlett Packard (Avondale, PA) HP 7673A automatic sampler, an HP 5890 gas-liquid chromatograph, an HP 3393A integrator, and a Nelson chromatography data system (Cupertino, CA). The last device collects, reintegrates, and stores the data. The
samples in the autosampler tray were cooled to keep
the sample temperature at approximately 10°C to
prevent FAME oxidation. We used 0.32-mm (internal diameter) 30-M (length) Supelcowax 10 (Supelco,
Bellefonte, PA) capillary columns for these analyses.
A flame ionization detector also was used. The gas
chromatograph was run using hydrogen as the carrier
gas with an approximate head pressure of 9 psi and a
flow rate of 4 ml/min. Other approximate conditions
were as follows: flame, hydrogen 20 psi at 25 ml/min
and air 40 psi at 385 ml/min; make up, nitrogen 27
ml/min; and temperature program, first ramp 150210°C with increases of 2°C/min, then plateau at
210°C for 10 min, third ramp 210-250°C with increases of 10°C/min, and then constant 250°C for 6
min. Both injector and detector were maintained at
270°C. The signal range and attenuation were both
zero. We injected 0.5-AI1 samples into the gas chromatograph in the splitless mode. Select standard
FAME were obtained from Nuchek Prep (Elysian,
MN), Sigma (St. Louis, MO), and Supelco. These
were used for calibration and peak identification.
FAME percentages were calculated by the Nelson
data system based on the fatty acid amount divided by
the total fatty acids.
Statistical Methods
Analysis of covariance was used to compare (1)
plasma fatty acid mole percentages in all patients with
Vol. 33
RP patients versus those in the group of normal subjects and (2) each group of patients by genetic type
versus the group of normal subjects, after controlling
for age and gender. We followed the SAS General Linear Model procedure (Statistical Analysis System Institute, Cary, NC). Fatty acid levels did not follow a normal distribution. To normalize the distribution, the
values were transformed using the natural logarithm
logg (x + 1) transformation in the analyses of covariance.28-29
In thefirstanalysis, a class variable with two categories was used to distinguish affected patients from
normal subjects. In the second analysis, differences
among each of the four genetic types versus those in
normal subjects were assessed using a class variable
withfivecategories. First, an F test was done to assess
overall group differences after controlling for age and
gender. Differences in the mean fatty acid percentages
between pairs of groups were analyzed using the leastsignificant difference approach. Contingency-table
methods were used to compare the percentage of patients with RP of specific genetic types who were below the tenth percentile in their amount of plasma
DHA. Fatty acid concentration data were reported in
moles percent (ie, milligrams per deciliter).
Results
The mean levels of fatty acids as a percentage (normalized mass percent) of total fatty acids in patients
with RP and in normal control subjects are shown in
Table 1. These fatty acid values reflect approximately
90% of all free and bound fatty acids in the plasma,
including those bound to albumin and those in triglycerides, phospholipids, and cholesterol esters. Patients
with RP had significantly lower (P < 0.01) percentages of alpha linolenic acid (18:3co3); 13,16,19 doco-
Table 1. Plasma fatty acid normalized mass percentage in retinitis pigmentosa patients and normal controls
Fatty acid
(n = 188)
x±SE
Normal
(n = 91)
x±SE
P*
16:0 (palmitic)
18:0(stearic)
18:l«9(oleic)
18:2w6(linoleic)
18:3w3 (alpha linolenic)
18:3w6 (gamma linolenic—GLA)
20:3co6 (dihomogamma linolenic—DGLA)
20:4w6 (arachidonic)
20:5w3 (5,8,11,14,17 eicosapentaenoic—EPA)
22:3o>3 (13,16,19 docosatrienoic)
22:5u>3 (7,10,13,16,19 docosapentaenoic)
22:6o>3 (4,7,10,11,16,19 docosahexaenoic—DHA)
20:5co3/22:6a>3t
21.81 ±0.15
10.19 ±0.23
18.17 ±0.22
27.53 ±0.35
0.93 ± 0.05
0.30 ±0.01
1.48 ±0.03
6.58 ±0.10
0.44 ± 0.03
0.22 ± 0.02
0.32 ± 0.02
1.10 ±0.05
0.46 ± 0.03
21.48 ±0.22
9.93 ± 0.27
17.24 ±0.26
28.15 ±0.44
1.27 ±0.09
0.32 ± 0.03
1.41 ±0.04
6.35 ±0.13
0.49 ± 0.04
0.47 ± 0.05
0.36 ± 0.03
1.34 ±0.06
0.40 ± 0.03
NS
NS
NS
NS
<0.001
NS
NS
NS
NS
<0.001
NS
0.005
NS
Affected
* P values for differences between affected and normal subjects after controlling for age and sex using the Statistical Analysis System General Linear
Model procedure.
