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4 Systemic Route for Retinal Drug Delivery: Role of the Blood-Retinal Barrier
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processes, namely, the uptake of these agents into the RVEC or RPE cells via
influx transporters and their subsequent excretion into the blood via efflux transporters. For example, the retina-to-blood efflux transport of several organic anions
has been supposed to involve the concerted actions of organic anion transporter 3
(OAT3, SLC22A8) and multidrug resistance-associated protein 4 (MRP4, ABCC4)
at the inner BRB (Barza et al. 1983; Hosoya et al. 2009). OAT3 and MRP4 share
substrates such as b-lactam antibiotics and the anticancer drug 6-mercaptopurine
(6-MP). In this case, inhibition of drug efflux transporters may lead to an
increased distribution of drug to the retina. Studies carried out by Kompella and
his coworkers have demonstrated that preadmi nistration of probenecid, an organic
anion transporter inhibitor, increases retinal concentration of N-4-benzoylaminophenylsulfonylglycine (BAPSG), a novel anionic aldose reductase inhibitor
(Sunkara et al. 2010). Taken collectively, development of drugs that are well distributed to the retina can be achieved by incorporating structures that are recognized by the blood-to-retina transport systems or are not recognized by the
retina-to-blood efflux systems. The success of retinal drug delivery may thus
depend on several factors: (1) identity of the transporters that are expressed specifically at the BRB, (2) differential localization of the transporters in the two
poles of the plasma membrane of the RVEC and RPE cells, and (3) substrate selectivity of the individual transporters, particularly differences in substrate specificity
between the influx transporters and the efflux transporters. These factors can be
exploited to our advantage to establish efficient strategies for optimal delivery of
clinically relevant therapeutic drugs into the retina (Mannermaa et al. 2006;
Hosoya and Tachikawa 2009).
4.2 Blood-Retinal Barrier Influx Transporters/Receptors
as a Potential Route for Retinal Drug Delivery
The BRB transporters play an essential role in the blood-to-retina transport of essential nutrients such as glucose, amino acids, vitamins, and nucleosides. The role of
transporters in this process has been assessed by the greater blood-to-retina permeability rates of these essential nutrients compared with that of mannitol, a marker of
passive non-carrier-mediated diffusion (Hosoya and Tachikawa 2009). The molecular identity of the transporters at the BRB has been established using a conditionally
immortalized rat retinal capillary endothelial cell line (TR-iBRB cells) as in vitro
model of inner BRB (Hosoya et al. 2001b) and primary cultures and cell lines of
RPE cells as in vitro model of outer BRB. A considerable amount of work on the
transport characteristics of RPE cells has also been carried out using the ARPE-19
cell line. Apical membrane vesicles from RPE cells and isolated RPE/choroid preparations have also been used for the directional transport studies. With the use of
these various approaches, a great deal of information is now available on the identity and characteristics of transporters at the BRB as summarized in Table 4.1.
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Transport system
Expression and localization
Endogenous and
potential drug substrates ReferencesInner BRB Outer BRB
SLC2A1
(GLUT1)
rt (LU, AL) rt (BL, AP) D-Glucose, dehy-
droascorbic acid
Hosoya et
al. (2004);
Minamizono et
al.
(
2006); Takata
et
al. (1992)
SLC5A8
(SMCT1)
rt (BL) Lactate, pyruvate, ketone
bodies, benzoate,
salicylate, 5-aminosalicylate,
3-bromopyruvate
Gopal et al. (2007);
Martin et
al.
(2007);
Thangaraju et al.
(2009)
SLC6A6
(TauT)
rt (in vitro) m, h
(in vitro)
Taurine, g-aminobutyric
acid
Bridges et al. (2001);
El-Sherbeny et al.
(2004); Tomi et al.
(2007b, 2008)
SLC6A8 (CRT) rt (LU, AL) Creatine Nakashima et al.
(2004)
SLC6A14
(ATB
0+
)
h (in vitro) Nitric oxide synthase
inhibitors, valacyclovir, valganciclovir
Hatanaka et
al. (2001,
2004); Nakanishi
et
al. (2001);
Umapathy et al.
(2004)
SLC7A5 (LAT1) rt h (in
vitro) L-Leucine,
L-phenylalanine,
L-DOPA, melphalan,
gabapentin
Goldenberg et
al.
(1979); Tomi et al.
