1Nuffield Laboratory of Ophthalmology, University of Oxford, Oxford; and 2AstraZeneca R&D Charnwood, Loughborough, United Kingdom
Submitted 20 January 2004 ; accepted in final form 22 September 2004
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ABSTRACT |
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retina; eye; immunohistochemistry; polymerase chain reaction
Monocarboxylates are transported across membranes via a family of proton-coupled carrier proteins known as monocarboxylate transporters (MCTs). Multiple MCT-related sequences have been identified, although only MCT1-MCT4 have been functionally characterized (15). To date, MCT1 is the best-characterized isoform and is found in the majority of tissues analyzed in the rat (15). In contrast, MCT2 has a more limited distribution in the rat and may well exist in several alternatively spliced forms (16). Rat MCT3 has been localized exclusively in the RPE (21), whereas rat MCT4 is particularly evident in tissues with high glycolytic rates, such as skeletal muscle (17). For clarification of the energy utilization requirements of the various ocular tissues, information as to the distribution of the different MCT subtypes in the eye is warranted. Despite the fact that lactate plays a central role in the metabolism of various ocular tissues, clear knowledge of the precise locations of the different MCT isoforms in the eye is lacking. Studies carried out to date have focused exclusively on the RPE (21) and retina (3, 12) and, with respect to the retina, have generated results that do not always correlate.
The distribution of the MCT1-MCT4 isoforms in the retina has thus far been studied using immunohistochemistry and immunoblotting. MCT1 is expressed throughout the retina, being particularly abundant in photoreceptor inner segments, Müller cells, and blood vessels (3, 12, 20). In contrast, data generated on MCT2 are contradictory: Gerhart et al. (12) showed MCT2-like staining to be associated with Müller cells and synaptic and nuclear layers of the retina, yet Bergersen et al. (3) found no evidence for MCT2-like immunoreactivity in the retina. Evidence to date indicates that MCT3 is not expressed in the retina (20), but some immunoreactivity for MCT4 has been reported (3).
In the rat RPE, Philp et al. (21) provided convincing data indicating that high levels of MCT 1 and MCT3 are polarized to the apical and basolateral membranes, respectively, of these cells. The localization of MCT1, as well as the absence of MCT2, was subsequently confirmed by the work of both Gerhart and Bergersen (3, 12). The latter group also concluded that a low level of MCT4-like immunoreactivity is associated with the RPE.
To our knowledge, no reports exist on the types of MCTs that are associated with the various structures of the anterior segment of the eye, yet there are valid reasons to warrant an investigation. Although historically it has been assumed that lactate diffuses out of the lens and cornea, it is more likely that lens epithelial cells and corneal epithelial or endothelial cells actively extrude lactate via one or more of the MCT isoforms. This idea is given significant weight by the finding in rabbit cornea of lactate transport mechanisms that are both carrier mediated and proton coupled (5, 13). With regard to the iris and ciliary epithelia, these tissues are derived from the same embryological structure as the RPE and, as a consequence, share a number of common properties. Indeed, iris pigment epithelium cells in culture can acquire many RPE properties (26). It is therefore of interest to determine whether a pattern of MCT expression exists in the iris and ciliary epithelium that is similar to that in the RPE.
The aim of the present study was to delineate the types of MCTs associated with the various tissues of the rat eye by using more sensitive technologies than previously employed. In particular, the objective was to provide information on the types of MCTs expressed by the different structures of the anterior uvea. To achieve these objectives, we used a combination of conventional and real-time RT-PCR in concert with immunoblotting and immunohistochemistry.
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MATERIALS AND METHODS |
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All experiments conformed to the Association for Research in Vision and Ophthalmology "Statement for the Use of Animals in Ophthalmic and Vision Research." Adult Wistar rats and Royal College of Surgeons (RCS) rats (200250 g) were housed in a temperature- and humidity-controlled room with a 12:12-h light-dark cycle and provided with food and water ad libitum. For bright light-induced injury, Wistar rats were exposed to evenly distributed bright light (2,000 lux) for 48 h and returned to their normal housing for 5 days before being killed. For kainate-induced injury, Wistar rats were injected intravitreally in one eye with 5 nmol of kainate (5 µl in sterile water) and returned to their normal housing for 3 days before being killed.
