糖尿病视网膜病变,动物模型 z-bo 2020-12-06 378

　　Introduction: Diabetic retinopathy occurs in approximately one-third of diabetic patients. This is the main cause of blindness in adults between 24 and 70 years old. The overall prevalence of type 1 diabetes patients seems to be higher than that of type 2 diabetes patients. Approximately 25% of patients with type 1 diabetes begin to develop symptoms of DR within 5 years after the onset of diabetes and increase to 80% within 15 years. Interestingly, although gender has not been found to have an effect on the susceptibility to type 1 or 2 races, the socio-economic status does affect the susceptibility to DR. The onset and progression of diabetes is caused by many factors, including prolonged course of diabetes, poor blood sugar control, and elevated blood pressure. Hyperglycemia can lead to microvascular disease, including microvascular tumors, bleeding, and thickening of the basement membrane. The result is increased blood retinal barrier permeability (BRB) causing leakage and diabetic macular edema (DME). Increased vascular permeability leads to capillary occlusion, leading to retinal ischemia, leading to increased levels of vascular endothelial growth factor (VEGF). Retinal ischemia and increased vegf levels promote neovascularization. Based on the typical form of neovascularization, DR can be classified as non-proliferative DR (NPDR) or hyperplasia (PDR). Due to microvascular injury and pericyte loss of NPDR, it can progress to proliferative DR, with microvascular tumors, spots and spotting, cotton wool spots, capillary non-perfusion areas. Changes in microglia and DME can also occur in NPDR. In PDR, new blood vessels cause hemorrhage of the retina and vitreous, and cause retinal detachment. DME may also occur in PDR. Disease models contribute to the understanding of the mechanisms leading to DR diseases. Several animal models have been developed to study the etiology and pathogenesis of DR, and to develop and test therapies to treat this disease. Since DR is a complex disease with both genetic and environmental impact, animal models are also developed through induction or gene mutation. The induced model is produced through surgery, drugs, diet, and laser or chemical damage. Use selective breeding and gene editing to create genetic models. Although a large number of species are used to generate DR models, including mice, rats, cats, dogs, pigs and non-human primates, mice and rats have been studied the most because of their small size and short life span , The reproduction speed is fast, and the most effective research can be carried out. Rodents have also been the focus of most genetic research, discovering certain genetic hyperglycemia or obesity. However, the DR phenotype in the canine model appears to be most similar to human DR. Surprisingly, non-human primates have been shown to be relatively resistant to induced DR. Cats generally do not suffer from cataracts. In contrast, the eye structure of pigs and zebrafish is similar to humans, the blood vessel structure is easy to visualize, the life span is short, and the reproduction capacity is large (zebrafish). Although there is no single animal model that represents the complete vascular and neurological complications of human DR in the early and late stages. The model described in this review can help determine the mechanism of DR and develop new therapies.

　　 Induction model: An induction model was established through five methods: pancreatic resection, alloxan administration, streptozotocin (STZ), high galactose diet, laser or chemical damage. Although all induction methods are still being studied, the most common is the use of streptozotocin, because it leads to the fastest rate of disease development. Alloxan is considered to be less efficient in diabetes induction, and dietary methods take the most time for disease progression. Models caused by surgery and injuries are technically the most challenging. The most commonly used models for inducing DR are mice and rats, but whole, cats, pigs, rabbits, monkeys, and zebrafish are also used. In larger animals, the pathology of inducible DR is usually slower, with rodents and zebrafish more favored.

　　 Pancreatectomy: One of the oldest methods of inducing diabetes in animal models is pancreatectomy or removal of β cells from the pancreas. Complete resection of the pancreas in adult dogs induces diabetes. This technique is usually applied to large animals such as cats and monkeys. In adult cats, hyperglycemia occurs within 3 weeks after surgery; combined use with alloxan can be shortened to within 1 week after surgery. Thickening of the basal capillary membrane can occur 3 months after the onset of hyperglycemia. Other symptoms of DR, including microaneurysms, non-perfusion areas of capillaries, and neovascularization usually take 5-9 years to form. Therefore, the maintenance of this model takes a long time. BRB leakage in the monkey model is within 1 year of the onset of hyperglycemia. However, after 10 years, the monkeys still did not develop proliferative retinopathy. Although the pancreas is removed and long-term diabetes, monkeys seem to be able to resist the induction of DR very well. Only 30% of diabetic patients develop DR, which suggests that primates may have additional biological mechanisms to re-regulate homeostasis in chronic insults. Understanding the regulatory factors that contribute to the physiological differences of species is important for the development of appropriate disease models.