(2005); Yamamoto
et al. (2010)
SLC7A7
(y
+
LAT1)
h (in vitro) L-Arginine, L-lysine,
L-ornithine,
L-leucine
Nakauchi et
al. (2003)
SLC7A8 (LAT2) h (in
vitro) L-Leucine,
L-phenylalanine,
L-alanine,
L-glutamine
Yamamoto et
al.
(2010)
SLC7A11 (xCT) rt (in
vitro) m, h
(in vitro)
L-Cystine, L-glutamate Bridges et al. (2001);
Dun et al. (2006);
Tomi et al. (2002)
SLC16A1
(MCT1)
rt (LU, AL) rt (AP), h
(AP)
Lactate, pyruvate, ketone
bodies, foscarnet,
salicylate, benzoate
Enerson and Drewes
(2003); Gerhart
et al. (1999); Philp
et al. (1998,
2003); Morris and
Felmlee (2008)
SLC16A8
(MCT3)
m (BL), rt
(BL), h
(BL)
Lactate, pyruvate Daniele et al. (2008);
Philp et al. (1998,
2003)
SLC19A1
(RFC1)
rt m (AP),
h (AP,
in vitro)
Folate, N
5
-methyltetrahydrofolate
(MTF), methotrexate
Chancy et al. (2000);
Hosoya et al.
(2008b)
(continued)
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Table 4.1 Expression of transporters/receptors at the blood-retinal barrier
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4 Systemic Route for Retinal Drug Delivery: Role of the Blood-Retinal Barrier
Transport system
Expression and localization
Endogenous and
potential drug substrates ReferencesInner BRB Outer BRB
Slco1a4
(Oatp1a4/
Oatp2)
rt rt (AP) Estradiol
17b-glucuronide,
digoxin
Gao et al. (2002); Ito
et
al. (2002)
Slco1c1
(Oatp14)
rt Estradiol
17b-glucuronide
Tomi and Hosoya
(
2004)
Slco4a1 (Oatp-E) rt Thyroid hormone Ito et al. (2003)
SLC22A3
(OCT3)
m, h (in vitro) Prazocin, clonidine,
cimetidine, verapamil,
imipramine,
desipramine,
quinine, nicotine,
methylenedioxymethamphetamine
Koepsell et
al. (2007);
Rajan et al. (2000)
SLC22A5
(OCTN2)
rt (in
vitro) Acetyl-L-carnitine,
L-carnitine,
cephaloridine,
tetraethylammonium,
pyrilamine, quinidine,
verapamil, valproate
Ganapathy et al.
(2000); Ohashi
et al. (1999);
Tachikawa et
al.
(2010)
SLC22A8
(OAT3)
rt (AL) p-Aminohippuric acid,
benzylpenicillin,
6-mercaptopurine
Hosoya et
al. (2009)
SLC23A2
(SVCT2)
h (AP,
in
vitro)
Ascorbic acid Ganapathy et al.
(2008)
SLC29A2
(ENT2)
rt (in vitro) Purine and pyrimidine
nucleosides,
3¢-azido-3¢deoxythymidine
(zidovudine, AZT),
2¢, 3¢-dideoxycytidine
(zalcitabine, ddC),
2¢,3¢-dideoxyinosine
(ddI), cytarabine,
gemcitabine
Baldwin et al. (2004);
Nagase et al.
(2006); Yao et al.
(2001)
SLC46A1
(PCFT)
m, h (in vitro) Folate, MTF,
methotrexate
Umapathy et al.
(2007)
ABCA3 m Tachikawa et al.
(2008)
ABCA9 m Tachikawa et al.
(2008)
ABCB1
(P-glycoprotein)
rt (LU),
m, b
(in vitro),
h (BL) Cyclosporine A,
daunorubicin,
doxorubicin,
irinotecan, paclitaxel,
quinidine, verapamil,
vinblastine
Hosoya and Tomi
(2005); Kennedy
and Mangini
(2002); Tomi and
Hosoya (2010)
Table 4.1 (continued)
(continued)
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M. Tachikawa et al.
Transport system
Expression and localization
Endogenous and
potential drug substrates ReferencesInner BRB Outer BRB
ABCC1 (MRP1) h (in
vitro) Fluorescein, daunorubi-
cin, doxorubicin,
vinblastine
Aukunuru et
al.
(
2001);
Mannermaa et
al.