Culture of Transformed Müller Cells
The RMC-1 transformed Müller cell line was obtained from Dr. V. Sarthy (Dept. of Ophthalmology, Northwestern Univ., Chicago, IL). These cells were grown in MEM supplemented with glucose (final concentration, 25 mM), 2.5 mg/ml amphotericin B, 100 µg/ml gentamicin, and 10% FBS. Cells were passaged at a ratio of 1:3 every 4872 h and used for the outlined PCR studies when confluent, after being scraped and centrifuged to pellets.
PCR Analysis
For RT-PCR studies, retina, RPE, iris, ciliary body, whole cornea, corneal epithelium, corneal endothelium, lens, heart, testis, and skeletal muscle were carefully dissected and homogenized directly in TRI reagent (Sigma). Total RNA was isolated according to the manufacturers instructions, and first-strand cDNA synthesis was performed on 2 µg of DNase-treated RNA as described previously (19).
For conventional RT-PCR, the individual cDNA species were amplified in a reaction containing the cDNA equivalent of 25 ng of total RNA, PCR buffer, MgCl2 (3.5 mM for MCT4, 4 mM for all other primers), dNTPs, the relevant sense and antisense primers, and AmpliTaq Gold (Applied Biosystems, Foster City, CA). Reactions were initiated by incubating at 94°C for 10 min, and PCRs (94°C for 15 s; 52, 53, or 55°C for 30 s; and 72°C for 30 s) were performed for a suitable number of cycles (Table 1), followed by a final extension at 72°C for 3 min. The PCR products of cyclophilin, rhodopsin, and MCT1-MCT4 yielded single bands corresponding to the molecular masses expected on the basis of the relevant sequence (see Table 1). The oligonucleotide primer sequences and their annealing temperatures are also shown in Table 1. PCR reaction products were separated on 1.5% agarose gels with the use of ethidium bromide for visualization.
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Rabbit anti-rat MCT1 (1:1,000) and rabbit anti-rat MCT2 (1:1,000) were both generous gifts from Luc Pellerin (Geneva, Switzerland). For detection of MCT3, two distinct antisera were employed: rabbit anti-rat MCT3 from Chemicon (1:200) and rabbit anti-rat MCT3 from ADI (1:200). Rabbit anti-rat MCT4 (1:200) and mouse anti-chicken actin (1:2,000) were also obtained from Chemicon.
Laser Capture Microdissection
After enucleation, retinas were removed and snap frozen, and 10-µm-thick sections were produced. Slides were fixed in 70% ethanol for 1 min and then dehydrated and cleared in xylene. Cells were isolated from the inner and outer nuclear layers of retinal sections by using a PixCell I laser capture microdissection system (Arcturus Engineering, Harpenden, UK). The cells attached to the capture film were removed from the film with lysis buffer, and total RNA was then extracted as previously described.
Immunohistochemistry
Rats were anesthetized with pentobarbital sodium and transcardially perfused with phosphate-buffered saline (PBS) and, subsequently, with Davidsons fixative. After enucleation, whole eyes were fixed in Davidsons fixative for 24 h, transferred to industrial methylated spirits, and processed for routine paraffin-embedded sections on an automated tissue processor. The eyes were embedded sagitally, and 5-µm serial sections were cut using a rotary microtome. Tissue sections were deparaffinized, rehydrated into 70% ethanol, and treated for 10 min with 0.5% H2O2 in absolute methanol to block endogenous peroxidases before being taken to deionized water. Where necessary, antigen retrieval was achieved by incubating the slides in boiling 10 mM citrate buffer (pH 6.0) at a pressure of 5 lb/in.2 for 5 min. After being washed in PBS-T (PBS containing 0.1% Triton X-100), tissue sections were then equilibrated for in PBS-T containing primary antibody. Sections analyzed for immunohistochemical localization of MCT1 were incubated with primary antisera for 30 min at 37°C, whereas detection of MCT3 and MCT4 was undertaken with overnight antisera incubations at 4°C. MCT2 was analyzed using both methodologies. Furthermore, the antigen retrieval procedure was not employed for detection of MCT3 or MCT4, because this was noted to diminish antisera labeling efficiency. After incubation with primary antisera, sections were washed in PBS-T and incubated in a solution of PBS-T containing appropriate secondary antibodies linked to biotin (1:100; Vector Labs, Peterborough, UK) and horse serum (1:100) for 30 min. After a further wash with PBS-T, sections were developed using a standard avidin-peroxidase detection system (Vector) with 0.1% (vol/vol) H2O2 and 0.1% (wt/vol) 3',3'-diaminobenzidine in PBS-T. Sections were mounted in PBS-glycerol and visualized using light microscopy.