t Molar ratio that allows for quantification of the relative amounts of poly-
unsaturated fats in the plasma, (n = 185 for 20:5OJ3/22:6CO3 for affected patients.)
NS, not significant.
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DOCOSAHEXAENOIC ACID IN RP / Gong er ol
No. 9
satrienoic acid (22:3oo3); and DHA (22:6a>3). These
omega-3 fatty acids were, respectively, 15%, 14%, and
10% lower than the mean normal values after controlling for age and gender. All patients and control subjects had plasma cholesterol values over 100 mg/dl,
excluding the diagnoses of abetalipoproteinemia and
homozygous hypobetalipoproteinemia.30'31 Phytanic
acid was not seen in any sample, excluding the diagnosis of Refsum's disease.
The mean levels of plasma fatty acids as percentages of total fatty acids by genetic type of RP (after
controlling for age and gender) are shown in Table 2.
For 52 patients with autosomal dominant and 48 patients with autosomal recessive RP, no significant differences from normal were found with respect to
mean DHA percentages. The patients with dominant
RP had significantly lower mean percentages of alpha
linolenic acid (18:3w3, P < 0.05) and 13,16,19 docosatrienoic acid (22:3u>3, P < 0.01). These fatty acids
were decreased 13% and 15%, respectively, below the
mean normal levels. The patients with recessive RP
had a significantly lower mean percentage of plasma
13,16,19 docosatrienoic acid (22:3<o3, P < 0.01).
Their mean percentage of plasma arachidonic acid
(2O:4co6) was also slightly higher than normal (P
<0.05).
By contrast, 45 patients with X-linked RP (Table 2)
had mean percentages of 13,16,19 docosatrienoic
acid (22:3a>3) and DHA (22:6w3) that were significantly lower than normal (P < 0.01 and P < 0.05,
respectively). These omega-3 fatty acids were 17% and
18% below the mean normal percentages, respectively. These patients also had a 12% higher mean
2599
percentage of oleic acid (18:lco9, P < 0.01) than the
control group (Table 2). Furthermore, the ratio of
5,8,11,14 eicosapentaenoic acid (EPA) to DHA
(20:5co3/22:6o;3) was 0.59 in the X-linked patients
versus 0.40 in the control subjects {P < 0.05). In this
group of patients with X-linked RP, the percentage of
those with DHA values below the tenth percentile of
normal was 13.3% (P not significant).
Forty-three patients with isolate RP (Table 2) also
had significantly lower (P < 0.01) mean percentages
of DHA (22:6co3). Their mean percentage was 17%
below the mean normal level. These patients also had
significantly lower (P < 0.01) mean percentages of
linolenic acid (18:3a>3), EPA (20:5co3), and 13,16,19
docosatrienoic acid (22:3co3). Their mean percentages
were reduced 27%, 10%, and 19%, respectively, below
the mean normal levels. In this group of patients with
isolate RP, the percentage of those with DHA values
below the tenth percentile of normal was 32.6% (P
< 0.05).