(
2009); Tomi and
Hosoya (
2010)
ABCC3 (MRP3) m Ethinyl estradiol
glucuronide,
etoposide
glucuronide,
fexofenadine
Tachikawa et al.
(2008); Tomi and
Hosoya (2010)
ABCC4 (MRP4) m (LU), rt h (in
vitro) p-Aminohippuric acid,
6-mercaptopurine
metabolite,
benzylpenicillin
Hosoya and
Tachikawa (
2009);
Mannermaa et al.
(2009); Tachikawa
et al. (2008);
Tagami et al.
(2009)
ABCC5 (MRP5) h (in vitro) Methotrexate Mannermaa et al.
(2009); Tomi and
Hosoya (2010)
ABCC6 (MRP6) m, rt BQ-123, etoposide Hosoya and
Tachikawa (2009);
Tachikawa et al.
(2008); Tomi and
Hosoya (2010)
ABCG2 (BCRP/
MXR/ABCP)
m (LU), rt
(in vitro)
Mitoxantrone, doxorubi-
cin, pheophorbide a
Asashima et al.
(2006); Tomi and
Hosoya (2010)
SR-BI rt Monkey, h
(in vitro)
a-Tocopherol Duncan et al. (2009);
Tachikawa et al.
(2007);
Tserentsoodol
et al. (2006)
Folate receptor a m (BL) Folate Chancy et al. (2000)
ABC ATP-binding cassette; AP apical membrane; b bovine; BL basolateral membrane; BRB blood-
retinal barrier; h human; LU luminal membrane; m mouse; rt rat; SLC solute carrier
Table
4.1 (continued)
4.2.1 Amino Acid-Mimetic Drugs
L-DOPA [levodopa, (-)-3-(3,4-dihydroxyphenyl)-L-alanine], the amino acid precursor of dopamine, and amino acid mustards are examples of amino acid mimetic
drugs. The Na+-independent amino acid transport system, system L recognizes neutral amino acids as endogenous substrates. At the molecular level, system L is
encoded by L (Leucine-referring)-type amino acid transporter (LAT) 1 (SLC7A5)
and LAT2 (SLC7A8). Retinal capillary endothelial cells exclusively express LAT1
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4 Systemic Route for Retinal Drug Delivery: Role of the Blood-Retinal Barrier
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protein (Tomi et al. 2005). RPE cells express mRNAs for LAT1 (Uchino et al. 2002)
and LAT2 (Nakauchi et al. 2003). LAT1 mRNA expression in ARPE-19 cells is
quantitatively 42.5-fold higher than that of LAT2 mRNA expression (Yamamoto
et al. 2010). Indeed, functional analysis suggests that the contribution of LAT1
to L-leucine uptake by ARPE-19 cells is 70% of the total L-leucine uptake
(Yamamoto et al. 2010). Many patients with Parkinson’s disease have blurred vision
or other visual disturbances, which are reflected in the reduced retinal dopamine
concentration and delayed visual evoked potentials (Bodis-Wollner 1997). L-DOPA
corrects these deficiencies (Bhaskar et
al. 1986). Since LAT1 transports L-DOPA as
a high-affinity substrate with the Michaelis constant of 34.2 mM (Uchino et al.
2002), LAT1 at the BRB provides an important route for the delivery of L-DOPA
into the retina. Melphalan (phenylalanine mustard), an alkylaing agent used in the
treatment of retinoblastoma (Kaneko and Suzuki 2003), and gabapentin (an analog
of g-aminobutyrate), a drug used in the treatment of acquired pendular nystagmus
(Averbuch-Heller et al. 1997), are also transported via LAT1 (Goldenberg et al.