Immunoblotting
Tissue and RPE culture samples were sonicated in homogenization buffer (20 mM Tris·HCl, pH 7.4, containing 2 mM EDTA, 0.5 mM EGTA, 1% SDS, 0.1 mM phenylmethylsulfonyl fluoride, 50 µg/ml aprotinin, 50 µg/ml leupeptin, and 50 µg/ml pepstatin A). The protein contents of the samples were then measured using standard methodology, and samples were approximately equalized. An equal volume of sample buffer (62.5 mM Tris·HCl, pH 7.4, containing 4% SDS, 10% glycerol, 10% -mercaptoethanol, and 0.005% bromphenol blue) was added, and the samples were left at room temperature overnight. Electrophoresis of samples was performed using 10% polyacrylamide gels containing 0.1% SDS. Proteins were then blotted onto nitrocellulose. The blots were incubated with MCT or actin antisera at the appropriate dilutions (see above) for 34 h at room temperature, and secondary antibodies conjugated to horseradish peroxidase were subsequently employed (Sigma). Nitrocellulose blots were developed with a 0.016% (wt/vol) solution of 3-amino-9-ethylcarbazole in 50 mM sodium acetate (pH 5.0) containing 0.05% (vol/vol) Tween 20 and 0.03% (vol/vol) H2O2.
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RESULTS |
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Localization of MCT2 has proved problematic in the past. To verify the specificity of the MCT2 and MCT4 antibodies for use with ocular sections, we performed preliminary staining using testis and skeletal muscle sections. These tissues are known to display discrete patterns of MCT2 (testis) and MCT4 (skeletal muscle) staining (14, 28). The MCT2 antibody stained specifically in sperm tails throughout the epididymis (Fig. 1A) but failed to label skeletal muscle (Fig. 1D), whereas incubation with the MCT4 antibody yielded strong staining in the sarcolemma of extraocular skeletal muscle (Fig. 1C) but no staining in the testis (Fig. 1B).
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To detect the presence of MCT14 mRNAs in rat samples, we first performed RT-PCR by using primers specific to the published sequence for each of the isoforms. To verify the efficiency of each of the primer pairs for amplification of the relevant mRNAs, we included a positive control cDNA sample appropriate for each MCT isoform with the ocular samples (see Fig. 2). For each of the mRNAs analyzed, amplification of samples resulted in a single detectable product whose size was equivalent to that predicted from the relevant sequence (see Table 1). Synthesis of each product was cycle dependent, with saturation of signal achieved by 40 cycles (data not shown), and contamination with genomic DNA was excluded by performing RT-PCR in the absence of reverse transcriptase (negative controls; see Fig. 2). In addition, for each mRNA amplified, nonspecific contamination was excluded by performing RT-PCR with the use of water instead of the cDNA sample (data not shown).
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Retina. Initial RT-PCR experiments showed clear signals for all four MCT mRNAs in normal retinal samples (Fig. 2). Real-time PCR experiments confirmed these results and demonstrated that the level of MCT1 mRNA is substantially higher than the levels of MCT2-4 mRNAs in the retina (Fig. 3). The presence of MCT3 mRNA in retinal samples was surprising, and to exclude the possibility of contamination from RPE, we utilized laser capture microdissection (LCM). Cells isolated from the ganglion cell layer and the inner and outer nuclear layers of the retina were pooled, the RNA was extracted, and real-time PCR was performed. The levels of all four MCT transcripts in the LCM sample were found to be similar to those measured in retinas dissected by conventional means (Fig. 4). Further information regarding the subretinal localization of the mRNAs encoding MCT1-MCT4 was provided by selectively killing certain cell types in the retina and examining whether the amounts of the MCT transcripts were altered. In 60-day-old RCS rats and in normal Wistar rats that had undergone exposure to intense light, there were marked losses of photoreceptors, which were verified by substantial reductions in the mRNA for the photoreceptor-specific marker rhodopsin (Table 3). Comparison of the abundance of the MCT mRNAs in these samples with that in age-matched control retinas revealed that in both groups of photoreceptor-deficient rats, there was a significant decrease in the level of MCT1 mRNA and considerable increases in the levels of MCT2, MCT3, and MCT4 mRNAs (Table 3). Conversely, in the retinas of rats given an intraocular injection of kainate, which kills cells in the inner retina but spares the outer retina, there was a decrease in the levels of MCT2, MCT3, and MCT4 but no change in the level of MCT1 (Table 3). Finally, cDNA samples were prepared from an immortalized rat Müller cell culture. Amplification of these samples resulted in a strong signal for MCT1 mRNA, weak signals for MCT2 and MCT4 mRNAs, and little or no signal for MCT3 mRNA (Fig. 2).