Data on each plasma fatty acid's absolute concentration, corrected for lack of complete recovery with
an internal standard, in patients with RP and control
subjects are shown in Table 3. In this analysis, patients with RP had significantly lower (P <, 0.013)
concentrations of plasma alpha linolenic acid, docosatrienoic acid, and docosahexaenoic acid than control subjects. Data on each plasma fatty acid's absolute concentrations in patients with specific genetic
subtypes of RP and control subjects are listed in Table
4. In the autosomal dominant type of RP, alpha linolenic acid concentrations were significantly lower
than normal (P < 0.05). In recessive RP, no differ-
Table 2. Plasma fatty acid normalized mass percentage by genetic type of retinitis pigmentosa
Fatty acid
Dominant
(n = 52)
x±SE
16:0 (palmitic)
21.45
18:0(stearic)
9.79
18:1 w9 (oleic)
17.84
18:2a>6 (linoleic)
27.67
18:3a>3 (alpha linolenic)
0.94
18:3o>6 (gamma linolenic—GLA)
0.28
20:3o)6 (dihomogamma linolenic—DGLA) 1.46
20Aw6 (arachidonic acid)
6.77
20:5«3 (5,8,11,14,17
eicosapentaenoic—EPA)
0.43
22:3a>3 (13,16,19 docosatrienoic)
0.21
22:5w3 (7,10,13,16 docosapentaenoic)
0.30
22:6«3 (4,7,10,11,16,19
docosahexaenoic—DHA)
1.27
20:5w3/22:6o>3§
0.35
Recessive
(n = 48)
x±SE
X-linked
(n = 45)
x±SE
Isolate
(n = 43)
x±SE
21.79 ±0.28 22.08 ± 0.27
9.59 ± 0.37
9.12 ±0.41
0.46 17.47 ±0.36 19.85 ±0.42f
0.72 28.66 ± 0.70 26.42 ± 0.64
0.081: 1.00 ±0.09
1.14 ± 0.10
0.29 ± 0.03
0.03
0.31 ±0.02
0.06
1.47 ±0.06
1.48 ±0.05
0.20 6.78 ±0.17$ 6.02 ±0.19
21.96 0.32
12.46 :O.58t
17.59 0.41
27.27 0.74
0.60 0.07f
0.30 0.02
1.52 0.06
6.71 0.18
0.33
0.38
Normal
(n = 91)
x±SE
F*
21.48 ±0.22 0.88
9.93 ± 0.27 8.56
17.24 ±0.26 4.15
28.15 ±0.44 0.80
1.27 ±0.08 7.26
0.32 ± 0.03 0.57
1.41 ±0.04 0.84
6.35 ±0.13 3.04
NS
<0.001
0.003
NS
<0.001
NS
NS
0.018
0.07
0.03f
0.02
0.47 ± 0.03
0.28 ± 0.04f
0.35 ± 0.03
0.52 ± 0.08
0.25 ± 0.04t
0.34 ± 0.02
0.35 :0.04t
0.14 •0.02t
0.30 0.05|
0.49 ±0.04
0.47 ±0.05
0.36 ±0.03
2.53
7.80
1.39
0.041
<0.001
NS
0.12
0.03
1.13 ±0.06
0.44 ± 0.03
0.98 ±0.09*
0.59 ± 0.06|
1.01
0.49
1.34 ±0.06
0.40 ± 0.03
3.57
2.93
0.007
0.021
* F statistic for the Type III sum of squares comparing overall differences
among genetic types after controlling for age and sex using the SAS GLM
procedure.
t P < 0.01 for different from normal.
t P < 0.05 for different from normal.
o.iot
0.12
§ Molar ratio that allows for quantification of the relative amounts of polyunsaturated fats in the plasma, (n = 51 for dominants and n = 41 for isolates
for 20:5<o3/22:6<o3).
NS, not significant.