1979). Tomi et al. (2005) and Hosoya et al. (2008a) have investigated the potential
participation of LAT1 in the retinal delivery of various amino acid mustards as alkylating agents using TR-iBRB cells. Since LAT1 is an obligatory amino acid
exchanger, influx of one amino acid substrate into cells is coupled obligatorily to
efflux of some other amino acid substrate. Interestingly, melphalan did not transstimulate the efflux of [3H]phenylalanine and [3H]L-leucine in TR-iBRB cells
(Hosoya et al. 2008a) and ARPE19 cells (Yamamoto et al. 2010), respectively. This
suggests that melphalan may not be a good substrate for LAT1. In support of this
notion, melphalan needs to be injected into the vitreous humor in patients with
retinoblastoma because it is not efficiently transported from the blood to the retina
through the BRB. In contrast, phenylglycine-mustard was very effective in inducing
the efflux of [3H]phenylalanine, suggesting that phenylglycine-mustard is an effective transportable substrate for LAT1. Even though L-DOPA and certain amino
acid-mustards are recognized as transportable substrates for LAT1, LAT1-mediated
delivery of drugs into the retina is likely to exhibit competition from its endogenous
amino acid substrates in vivo. LAT1 possesses high affinity for its endogenous substrates; the Michaelis constants for L-leucine in TR-iBRB cells and ARPE19 cells
are ~15 mM (Tomi et al. 2005) and 8.7 mM (Yamamoto et al. 2010), respectively.
The normal plasma concentration of L-leucine (80–160 mM) is several-fold higher
than this value. Furthermore, the other endogenous amino acids such as L-isoleucine,
L-valine, L-phenylalanine, L-tyrosine, and L-tryptophan, which are transportable
substrates for LAT1, are present in the plasma at the concentration range of
52–220 mM. These amino acids in plasma may saturate LAT1-mediated transport at
the BRB. RPE cells express mRNA for y+LAT1 (SLC7A7) (Nakauchi et al. 2003),
which transports cationic amino acids such as L-arginine and L-lysine in an Na+-
+
independent manner and neutral amino acids in an Na
-dependent manner. Under
physiologic conditions, the transport process mediated by y+LAT1 involves Na+dependent entry of neutral amino acids into cells coupled with exit of cationic amino
acids from the cells. Therefore, there may be a functional coupling between LAT1/
LAT2 and y+LAT1 in the vectorial transfer of amino acid-mimetic drugs across the
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outer BRB, but the localization of these transporters needs to be investigated. RPE
cells also express the Na+- and Cl−-dependent amino acid transporter ATB
(SLC6A14) (Nakanishi et al. 2001). ATB
0,+
transports a wide variety of drugs and
0,+
prodrugs (Ganapathy and Ganapathy 2005), including nitric oxide synthase inhibitors (Hatanaka et al. 2001) and amino acid derivatives of antiviral agents such as
valacyclovir (Hatanaka et al. 2004) and valganciclovir (Umapathy et al. 2004).
4.2.2 Monocarboxylic Drugs
Endogenous monocarboxylates such as lactate, pyruvate, and ketone bodies (b-
hydroxybutyrate and acetoacetate) are transported via two types of transporters, e.g.,
the H+-coupled monocarboxylate transporters (MCTs) belonging to the SLC16 family and the Na+-coupled monocarboxylate transporters (SMCTs) belonging to the
SLC5 family. MCT1 (SLC16A1) is localized in both the luminal and abluminal
membranes of RVEC (Gerhart et al. 1999). RPE cells express MCT1, MCT3
(SLC16A8), and SMCT1 (SLC5A8). MCT1 is expressed exclusively in the apical
membrane of RPE cells whereas MCT3 is expressed exclusively in the basolateral
membrane (Philp et al. 1998, 2003; Daniele et al. 2008; Deora et al. 2005). SMCT1
is expressed only in the basolateral membrane of RPE cells (Martin et al. 2007).
Thus, MCT1 at the inner BRB and SMCT1 at the outer BRB can be exploited to take
up their substrates from the circulating blood into the cells. Several monocarboxylic
drugs have been shown to be substrates for MCT1; this includes foscarnet, salicylate, benzoate, and a prodrug of gabapentin (Enerson and Drewes 2003; Morris and
Felmlee 2008). The monocarboxylic drugs that are recognized by SMCT1 include
benzoate, salicylate, 5-aminosalicylate (Gopal et al. 2007), and 3-bromopyruvate
(Thangaraju et al. 2009). Therefore, MCT1 and SMCT1 at the BRB have potential
for the delivery of monocarboxylic drugs into retina. Nonsteroidal anti-inflammatory drugs (e.g., ibuprofen, ketoprofen), which are also monocarboxylates, are not
recognized by SMCT1 as transportable substrates, but these drugs function as blockers of the transporter (Itagaki et
that disruption of MCT3 gene impairs visual function presumably as a consequence
of reduction in subretinal space pH (Daniele et al. 2008). This suggests that MCT3
at the outer BRB is critically positioned to facilitate transport of lactate out of the
retina and plays a role in pH homeostasis of the retina (Daniele et al. 2008).
