The Predictive Role of Thyroid Hormone Levels for Early Diabetic Retinal Changes in Experimental Rat and Human Diabetes.

Purpose: In diabetic subjects, early visual functional alterations such as color vision deficiencies (CVDs) are known to precede clinically apparent diabetic retinopathy. Prominent photoreceptor outer segment degeneration and an increase in the number of retinal dual cones (co-expressing S- and M-opsins simultaneously) have been described in diabetic rat models, suggesting a connection with the development of CVDs. As cone opsin expression is controlled by thyroid hormones, we investigated the diabetic retina in association with thyroid hormone alterations. Methods: In rat models of type 1 and 2 diabetes, dual cones were labeled by immunohistochemistry, and their numbers were analyzed in relation to free triiodothyronine (fT3) and free thyroxine (fT4) levels. Quantification of dual cones was also performed in human postmortem retinas. Additionally, a cross-sectional case-control study was performed where thyroid hormone levels were measured and color vision was assessed with Lanthony desaturated D15 discs. Results: A higher number of dual cones was detectable in diabetic rats, correlating with fT4 levels. Dual cones were also present in postmortem human retinas, with higher numbers in the three diabetic retinas. As expected, age was strongly associated with CVDs in human patients, and the presence of diabetes also increased the risk. However, the current study failed to detect any effect of thyroid hormones on the development of CVDs. Conclusions: Our results point toward the involvement of thyroid homeostasis in the opsin expression changes in diabetic rats and human samples. The evaluation of the possible clinical consequences warrants further research.

Detailed Evaluation of Possible Ganglion Cell Loss in the Retina of Zucker Diabetic Fatty (ZDF) Rats.

A thinning of the inner retina is one of the earliest potential markers of neuroretinal damage in diabetic subjects. The histological background is uncertain; retinal ganglion cell (RGC) loss and changes in the structure or thickness of the inner plexiform layer (IPL) have been suspected. Studies conducted on animal models on RGC pathology gave contradictory results. Hereby we present RGC numbers, distribution patterns and IPL thickness from Zucker Diabetic Fatty (ZDF) rats. After labelling RGCs on retinal whole mounts, isodensity maps were constructed, RGC numbers and distribution patterns analysed using a custom-built algorithm, enabling point-by-point comparison. There was no change in staining characteristics of the antibodies and no significant difference in average RGC densities was found compared to controls. The distribution patterns were also comparable and no significant difference was found in IPL thickness and stratification or in the number of apoptotic cells in the ganglion cell layer (GCL). Our results provide a detailed evaluation of the inner retina and exclude major RGC loss in ZDF rats and suggest that other factors could serve as a potential explanation for inner retinal thinning in clinical studies. Our custom-built method could be adopted for the assessment of other animal or human retinas.

Targeting neuronal and glial cell types with synthetic promoter AAVs in mice, non-human primates and humans.

Targeting genes to specific neuronal or glial cell types is valuable for both understanding and repairing brain circuits. Adeno-associated viruses (AAVs) are frequently used for gene delivery, but targeting expression to specific cell types is an unsolved problem. We created a library of 230 AAVs, each with a different synthetic promoter designed using four independent strategies. We show that a number of these AAVs specifically target expression to neuronal and glial cell types in the mouse and non-human primate retina in vivo and in the human retina in vitro. We demonstrate applications for recording and stimulation, as well as the intersectional and combinatorial labeling of cell types. These resources and approaches allow economic, fast and efficient cell-type targeting in a variety of species, both for fundamental science and for gene therapy.

Histological Evaluation of Diabetic Neurodegeneration in the Retina of Zucker Diabetic Fatty (ZDF) Rats.

In diabetes, retinal dysfunctions exist prior to clinically detectable vasculopathy, however the pathology behind these functional deficits is still not fully established. Previously, our group published a detailed study on the retinal histopathology of type 1 diabetic (T1D) rat model, where specific alterations were detected. Although the majority of human diabetic patients have type 2 diabetes (T2D), similar studies on T2D models are practically absent. To fill this gap, we examined Zucker Diabetic Fatty (ZDF) rats - a model for T2D - by immunohistochemistry at the age of 32 weeks. Glial reactivity was observed in all diabetic specimens, accompanied by an increase in the number of microglia cells. Prominent outer segment degeneration was detectable with changes in cone opsin expression pattern, without a decrease in the number of labelled elements. The immunoreactivity of AII amacrine cells was markedly decreased and changes were detectable in the number and staining of some other amacrine cell subtypes, while most other cells examined did not show any major alterations. Overall, the retinal histology of ZDF rats shows a surprising similarity to T1D rats indicating that despite the different evolution of the disease, the neuroretinal cells affected are the same in both subtypes of diabetes.

Study of retinal neurodegeneration and maculopathy in diabetic Meriones shawi: A particular animal model with human-like macula.

The purpose of this work was to evaluate a potentially useful animal model, Meriones shawi (M.sh)-developing metabolic X syndrome, diabetes and possessing a visual streak similar to human macula-in the study of diabetic retinopathy and diabetic macular edema (DME). Type 2 diabetes (T2D) was induced by high fat diet administration in M.sh. Body weights, blood glucose levels were monitored throughout the study. Diabetic retinal histopathology was evaluated 3 and 7 months after diabetes induction. Retinal thickness was measured, retinal cell types were labeled by immunohistochemistry and the number of stained elements were quantified. Apoptosis was determined with TUNEL assay. T2D induced progressive changes in retinal histology. A significant decrease of retinal thickness and glial reactivity was observed without an increase in apoptosis rate. Photoreceptor outer segment degeneration was evident, with a significant decrease in the number of all cones and M-cone subtype, but-surprisingly-an increase in S-cones. Damage of the pigment epithelium was also confirmed. A decrease in the number and labeling intensity of parvalbumin- and calretinin-positive amacrine cells and a loss of ganglion cells was detected. Other cell types showed no evident alterations. No DME-like condition was noticed even after 7 months. M.sh could be a useful model to study the evolution of diabetic retinal pathology and to identify the role of hypertension and dyslipidemia in the development of the reported alterations. Longer follow up would be needed to evaluate the potential use of the visual streak in modeling human macular diseases.