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INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / August 1992
Vol. 33
Table 3. Plasma fatty acid concentrations in retinitis pigmentosa and normal controls
Affected
(n = 755;
Normal
(n = 91)
x±SE
x±SE
P*
81.87 ±2.04
37.66 ± 1.04
68.91 ±2.03
100.26 ± 1.66
3.52 ± 0.20
1.13 ±0.05
76.38 ±2.91
34.97 ± 1.37
62.16 ± 2 . 9 5
NS
NS
NS
NS
0.010
NS
NS
0.032
NS
<0.001
NS
0.013
NS
Fatly acid
16:0 (palmitic)
18:0 (stearic)
18:1OJ9 (oleic)
18:2co6 (Hnoleic)
18:3co3 (alpha linolenic)
18:3u)6 (gamma linolenic—GLA)
20:3co6 (dihomogamma linolenic—DGLA)
20:4o>6 (arachidonic)
20:5o>3 (5,8,11,14,17 eicosapentaenoic—EPA)
22:3co3 (13,16,19 docosatrienoic)
22:5w3 (7,10,13,16,19 docosapentaenoic)
22:6co3 (4,7,10,11,16,19 docosahexaenoic—DHA)
20:5u>3/22:6co3t
5.44
23.98
1.65
0.80
1.19
4.01
0.46
97.20 ± 2.24
4.50 ±0.33
1.11 ±0.09
4.95 ±0.19
21.69 ±0.50
1.71 ±0.13
±0.14
± 0.44
±0.12
± 0.06
±0.06
±0.17
± 0.03
1.56 ± 0 . 1 6
1.21 ±0.08
4.55 ± 0.22
0.40 ± 0.03
* P values for differences between affected and normal subjects after controlling for age and sex using the SAS GLM procedure.
t Molar ratio that allows for quantification of the relative amounts of poly-
unsaturated fats in the plasma, (n = 185 for 20:5OJ3/22:6W3 for affected patients.)
NS, not significant.
ences from control levels for any variables were found
except there was an increase in arachidonic acid (P
<: 0.01). With respect to X-linked RP, palmitic (P
^ 0.01) and oleic (P < 0.01) acid values were significantly higher than control levels. Docosatrienoic values were significantly lower (P < 0.01). By contrast
with the percentage values, the actual concentrations
of DHA in X-linked RP were not significantly different from those in control subjects. In isolate RP,
plasma stearic acid values were significantly higher (P
< 0.01). Alpha linolenic acid, EPA, docosatrienoic
acid, docosapentaenoic acid, and DHA were all significantly lower than normal (P < 0.01).
Discussion
Our study showed that mean plasma DHA levels
(as percentages of total plasma fatty acids in plasma)
were reduced below the mean normal levels in patients with X-linked and isolate forms of RP. Although the reductions were statistically significant,
the actual levels were reduced only 17-18% below
normal. Because DHA normally comprises approximately 3% of the total plasma fatty acids, it is unknown whether these observed differences in the Xlinked and isolate forms of RP have any clinical significance. Most patients with isolate RP are thought to
Table 4. Plasma fatty acid concentration by genetic type of retinitis pigmentosa
Fatty acid
16:0 (palmitic)
18:0 (stearic)
18:1 w9 (oleic)
18:2co6 (linoleic)
18:3o>3 (alpha linolenic)
18:3OJ6 (gamma linolenic—GLA)
20:3OJ6 (dihomogamma linolenic—
DGLA)
20:4OJ6 (arachidonic acid)
20:5co3 (5,8,11,14,17
eicosapentaenoic—EPA)
22:3u>3 (13,16,19 docosatrienoic)
22:5o)3 (7,10,13,16 docosapentaenoic)
22:6co3 (4,7,10,11,16,19
docosahexaenoic—DHA)
20:5o>3/22:6o;3§
Dominant
(n = 52)
Recessive
(n = 48)
X-linked
(n = 45)
Isolate
(n = 43)
x±SE
x±SE
x±SE
x±SE
79.56
35.91
67.33
99.44
3.47
1.08
± 3.27
82.11 ± 3 . 3 3
35.86 ± 1.85
± 1.77
± 3.69
66.15 ±2.94
±2.71
105.22 ± 3.26
± 0.36*
3.81 ± 0 . 3 5
1.14 ± 0.12
±0.11
5.38 ± 0 . 2 9
24.53 ± 0.94
1.57 ±0.26
0.74 ±0.1 If
1.12 ± 0 . 0 8
4.48 ± 0.39
0.35 ± 0.03
90.31
36.08