al. 2006). A study on MCT3 knockout mice reveals
4.2.3 Nucleoside Analogs
The Na+-independent equilibrative nucleoside transporters (ENTs) belonging to
the SLC29 family accepts purine and pyrimidine nucleosides as endogenous
substrates. ENT1 (SLC29A1) and ENT2 (SLC29A2) transport several antiviral
and anticancer nucleoside drugs such as 2¢, 3¢-dideoxycytidine (zalcitabine, ddC),
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4 Systemic Route for Retinal Drug Delivery: Role of the Blood-Retinal Barrier
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2¢,3¢-dideoxyinosine (ddI), cytarabine, and gemcitabine (Baldwin et al. 2004; Yao
et al. 2001). ENT2 also transports 3¢-azido-3¢-deoxythymidine (zidovudine, AZT)
(Baldwin et al. 2004; Yao et al. 2001). The blood-to-retina transport of [3H]adenosine, a purine nucleoside, is carrier-mediated and is inhibited competitively by unlabeled adenosine and thymidine but not by cytidine (Nagase et al. 2006). Similar
features are evident for adenosine transport in TR-iBRB cells which express ENT2
mRNA (Nagase et al. 2006). Adenosine plays an important role in retinal neurotransmission, blood flow, vascular development, and cellular response to ischemia. The delivery of adenosine into the retina across the inner BRB is therefore
an important physiological process. The Michaelis constant for adenosine for
transport via ENT2 is ~30
in plasma (~0.1 mM), indicating that ENT2 is not saturated with adenosine under
physiologic conditions. Since the nucleoside analogs described earlier are substrates for ENT2 (Baldwin et al. 2004; Yao et al. 2001), this transporter is potentially involved in the delivery of such drugs into the retina. Although the expression
of ENTs has not been examined in RPE cells, ARPE-19 cells exhibit nitrobenzylmercaptopurine riboside-sensitive uptake of nucleoside (Majumdar et al. 2004).
Therefore, it is very likely that the nitrobenzylmercaptopurine-sensitive nucleoside
transporter ENT1 is expressed in these cells. ENTs at the BRB may be a potential
route for the delivery of nucleoside drugs from the circulating blood to the retina.
mM, which is much higher than adenosine concentration
4.2.4 Folate Analogs
The reduced folate carrier (RFC1, SLC19A1), folate receptor a, and the H+coupled folate transporter (PCFT, SLC46A1) play a role in the uptake of folate and
its analogs (Zhao et al. 2009). Tetrahydrofolate functions as a cofactor for de novo
synthesis of purines and pyrimidines, and also as a critical component in the metabolism of the sulfur-containing amino acids methionine and homocysteine. Folate
deficiency causes visual dysfunction; therefore, neural retina must possess mechanisms to obtain this essential vitamin. This implies that the cells constituting the
BRB must express transport systems for folate. Since folate exists predominantly as
the methyl derivative of the reduced folate (N5-methyltetrahydrofolate, MTF) in the
plasma, MTF would be the principal substrate for the folate transport proteins.
RFC1 mediates MTF uptake by TR-iBRB cells, being inhibited by folate analogs
such as methotrexate and formyltetrahydrofolate (Hosoya et al. 2008b). RFC1
mRNA is expressed abundantly in freshly isolated rat RVEC (Hosoya et al. 2008b).
The outer BRB is capable of transcellular transfer of folate in the blood-to-retina
direction, indicating that the folate transport proteins are expressed in RPE cells in
a polarized manner (Bridges et al. 2002). Folate receptor a is expressed in the basolateral membrane whereas RFC1 is expressed exclusively in the apical membrane
(Chancy et al. 2000). Folate receptor a would be involved in the uptake of folate
across the basolateral membrane as the first step in the folate transport across RPE
cells from the circulating blood to the retina. RFC1 in the apical membrane will then
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facilitate the delivery of folate from the cytoplasm into the retina, thus completing
the transcellular transfer. Since the folate receptor a-mediated entry into cells results
in the delivery of folates into endosomes, an additional step is needed to deliver
folates from the endosomes into cytoplasm. PCFT is a candidate responsible for this
process (Qiu et al. 2006). Since there exists an H+ gradient across the endosomal
membrane in the endosome-to-cytoplasm direction, folate will get transferred into
the cytoplasm via PCFT. PCFT is indeed expressed in RPE cells (Umapathy et al.