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DISCUSSION |
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The results presented in this study demonstrate that the retina expresses MCT1, MCT2, and MCT4. MCT1 was localized throughout the retina but was particularly abundant in the outer retina, whereas MCT2 and MCT4 were located preferentially in the plexiform layers.
In agreement with the results of previous studies, we found MCT1 immunoreactivity in the rat retina to be strongly associated with photoreceptor inner segments (3, 12). The abundance of MCT1 at this site is illustrated by the substantial reduction of retinal mRNA content in the retinas of photoreceptor-deficient rats. Lactate is thought to be an important energy substrate of photoreceptor inner segments (23), and MCT1 probably facilitates lactate uptake for oxidative phosphorylation. Studies performed with retinal preparations in vitro have identified the Müller cells as the source of the lactate used by photoreceptors (23, 29). Astrocytes play a similar role in the brain, exporting lactate for use by neurons (8), and recent work indicated that the MCT1 and MCT4 isoforms facilitate this release of lactate (10). The MCT isoform responsible for lactate efflux from Müller cells has not, however, been definitively established. The current study sheds some light on the subject of which MCT isoforms are expressed by retinal Müller cells. Müller cells are identified by their processes, which insinuate themselves between neuronal cell bodies in the nuclear layers, ramify laterally in the plexiform layers, and terminate proximally and distally as end-feet in the nerve fiber layer and outer limiting membrane, respectively. In retinal sections, MCT1, MCT2, and MCT4 immunoreactivities were all clearly detectable in the nerve fiber layer and in both plexiform layers, and it may be inferred therefore that all three isoforms are localized to Müller cells. However, the presence of MCT isoforms in the nerve fiber layer may reflect expression by astrocytes rather than by Müller cell end-feet, and many classes of neurons extend processes into the plexiform layers. Thus, to state with any real confidence that a protein is expressed by Müller cells, it is important to examine the nuclear layers and outer limiting membrane, as well. MCT1 labeling was apparent throughout both nuclear layers and at the outer limiting membrane, and MCT4 labeling was present throughout both nuclear layers and at the outer limiting membrane but was less abundant than MCT1, whereas MCT2 labeling was not seen within either nuclear layer or distally. In cultured, transformed Müller cells, MCT1 was the only MCT transcript to be strongly expressed. The combined results suggest that MCT1 is the dominant MCT isoform found in Müller cells, with a limited expression of MCT4.
As mentioned in the Introduction, contradictory results have been reported regarding the localization of MCT2 in the retina: Gerhart et al. (12) showed MCT2-like staining to be associated with Müller cells and synaptic and nuclear layers of the retina, yet Bergersen et al. (3) found no evidence for MCT2-like immunoreactivity in the tissue. Indeed, in a recent paper, Bergersen et al. (4) suggested that the antibody used by Gerhart may recognize MCT4 rather than MCT2 and, by implication, that MCT2 may not be present in the retina. In this study, we used an MCT2 antibody that was previously successful for localization of MCT2 in mouse (10) and rat tissue (2). To verify its suitability for use in rats, we incubated testis and skeletal muscle sections with this MCT2 antibody. Specific staining was observed in sperm tails throughout the epididymis, but no immunolabeling was detectable in the sarcolemma of the extraocular skeletal muscle. Under the conditions employed in our study, therefore, the antibody is specific for MCT2. The results of our study show that MCT2 is present on the processes of neurons in the inner retina rather than in Müller cells.