82.07
103.53
4.71
1.25
±6.15f
± 1.96
± 5.96f
±4.28
±0.53
±0.10
75.57
43.44
60.11
92.30
±2.64
± 2.62f
±2.01
±2.71$
2.02 ±0.2If
1.04 ±0.08
Normal
(n = 91)
x±SE
F*
P
97.20 ± 2.24
4.50 ± 0.33
1.11 ± 0 . 0 9
2.37
2.25
4.33
3.44
7.42
0.76
0.053
NS
0.002
0.009
<0.00l
NS
4.95 ± 0 . 1 9
21.69 ± 0 . 5 0
1.80
2.02
NS
5.08
7.99
2.43
<0.001
<0.001
0.048
4.07
2.96
0.003
0.020
76.38 ±2.91
34.97 ± 1.37
62.16 ±2.95
5.38 ±0.27
25.32 ± 0.98f
5.83 ± 0.29f
23.19 ±0.84
1.74 ± 0 . 1 3
1.08 ± 0 . 1 5
1.27 ±0.11
2.11 ±0.34
0.88 ± 0.13f
1.36 ±0.11
0.48 ± 0.08t
1.02 ±0.16t
1.71 ± 0 . 1 3
1.56 ± 0.16
1.21 ±0.08
4.19 ±0.23
3.82 ±0.37
0.59 ± 0.06t
3.43 ± O.35f
0.48 ±0.11
4.55 ± 0.22
0.40 ± 0.03
0.44 ± 0.03
* F statistic for the Type HI sum of squares comparing overall differences
among genetic types after controlling for age and sex using the SAS GLM
procedure.
f P < 0.01 for different from normal.
% P < 0.05 for different from normal.
5.19 ± 0 . 2 3
22.63 ± 0.60
1.17±0.14t
NS
§ Molar ratio that allows for quantification of the relative amounts of polyunsaturated fats in the plasma, (n = 51 for dominants and n = 41 for isolates
for 20:5«3/22:6«3).
NS, not significant.
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No. 9
DOCOSAHEXAENOIC ACID IN RP / Gong er ol
inherit their condition by an autosomal recessive
mode, and it is not clear why the patients with autosomal recessive RP do not show this deficiency. The
finding of normal levels of DHA in our group of patients with dominant RP contrasts with previous reports of reduced levels in one group2 and elevated
levels in another group,10 both with dominant RP.
There is no evidence in humans that photoreceptor
outer segment lipids are altered significantly in the
those with modest changes in plasma fatty acid levels.
Studies in rhesus monkeys have documented that a
low dietary intake of omega-3 fatty acids during prenatal and postnatal development leads to reductions of
80-90% in the DHA content of plasma and tissues
and compensatory increases in long-chain omega-6
fatty acids.32"34 Such increases were not observed in
the plasma in our patients with RP. The visual acuity
of omega-3 fatty acid-deficient infant monkeys was
subnormal.32 These data suggest that DHA depletion
leads to defects in the structure and functioning of
central retinal cones. In the common forms of human
RP, rod function is affected early, and central cone
function is relatively preserved. Similarly, the miniature poodles with rod-cone degeneration and decreases in plasma DHA levels had central degeneration at an early stage; this differed from thefindingsin
common forms of human RP.11
In our studies of patients with X-linked RP, there
was no deficiency in any of the detectable precursors
of DHA, namely 18:3co3, 20:5co3, or 22:5co3, suggesting that some patients with this form of RP may have
a block in the pathways for converting 22:5a>3 to
22:6w3. These pathways recently were studied extensively.35 The possibility of a block was supported by a
significantly higher ratio of 20:5o>3 to 22:6co3 in this
group than in normal subjects. By contrast, in isolate
RP, all omega-3 fatty acids were decreased, including
the precursor 18:3a>3. This pattern raised the possibility of either a dietary deficiency of 18:3co3 or possibly
enhanced utilization of all of the omega-3 fatty acids
by the body in isolate RP.
