Deletion of GLUT1 in mouse lens epithelium leads to cataract formation

Abstract

The primary energy substrate of the lens is glucose and uptake of glucose from the aqueous humor is dependent on glucose transporters. GLUT1, the facilitated glucose transporter encoded by Slc2a1 is expressed in the epithelium of bovine, human and rat lenses. In the current study, we examined the expression of GLUT1 in the mouse lens and determined its role in maintaining lens transparency by studying effects of postnatal deletion of Slc2a1. In situ hybridization and immunofluorescence labeling were used to determine the expression and subcellular distribution of GLUT1 in the lens. Slc2a1 was knocked out of the lens epithelium by crossing transgenic mice expressing Cre recombinase under control of the GFAP promoter with Slc2a1 mice to generate Slc2a1;GFAP-Cre (LensΔGlut1) mice. LensΔGlut1 mice developed visible lens opacities by around 3 months of age, which corresponded temporally with the total loss of detectable GLUT1 expression in the lens. Spectral domain optical coherence tomography (SD-OCT) imaging was used to monitor the formation of cataracts over time. SD-OCT imaging revealed that small nuclear cataracts were first apparent in the lenses of LensΔGlut1 mice beginning at about 2.7 months of age. Longitudinal SD-OCT imaging of LensΔGlut1 mice revealed disruption of mature secondary fiber cells after 3 months of age. Histological sections of eyes from LensΔGlut1 mice confirmed the disruption of the secondary fiber cells. The structural changes were most pronounced in fiber cells that had lost their organelles. In contrast, the histology of the lens epithelium in these mice appeared normal. Lactate and ATP were measured in lenses from LensΔGlut1 and control mice at 2 and 3 months of age. At 2 months of age, when GLUT1 was still detectable in the lens epithelium, albeit at low levels, the amount of lactate and ATP were not significantly different from controls. However, in lenses isolated from 3-month-old LensΔGlut1 mice, when GLUT1 was no longer detectable, levels of lactate and ATP were 50% lower than controls. Our findings demonstrate that in vivo, the transparency of mature lens fiber cells was dependent on glycolysis for ATP and the loss of GLUT1 transporters led to cataract formation. In contrast, lens epithelium and cortical fiber cells have mitochondria and could utilize other substrates to support their anabolic and catabolic needs.

Modulating GLUT1 expression in retinal pigment epithelium decreases glucose levels in the retina: impact on photoreceptors and Müller glial cells

“…For example, Fidler et al ( 10) observed a similar metabolic flexibility when they deleted GLUT1 from platelets, which subsequently increased their use of amino acids. In addition, we observed a similar compensation by amino acids when we deleted GLUT1 from mouse lens epithelium and found a decrease in amino acids in the aqueous humor of transgenic mice, indicating an increase in amino acid oxidation by the mouse lens under glucose deprivation (46). Alternatively, glucose derived from the inner retinal vasculature, perhaps along with amino acid oxidation, could be sufficient to maintain the observed degree of photoreceptor viability.…”

“…Sections were incubated at room temperature with secondary antibodies (Table 1) and with DAPI, and then imaged on an LSM 780 NLO laser scanning microscope (Carl Zeiss, Oberkochen, Germany) using ApoPlan ϫ63/1.4 objective and EC NeoPlan ϫ10/0.3 objective. For hematoxylin and eosin (H&E)-stained sections, methanol:acetic acid fixed eyes were embedded in paraffin and 10-m sections were cut as described previously (46). Paraffin sections were deparaffinized using xylene and rehydrated in a graded series of ethanol then water and used for immunofluorescence labeling as described above.…”

Mechanosensitive collaboration between integrins and connexins allows nutrient and antioxidant transport into the lens

“…Another study showed glucose concentrations in anterior aqueous humor of 137 mg/100 ml (7.6 mM), and it was speculated that glucose concentration was ∼100 mg/100 ml (5.5 mM) in extracellular lens fluid ( Kuck, 1965 ). Glucose in the lens cell is primarily from aqueous humor ( Swarup et al, 2018 ), and glucose is converted into glucose-6-phosphate rapidly inside cells ( Kinoshita, 1965 ). Although the gradient of glucose from the cortex to the nucleus has not been directly measured, we would expect that glucose in the microfluidic solution is higher than that inside the lens cells, and like GSH, there is a gradient, with the highest level of glucose found in cortical fibers.…”

Loss of MPC1 reprograms retinal metabolism to impair visual function

“…Mouse experiments were performed in accordance with the National Institutes of Health guidelines and the protocol approved by the Institutional Animal Care and Use Committee of West Virginia University. MPC1 RNA in situ hybridization, metabolite analysis by mass spectrometry, EM, immunohistochemistry, and other methods were performed as reported (38,39) and described in details in SI Appendix.…”