The retina is among the most metabolically active tissues in the body, requiring a constant supply of blood glucose to sustain function. We assessed the impact of low blood glucose on the vision of C57BL/6J mice rendered hypoglycemic by a null mutation of the glucagon receptor gene, Gcgr. Metabolic stress from moderate hypoglycemia led to late-onset loss of retinal function in Gcgr ؊/؊ mice, loss of visual acuity, and eventual death of retinal cells. Retinal function measured by the electroretinogram b-wave threshold declined >100-fold from age 9 to 13 months, whereas decreases in photoreceptor function measured by the ERG a-wave were delayed by 3 months. At 10 months of age Gcgr ؊/؊ mice began to lose visual acuity and exhibit changes in retinal anatomy, including an increase in cell death that was initially more pronounced in the inner retina. Decreases in retinal function and visual acuity correlated directly with the degree of hypoglycemia. This work demonstrates a metabolic-stress-induced loss of vision in mammals, which has not been described previously. Linkage between low blood glucose and loss of vision in mice may highlight the importance for glycemic control in diabetics and retinal diseases related to metabolic stress as macular degeneration.C57BL/6J mice ͉ cell death ͉ glucagon receptor gene ͉ retinal function ͉ visual acuity C hanges in metabolism can affect vision. Lowering blood glucose (BG) can decrease human visual sensitivity (1-3) as does reducing the partial pressure of inhaled oxygen (4). Natural nighttime decreases in glucose availability (5, 6) parallel a decline in visual sensitivity that can be restored by glucose ingestion (7). In the cat, acute decreases in glucose supply can transiently reduce retinal sensitivity (8) and exacerbate the effects of hypoxia on the retina (9). The effects of metabolite supply on vision are not surprising in view of the high energy consumption by the retina (10, 11). Although the retina's high metabolic activity has been known for Ͼ40 years (12), the consequences of an inadequate supply of metabolites are not completely understood.We report here that a chronic decrease in BG in mice decreases retinal function, leading to a loss of vision and eventual degeneration of the retina. We observed decreases in both electroretinogram (ERG) a-and b-waves, as well as a loss in visual acuity. Retinal cell death, assayed by TUNEL, was increased in Gcgr Ϫ/Ϫ mice, and decreases in cell number were detected. These data indicate that a chronic decrease in BG leads to loss of vision and cell death in mice and highlight the possibility that the human retina may likewise be susceptible to hypoglycemia. ResultsGlucagon Receptor and Changes in BG. Hypoglycemia was induced in C57BL/6J mice by a null mutation of the glucagon receptor gene, Gcgr (13). Among its actions, the glucagon receptor under control of glucagon regulates gluconeogenesis to increase BG levels. Liver and kidney abundantly express Gcgr, and PCR analysis reveals trace levels of receptor mRNA in the retina of wi...

The use of the edible photosynthetic cyanobacterium Arthrospira platensis (spirulina) as a biomanufacturing platform has been limited by a lack of genetic tools. Here we report genetic engineering methods for stable, high-level expression of bioactive proteins in spirulina, including large-scale, indoor cultivation and downstream processing methods. Following targeted integration of exogenous genes into the spirulina chromosome (chr), encoded protein biopharmaceuticals can represent as much as 15% of total biomass, require no purification before oral delivery and are stable without refrigeration and protected during gastric transit when encapsulated within dry spirulina. Oral delivery of a spirulina-expressed antibody targeting campylobacter—a major cause of infant mortality in the developing world—prevents disease in mice, and a phase 1 clinical trial demonstrated safety for human administration. Spirulina provides an advantageous system for the manufacture of orally delivered therapeutic proteins by combining the safety of a food-based production host with the accessible genetic manipulation and high productivity of microbial platforms.