It is unlikely that any of the genetic types of RP
studied had a significant abnormality in fat absorption. One of the most sensitive indicators of fat malabsorption and dietary essential fatty acid deficiency is a
decreased plasma percentage of linoleic acid (18:
2co6).26'34-36 In all types of RP, normal percentages of
this major essential fatty acid (approximately 27% of
total fatty acids) were found. Similarly, it is unlikely
that these patients with RP consumed a substantially
different diet with regard to saturated fatty acid intake
than do normal subjects. The major saturated fatty
acid in human plasma, palmitic acid (16:0), which
comprises approximately 22-23% of plasma fatty
acids, was virtually identical in all types of RP compared with that in the group of normal subjects.
2601
It is known that the major essential fatty acids (linoleic and alpha linolenic acid) are converted to derivative fatty acids. An interesting feature of our data was
that, in all categories of patients with RP, there was a
consistent decrease in the percent of plasma fatty
acids as 22:3co3, a minor fatty acid (Table 2). This
fatty acid is produced as a result of a minor fatty acid
elongation pathway, whereby alpha linolenic acid
(18:3o;3) is elongated directly to 11,14,17 eicosatrienoic acid (20:3co3) and then to 13,16,19 docosatrienoic
acid (22:3co3) instead of being converted to EPA and
DHA (the major pathway). Therefore, in some patients with RP, there may be excess conversion of
18:3co3 to these major derivatives (EPA and DHA)
instead of the minor derivative, 22:3a>3, perhaps signaling a channeling of precursors to maximize the
EPA and DHA available to target tissues. With regard
to the major elongation and desaturation product of
18:3co3, namely 20:5o;3, only in isolate types of RP
was there a significant decrease. In all other forms of
RP, the percentage of plasma fatty acids as EPA was
similar to normal. This was also the case for the next
elongation product of 20:5a>3, which is 22:5a>3. This
fatty acid was decreased on a percentage basis only in
isolate RP but not in the other forms of RP.
Our results differ from an earlier study2 as they relate to dominant RP, even though both studies were
done on outpatient populations with the common
forms of RP and no known clinical differences. Moreover, we found no definitive evidence that these patients had a clinically significant deficiency of fatty
acids. Patients with essential fatty acid deficiency secondary to intestinal disease with no known retinal degeneration have had percentages of plasma linoleic
acid that were approximately one third of those observed in normal subjects.26 These differences were
much greater than those we observed comparing patients with RP and normal subjects.
In addition, we examined plasma fatty acids and
not fatty acids in specific lipid classes. Lecithin (or
phosphatidylcholine) is the major plasma phospholipid; phosphatidylethanolamine and phosphatidylserine are the major cell membrane phospholipids.
Erythrocyte membrane phospholipid fatty acids are
similar in composition to those of brain and retina,
and they change in parallel, at least in rats, after dietary manipulation.17 Studies are ongoing to determine if the small plasma fatty acid percentage changes
we observed in X-linked RP can be correlated with
alterations in fatty acids in erythrocyte membrane
phospholipids in these same patients.
Key words: retinitis pigmentosa, docosahexaenoic acid
(22:6co3; 4,7,10,13,16,19-docosahexaenoic acid), fatty
acids, retinal degenerations, plasma
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2602
INVESTIGATIVE OPHTHALMOLOGY G VISUAL SCIENCE / August 1992
Acknowledgments
The authors thank Patty Wedge, Frank Weaver, and Kevin McDermott for their assistance in preparing the manuscript.
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