Lessons from Genetically Modified Diabetic Animals

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Mice with a genetic ablation of PDGF-B exhibit several vascular phenotypes that are highly reminiscent of certain characteristics of diabetic retinopathy, including microvascular leakage and hemorrhage, pericyte deficiency, and microaneurysms in brain capillaries. A similar phenotype is observed when the PDGF-receptor b is absent, suggesting that this system is crucially involved in the recruitment of pericytes to developing vessels. Moreover, experiments in postnatal mice blocking pericyte recruitment by antibody inhibition of PDGF-receptor b revealed impaired remodeling, distortion and larger diameters of capillaries, and leaki-ness of retinal vessels. Of note, no signs of apoptosis or acellular occluded vessels were present. When administered to older animals, the inhibition of the PDGF-B/receptor b system had no effect on pericyte recruitment, suggesting that the system is only important for recruiting but not for maintaining pericytes in the capillaries. This is to be expected given the redundancy of factors involved in peri-cyte recruitment and attachment, such as angiopoietin-1, TGF-b, and tissue factor. In the absence of pericytes, changes in capillary diameters, with both increased and decreased luminal diameters, are reported. It is an open and largely disputed question whether this is attributable to the propensity of pericytes to contract. However, independent from these open questions, it is concluded that pericytes have a specific and profound role in sprouting angiogenesis in the developing retina, and that pericytes inhibit endothe-lial proliferation, as endothelial cell hyperplasia is seen in mice that lack pericytes.

Note that diabetic retinopathy initiates in the adult, mature retina, which is mostly resistant to manipulations such as hyperoxia or growth factor withdrawal, suggesting that important functions of pericytes and system that affect pericyte recruitment may vary.

Some indication of pericyte function in the adult vessel comes from experiments using retinal digest preparations from mice in which PDGF-B is reduced by half (heterozygous PDGF-B mice). These mice had a 28 percent reduction in pericyte numbers, and a moderate but significant increase in acellular capillaries, suggesting that pericytes function as survival factors for endothelial cells. In diabetic animals, the degree of pericyte loss was increased as expected, but the number of acellular capillaries increased exponentially. This means that with more pericyte loss, PDGF-B is not only a pericyte recruiting factor, but also an indirect survival factor for the stressed endothelium in diabetes.

In endothelial specific PDGF-B knockout mice, a heterogeneous degree of pericyte loss was observed, when assessed in several areas of the brain. In the retina, the numbers of capillary occlusions in the intermediate and deep capillary layers was inversely correlated with the numbers of pericytes on these vessels. The more pronounced the per-icyte deficit, the more capillary occlusions occurred. However, mice with the least pericyte coverage did not develop any capillary layers other than the superficial, and developed proliferative signs in their remaining superficial layer. This indicates that pericytes are important cells for developmental angiogenesis. However, their inhibitory effect on endothelial proliferation needs revision, as pericytes do accompany proliferating vessels in this (pathological) and in other (physiological) settings.

Apart from the PDGF-B/PDGF-bR system, other ligand-receptor systems may be involved in the developmental recruitment of pericytes. Angiopoietin-1, which signals via the endothelial specific tyrosine kinase receptor Tie-2, determines capillary sprouting, endothelial cell survival, and vascular remodeling and has been implicated in the stabilization of vessels by recruiting pericytes. Angiopoietin-2 can act as a natural antagonist of Ang-1. In a rat model of experimental diabetic retinopathy, Ang-2 is upregulated manyfold prior to the onset of pericyte dropout. The upregulation of Ang-2 persists over time, suggesting a nontransient effect of hyperglycemia on Ang-2 transcription. Confirmatory data come from a mouse model in which a reporter construct is expressed under the control of the Ang-2 promoter (Ang-2 LacZ knockin mice). Intravitreal application of Ang-2 mimics the pericyte-depleting effect of hyperglycemia without affecting retinal capillary diameters or endothelial cell survival. When Ang-2 LacZ knockin mice were maintained hyperglycemic for 6 months, they did not develop pericyte loss, as did the wild-type controls, suggesting that Ang-2 plays a significant role in the early peri-cyte loss by hyperglycemia. Preservation of pericytes in this diabetic model was accompanied by a partial reduction of acellular capillaries compared with nontransgenic diabetic mice. Pericytes partially protect endothelial cells in chronic hyperglycemia, but other mechanisms, related to the newly formulated hypothesis of the unifying concept of vascular damage in diabetes by Brownlee (see later discussion), have been proposed.

Confirmation of the role of Ang-2 in pericyte regulation of retinal capillaries also comes from experiments in which Ang-2 was overexpressed in the photoreceptor using a pho-toreceptor-specific promoter. The resulting increase in Ang-2 in the deep layers of the retina induce a selective moderate pericyte loss of the deep capillary network, while leaving the superficial network unaffected. Hyperglycemia in these mice induces a premature damage of the deep capillary layer, as indicated by an untimely formation of acellular capillaries.

Pericytes and acellular capillaries are unevenly distributed within the retina. Whereas pericyte ghosts, which are empty basement membrane pockets suggesting pericyte loss, are similar in all four quadrants of a diabetic dog's retina, acellular capillaries are most frequent in the superior temporal area. A number of questions need to be addressed before these findings are accepted as contradiction against pericytes being survival factors for endothelial cells in the diabetic retina, such as the local production of growth factors, the main subpopulation of pericytes leaving ghosts behind, and the possible contribution of local blood flow in different areas of the retina.

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Diabetes 2

Diabetes 2

Diabetes is a disease that affects the way your body uses food. Normally, your body converts sugars, starches and other foods into a form of sugar called glucose. Your body uses glucose for fuel. The cells receive the glucose through the bloodstream. They then use insulin a hormone made by the pancreas to absorb the glucose, convert it into energy, and either use it or store it for later use. Learn more...

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