2007). Electron microscopic analysis shows that folate receptor a and PCFT colo-
calize in Müller cells on the endosomal membrane as well as on the plasma membrane (Bozard et
receptor a in the basolateral membrane of RPE cells and that the two proteins work
together in the delivery of folates from choroidal circulation into RPE cells. Folate
analogs that serve as antifolates (e.g., methotrexate, pemetrexed) can be delivered
into neural retina across the BRB via the concerted actions of the three folate transport proteins. RFC1 transports methotrexate, an antifolate, used in eyes with primary CNS lymphoma, uveitis, and proliferative diabetic retinopathy (Hardwig et al.
2008). However, it would be necessary to consider that the retinal transport of folate
analogs via the folate transport proteins may interfere with the entry of the physiologic substrate MTF into retina.
al. 2010). Thus, it is conceivable that PCFT colocalizes with folate
4.2.5 Organic Cationic Drugs
A variety of organic cation transporters (OCTs and OCTNs) accept endogenous
and exogenous organic cations as substrates (Koepsell et al. 2007). RVEC express
OCTN2 (SLC22A5) (Tachikawa et al. 2010) and RPE cells express OCT3
(SLC22A3) (Rajan et al. 2000). OCTN2 at the inner BRB mediates the bloodto-retina transport of acetyl-L-carnitine (Tachikawa et al. 2010). Acetyl-Lcarnitine is effective in improving visual function in patients with early age-related
macular degeneration. The Michaelis constant for the transport of acetyl-Lcarnitine via OCTN2 in TR-iBRB cells is ~30
ological levels of these compounds in plasma (carnitine, ~50 mM; acetylcarnitine,
~20 mM) (Tachikawa et al. 2010). Exogenous administration of acetyl-Lcarnitine via a systemic route would therefore be able to increase the retinal
levels of acetyl-L-carnitine through OCTN2-mediated transport at the inner
BRB. Several other cationic and zwitterionic drugs including b-lactam antibiotics such as cephaloridine (Ganapathy et al. 2000), tetraethylammonium, pyrilamine, quinidine, verapamil, and valproate (Ohashi et al. 1999) are transportable
substrates for OCTN2. Although the localization of OCT3 in RPE cells remains
unknown, its substrates of pharmacological significance include prazocin
(a-adrenoceptor antagonist), clonidine (a-adrenoceptor agonist), cimetidine
(histamine H1 receptor antagonist), verapamil (calcium channel blocker), imipramine and desipramine (antidepressants), quinine (antimalarial drug), and nicotine and methylenedioxymethamphetamine (an addictive drug) (Koepsell et al.
mM, a value similar to the physi-
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4 Systemic Route for Retinal Drug Delivery: Role of the Blood-Retinal Barrier
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2007). There is also evidence of a novel organic cation transporter in RPE cells
that has not been characterized at the molecular level (Han et al. 2001). This
transport system recognizes verapamil, diphenhydramine, pyrilamine, quinidine,
quinacrine, and brimonidine (Zhang et al. 2006), an a2-adrenergic agonist
approved for the treatment of open-angle glaucoma. Since systemically administrated brimonidine can reach the back of the eye at concentrations sufficient to
activate a2-adrenergic receptors (Acheampong et al. 2002), the novel organic cation transporter in RPE cells regulates the brimonidine concentration in the retina.
Uptake of the endogenous organic cation choline by TR-iBRB cells is Na
+
independent and potential-dependent, indicating that a specific carrier exists at
the inner BRB for the transfer of choline into the retina (Tomi et al. 2007a). The
features of this uptake process are distinct from those of choline uptake mediated
by other known organic cation transporters although the molecular identity of the
transporter remains to be established. Considering that organic cation transporters exhibit broad substrate selectivity, the transport systems responsible for the
transfer of organic cations across the BRB hold great potential for delivery of
various organic cationic drugs into retina.