Purpose Age-related macular degeneration (AMD) is a condition that progressively reduces central vision in elderly individuals, resulting in a reduced capacity to perform many daily activities and a diminished quality of life. Recent studies identified clinical treatments that can slow or reverse the progression of exudative (wet) AMD and ongoing research is evaluating earlier interventions. Because early diagnosis is critical for an optimal outcome, the goal of this study is to assess psychophysical orientation discrimination for randomly positioned short line segments as a potential indicator of subtle macular changes in eyes with early AMD. Methods Orientation discrimination was measured in a sample of 74 eyes of patients aged 47 to 82 years old, none of which had intermediate or advanced AMD. Amsler-grid testing was performed as well. A masked examiner graded each eye as level 0, 1, 2, or 3 on a streamlined version of the Age-Related Eye Disease Study (AREDS) scale for AMD, based on the presence and extent of macular drusen or retinal pigment epithelium (RPE) changes. Visual acuity in the 74 eyes ranged from 20/15 to 20/40+1, with no significant differences among the grading levels. Humphrey 10–2 and Nidek MP-1 micro-perimetry were used to assess retinal sensitivity at test locations 1° from the locus of fixation. Results Average orientation-discrimination thresholds increased systematically from 7.4° to 11.3° according to the level of macular changes. In contrast, only 3 of 74 eyes exhibited abnormalities on the Amsler grid and central-field perimetric defects occurred with approximately equal probability at all grading levels. Conclusions In contrast to Amsler grid and central-visual-field testing, psychophysical orientation discrimination has the capability to distinguish between eyes with and without subtle age-related macular changes.

Arthrospira platensis (commonly known as spirulina) is a photosynthetic cyanobacterium1. It is a highly nutritious food that has been consumed for decades in the US, and even longer by indigenous cultures2. Its widespread use as a safe food source and proven scalability have driven frequent attempts to convert it into a biomanufacturing platform. But these were repeatedly frustrated by spirulina’s genetic intractability. We report here efficient and versatile genetic engineering methodology for spirulina that allows stable expression of bioactive protein therapeutics at high levels. We further describe large-scale, indoor cultivation and downstream processing methods appropriate for the manufacturing of biopharmaceuticals in spirulina. The potential of the platform is illustrated by pre-clinical development and human testing of an orally delivered antibody therapeutic against campylobacter, a major cause of infant mortality in the developing world and a growing antibiotic resistance threat3,4. This integrated development and manufacturing platform blends the safety of food-based biotechnology with the ease of genetic manipulation, rapid growth rates and high productivity characteristic of microbial platforms. These features combine for exceptionally low-cost production of biopharmaceuticals to address medical needs that are unfeasible with current biotechnology platforms.

Diabetes in pregnancy is associated with microvascular complications and a higher incidence of preeclampsia. The regulatory signaling pathways involving nitric oxide, cGMP, and cGMP-dependent protein kinase (PKG) have been shown to be down-regulated under diabetic conditions and contribute to the pathogenesis of vascular complications in diabetes. The present study was undertaken to investigate how high glucose concentrations regulate PKG expression in cytotrophoblast cells (CTBs). Human CTBs (Sw. 71) were treated with 45, 135, 225, 495, or 945 mg/dL glucose for 48 h. Some cells were pretreated with a p38 inhibitor (10 μM SB203580) or 10 μM rosiglitazone. After treatment, the cell lysates were subjected to measure the expression of protein kinase G1α (PKG1α), protein kinase G1β (PKG1β), soluble guanylate cyclase 1α (sGC1α), and soluble guanylate cyclase 1 β (sGC1β) by Western blot. Statistical comparisons were performed using analysis of variance with Duncan's post hoc test. The expressions of PKG1α, PKG1β, sGC1α, and sGC1β were significantly down-regulated (p < 0.05) in CTBs treated with >135 mg/dL glucose compared to basal (45 mg/dL). The hyperglycemia-induced down-regulation of cGMP and cGMP-dependent PKG were attenuated by the SB203580 or rosiglitazone pretreatment. Exposure of CTBs to excess glucose down-regulates cGMP and cGMP-dependent PKG, contributing to the development of vascular complications in diabetic mothers during pregnancy. The attenuation of hyperglycemia-induced down-regulation of PKG proteins by SB203580 or rosiglitazone pretreatment further suggests the involvement of stress signaling mechanisms in this process.