With regard to MCT4, the combined PCR and immunohistochemistry results indicate that this isoform is likewise concentrated in the inner retina, specifically in the two plexiform layers. The lack of MCT4 in the photoreceptors is in agreement with the data of Bergersen et al. (3), whereas the presence of this isoform in the inner retina concurs with the unpublished data of Philp et al. in the mouse (see Ref. 20). We suggest that MCT4 is expressed by Müller cells, albeit at lower levels than MCT1, but that it is also localized to neuronal processes within the inner retina.
The finding in this study that the retina expresses significant amounts of MCT3 mRNA was surprising because previous work has shown this isoform to be absent from the neural retinas of rats (21) and mice (20, 22). Initially, we considered that the positive signals might have resulted from amplification of mRNA emanating from a few RPE cells that had inadvertently contaminated the retinal samples during the dissection process. However, through the use of LCM, we were able to eliminate this possibility. In previous studies, detection of MCT3 mRNA in the retina was not attempted using PCR but with the very much less sensitive technique of Northern blot analysis. Because the levels of MCT3 transcript are substantially less in the retina than in the RPE, they may well be below the level detectable by Northern blot analysis. Nevertheless, although we detected MCT3 mRNA in the retina, immunohistochemistry experiments failed to identify the corresponding protein. These results are in agreement with those of Philp and colleagues (2022) and point to one of two explanations for the discrepancy: either the MCT3 antibodies used are not sensitive enough to detect low levels of the protein, or the MCT3 mRNA is, for some unknown reason, not translated into immunodetectable protein.
Retinal Pigment Epithelium
Previous work has shown that two MCT isoforms are expressed in significant levels by the adult rat RPE: MCT1 is polarized to the apical membrane, whereas MCT3 is restricted to the basal membrane (3, 21). The coordinated activity of these two transporters is suggested to allow the rapid transport of lactate from the subretinal space across the RPE and into the blood. The results of the current study are in complete agreement with these reported findings. The present study also provides tentative evidence that the MCT2 subtype is expressed at low levels in this cell type. The reason for the inability of previous investigators (3, 12) to detect MCT2 in the RPE is unclear but, as previously mentioned, is probably related to the particular antibodies used in these studies.
Iris and Ciliary Body
The present study shows that the iris expresses the mRNAs encoding all four MCT isoforms. The levels of the MCT1 and MCT2 transcripts were higher than those of MCT3 and MCT4, and immunohistochemistry was only able to show staining for MCT1 and MCT2 in iris tissue. The most prominent immunolabeling for both isoforms was seen in the posterior epithelium. Although little is known about transport processes across the iris, experiments have indicated that up to 45% of the total lactate content of the aqueous humor is lost to the blood vessels of the surrounding iris (24). Historically, this process has been considered to occur via passive diffusion, but in the light of these results, roles for MCT1 and MCT2 in the transport of lactate across the tissue and into the blood vessels the must be considered.
The results obtained with the ciliary body were broadly similar to those found in the iris. Ciliary body samples expressed mRNAs encoding all four MCTs, yet immunolocalization experiments were only able to show staining for MCT1 and MCT2. As in the iris, MCT1 and MCT2 immunolabeling was most intense in the epithelium of the tissue. The ciliary processes consist of a highly vascular zed stromal core and a specialized double layer of epithelium, which is responsible for the secretion of aqueous humor. The inner nonpigmented epithelium is directly adjacent to the posterior chamber, whereas the outer pigmented epithelium rests on the stromal. The patterns of MCT1 and MCT2 staining indicate that it is the nonpigmented cells that express these transporters. Formation of aqueous humor is a highly active process, and the ciliary epithelium meets its considerable energy demands partly through aerobic glycolysis. In fact, the tissue produces lactate in vitro at a rate similar to that in the cornea (25), and the majority of the lactate is generated by the more active nonpigmented epithelium. Lactate produced by the ciliary epithelium can be lost directly to the blood, unlike the situation in the cornea, but experiments have shown that >50% of the lactate produced by the ciliary body is released into the aqueous humor (7, 24). Transport of lactate between these cell layers and the blood and aqueous humor may well be facilitated by MCT1 and MCT2.