4.2.6 Opioid Peptides and Peptidomimetic Drugs
It has been proposed that RPE cells possess two novel oligopeptide transport systems
that accept opioid peptides and the peptide fragments of human immunodeficiency
virus HIV-1 Tat (e.g., Tat
2003; Chothe et al. 2010). Although these transporters have not been characterized at
the molecular level, it has been shown that peptides consisting of up to 25 amino acids
interact with these transport systems. The two transport systems are called Na+coupled oligopeptide transporters SOPT1 and SOPT2, which are distinct from the
H+-coupled peptide transporters PEPT1 (SLC15A1) and PEPT2 (SLC15A2).
Although there is a marked overlap between SOPT1 and SOPT2 in substrate specificity, dipeptides and tripeptides stimulate the activity of SOPT1 but inhibit the activity
of SOPT2 (Chothe et
al. 2010; Thakkar et al. 2008). SOPT1 and SOPT2 have poten-
tial for the transport of peptide and peptidomimetic drugs into RPE cells.
) and HIV-1 Rev (e.g., Rev
47–57
) as substrates (Hu et al.
34–50
-
4.2.7 Antioxidants
The retina has an obligate need for antioxidants for protection against light-induced
damage to the cells. Indeed, retinal diseases such as diabetic retinopathy and age-related
macular degeneration have oxidative insults as a pathological component. Vitamin C,
vitamin E, and glutathione are important antioxidants that may have potential in the
treatment of these retinal diseases. Understanding the transport characteristics of these
antioxidants at the BRB may assist in the design and development of suitable therapy
with appropriate antioxidants for treatment of the retinal diseases.
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4.2.7.1 Vitamin C
Vitamin C exists in plasma as the oxidized form (dehydroascorbic acid, DHA) as
well as the reduced form (ascorbic acid). The concentration of ascorbic acid in
plasma is in the range of 50–100 mM whereas DHA is present in the circulation
at much lower levels (~10 mM). However, the influx permeability rate of DHA
across the BRB is ~40-fold greater than that of the reduced form ascorbic acid
(Hosoya et al. 2004). The facilitative glucose transporter GLUT1 (SLC2A1) at
the BRB is responsible for DHA transport in the blood-to-retinal direction.
GLUT1 is expressed on both the luminal and abluminal membranes of the
endothelial cells (Hosoya et al. 2004; Takata et al. 1992). RPE cells also express
GLUT1 which is present both at the apical membrane and basolateral membrane
(Takata et al. 1992). After entering the retina, DHA is reduced into ascorbic acid
for subsequent use in photoreceptors and other retinal cells as an antioxidant.
Since the primary function of GLUT1 at the BRB is to transport glucose from
blood into retina, the fact that GLUT1 is responsible for the transport of both
glucose and DHA to the retina across the inner BRB is very relevant to diabetic
retinopathy. The Michaelis constant for GLUT1 for the transport of glucose is
5–8 mM, which is similar to the physiological plasma concentration of glucose
(~5 mM). This suggests that GLUT1 is not completely saturated with its physiologic substrate in vivo under physiological conditions. However, the blood-toretina transfer of DHA via GLUT1 at the BRB may be impaired significantly in
diabetes because plasma levels of glucose rise markedly in untreated diabetes.
The resultant deficiency of antioxidant machinery may contribute to the pathology of diabetic retinopathy (Minamizono et al. 2006). The reduced form of the
vitamin C, known as ascorbic acid, is transported via the Na+-dependent vitamin
C transporters SVCT1 (SLC23A1) and SVCT2 (SLC23A2). RPE cells express
predominantly SVCT2 (Ganapathy et al. 2008). Functional studies have shown
that the Na+-dependent uptake of ascorbic acid by RPE cells occurs predominantly at the apical membrane (Khatami et al. 1986; DiMattio and Streitman
1991; Lam et al. 1993). Thus, ascorbic acid also enters RPE cells from subretinal
space via SVCT2 at the apical membrane.
4.2.7.2 Vitamin E
Vitamin E has preventive and therapeutic effects in human retinopathies. Among
the members of the vitamin E family, a-tocopherol has the highest biologic activity, and is exclusively associated with high-density lipoprotein (HDL) in the blood
(Goti et al. 2001). Uptake of HDL-associated a-tocopherol into TR-iBRB cells is
most likely mediated by scavenger receptor class B type I (SR-BI) (Tachikawa
et al. 2007). RPE cells express SR-BI and its splice variant SR-BII (Duncan et al.
2009; Tserentsoodol et al. 2006). It is likely that SR-BI at the BRB functions
as an efficient pathway for the supply of a-tocopherol from the blood to retina.
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