The finding that significant levels of MCT3 mRNA but not MCT3 protein are present in the iris and ciliary body is curious. Embryologically, the pigmented ciliary epithelium and the posterior pigmented iris epithelium are the anterior continuation of the RPE. These cell types share many similarities, and indeed, the iris epithelium offers potential as a candidate for transplantation in cases of RPE degeneration (26). Thus it is not surprising that the cells express a similar complement of mRNAs. Aside from the RPE, the only other tissue that expresses functional MCT3 is the structurally and functionally similar choroid plexus (22). Interestingly, compared with the RPE, choroid plexus expresses a similarly high level of MCT3 transcript yet substantially lower amounts of MCT3 protein. Philp et al (22) suggested that expression of MCT3 is regulated at the posttranscriptional level, and it appears that this is true for the iris and ciliary body.
Smooth Muscle
The present study demonstrates that, in the eye, the MCT4 isoform is not only localized to the sarcolemma of extraocular skeletal muscle but also strongly expressed by the smooth muscle lining of larger blood vessels within the iris, ciliary body, and choroid. Of particular note is the expression by vessels comprising the short posterior ciliary artery system. To our knowledge, this is the first demonstration in any tissue of MCT4 expression by vascular smooth muscle cells. Interestingly, MCT4 was not identified in the dilator or sphincter smooth muscle fibers of the iris or in any of the three types of smooth muscle fibers in the ciliary body. An explanation for the presence of MCT4 in ocular vessels is currently a matter for speculation, but given that such large quantities of lactate are produced by virtually all of the ocular tissues, it would be logical if MCT4 were to facilitate the rapid transfer of lactate from these tissues into the vascular system and thereby its removal from the eye. Future work is needed to determine whether MCT4 is expressed by vascular smooth muscle of nonocular tissues.
Cornea
The corneal epithelium is a highly glycolytic tissue that generates significant quantities of lactate. Generated lactate cannot pass into the ocular tear film because of the presence of numerous tight junctions, and kinetic studies have indicated that proton-coupled facilitated transport mechanisms exist for the passage of lactate from the corneal epithelium to the stroma (5) and from the stroma across the corneal endothelium to the aqueous humor (13). The results presented in this article provide evidence for the presence of both the MCT1 and MCT2 isoforms on corneal epithelium and corneal endothelium. The abundance of the MCT2 transcript in the cornea suggests that this isoform plays a key role in the transport of lactate, and it appears to be the dominant isoform for removal of lactate from the basal cells of the corneal epithelium. However, the remarkably high levels of MCT2 may reflect the fact that this isoform has a much lower substrate capacity than the other MCTs (15) and that higher levels are needed to ensure that lactate does not accumulate intracellularly and result in intracellular acidosis, fluid buildup, and light scatter.
The cornea, like the iris, ciliary body, and retina, expressed significant amounts of MCT3 mRNA. Unlike these latter tissues, however, tentative evidence was found to indicate that the mRNA is translated into immunodetectable protein. Using the Chemicon MCT3 antiserum, we detected some specific labeling in the basolateral membrane of the basal cells of the epithelial layer. However, we observed no similar staining with the ADI MCT3 antiserum, despite the fact that both MCT3 antisera labeled the RPE intensely. Future work is needed to examine in more detail than possible here whether the cornea expresses a physiologically relevant amount of MCT3. The finding is potentially interesting, because the cornea appears to be only the third structure after the RPE and choroid plexus to express MCT3. We had anticipated that the MCT4 isoform might be the predominant MCT isoform in the cornea because of the highly glycolytic nature of the tissue, but scant evidence could be found to support this idea.
Lens
To fulfill its major function of refracting light, the lens must remain transparent. As a consequence, it is both avascular and devoid of major organelles and meets the bulk of its energy demands through anaerobic metabolism of glucose supplied by the aqueous and vitreous humors. The major end product, lactate, has traditionally been thought to diffuse out of the lens, but indirect evidence generated using rabbit and bovine lenses points toward a hydrogen ion-coupled mechanism of lactate extrusion (1, 27). The combined PCR and immunohistochemistry results generated in this study indicate that MCT1 and MCT4 are located in this tissue. PCR analysis provided clear evidence for expression of MCT4 mRNA by the lens and more ambiguous evidence for expression of MCT1 mRNA. The immunohistochemical results show that both MCT1 and MCT4 immunoreactivities are associated with the membrane of lens epithelial cells. The MCT4 isoform has an extremely high capacity for lactate and has a marked preference for lactate over pyruvate as a physiological substrate (11). These features specialize MCT4 for the release of lactate from highly glycolytic cell types such as skeletal muscle (6). The expression of MCT4 by lens epithelial cells thus would be entirely consistent with this function.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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