　　 Alloxan: Alloxan is a derivative of uric acid that directly acts on β cells in the pancreas. Injection of alloxan in the tail vein causes hypoglycemia due to necrosis of the islets in the pancreas. Beta cell death causes the release of insulin stored in these cells, causing hypoglycemia, followed by diabetes within 24 hours. Dunn and McLetchie also created a diabetic rat model induced by intraperitoneal injection of alloxan. Diabetic rabbits showed fatigue and weight loss. Rats fed alloxan showed polydipsia, polyuria, diabetes, and hyperglycemia. . Alloxan-directed cell death is mediated by inhibition of glucokinase, an enzyme in the glucose-insulin regulatory pathway, expressed in the liver and pancreas. The drug can cause toxicity to liver and kidney cells, but the appropriate dose can avoid toxicity. Alloxan has a specific effect on β cells in the pancreas, and has no toxic effect on α, δ or pancreatic exocrine cells. This compound is also unstable at room temperature and room temperature. Alloxan has been used to induce a variety of animals, including mice, rats, dogs and pigs, and rabbits. All models have damage to pancreatic β cells. Mice aged 8 to 10 weeks can be given a single dose of alloxan to induce hyperglycemia and cause diabetes. It was previously believed that alloxan-induced diabetic mice did not develop cell or vascular disease, but a recent study found that alloxan induced the loss of retinal ganglion cells (RGC) within 7 days. Alloxan also induces changes in microglia. Three-month-old animals have thicker cell bodies and shorter dendrites. Rats developed hyperglycemia and diabetes within one week of using alloxan. Neovascularization occurred 2-9 months after induction, and cataract occurred within one year. 15 months later, the thickening of the capillary basement membrane was observed. Alloxan in puppies induces diabetes for 5 consecutive weeks once a week. This causes retinopathy to be very similar to human DR, but it takes up to 53 to 69 months for dogs to develop DR after the onset of alloxan diabetes. Hemorrhage, loss of acellular capillary pericytes and microaneurysms were observed following the onset of alloxan administration. This is a feasible PDR model. This phenotype lasted 11 months. Compared with the dog model, the alloxan-induced pig DR model showed hyperglycemia within 48 hours. Alloxan-induced pigs also developed cataracts and capillary collapse 60 days after using alloxan.

　　 Streptozotocin (STZ): Streptozotocin is an antibiotic used in cancer chemotherapy. Intraperitoneal injection of rat streptozotocin and intravenous injection of canine streptozotocin resulted in persistent hyperglycemia in each species, along with diabetic polydipsia and polyuria. The mechanism of STZ inducing diabetes is due to the destruction of islet cells and the loss of β cells. Beta cells take up STZ because they express low-affinity glucose transporter 2 (GLUT2), and the structure of streptozotocin is similar to glucose and N-acetylglucosamine. Other cells, including hepatocytes and renal tubular cells, also express GLUT2 and experience damage similar to STZ. The mechanism of action of streptozotocin is cell death caused by DNA fragmentation. Streptozotocin-induced DR has been observed in various animal models such as mice, rabbits, pigs, rats, dogs, zebrafish and monkeys. The effect of STZ is generally better than that of alloxan. Several STZ-induced diabetes programs have been developed in mice, providing a total of 150 to 400 mg/kg of streptozotocin in 5 doses. Hyperglycemia usually occurs within 2 weeks and can last up to 22 months regardless of the dosage. The DR phenotype of STZ-induced mice showed increased astrocytes after 4-5 weeks of hyperglycemia. RGC loss occurred at 6 weeks, thinning of the inner nuclear layer (INL) and outer nuclear layer (ONL) of the retina at 10 weeks, new blood vessels formed at 16 weeks, and acellular capillaries appeared within 6 months. Compared with mice, rats require low doses of streptozotocin to develop diabetes. Although rodents are commonly used for streptozotocin-induced diabetes, several other models with different outcomes and incidence rates have been studied. Adult zebrafish, 4-6 months of age, one or several weeks of multiple doses of STZ intraperitoneal injection or direct injection into the caudal fin, hyperglycemia appeared within 3 weeks, 4 weeks showed thinning of the inner plexiform layer (IPL), Photosensitive segment layer (PSL) thins, cone receptor dysfunction, neuron damage. The model is maintained for approximately 80 days after diabetes induction. Larger animal models, such as rabbits, dogs, and non-human primates, are induced with a single dose of streptozotocin. A single dose of STZ induced hyperglycemia in rabbits weighing 1.5 kg. Retina and preretinal hemorrhage, vascular disease, venous thrombosis and proliferative retinopathy were found after 135 days. Dogs from 4.5 to 17 months of age, weighing 11 kg and 24 kg, were given a single dose of alloxan/STZ and developed hyperglycemia within 2 days. Alloxan/STZ-induced diabetic dogs have basement membrane thickening in monkeys at 1 year, and loss of shadow cells and smooth muscle cells after 4–5 years. A 12-year-old monkey will develop diabetes with a single dose of streptozotocin, and develop cotton wool spots and intense fluorescent spot ischemic retinopathy after 10 years.

　　 High-sugar diet: Several high-sugar diet models have been developed, including mice, rats, rabbits, dogs, and zebrafish. Continuous exposure to galactose-induced retinopathy is similar to human diabetes. Maintaining galactose causes the disease to continue to progress. However, galactose-fed animals lack the metabolic abnormalities of diabetes. The mice developed hyperglycemia at 6 weeks of age after the high-galactose diet. 15 months after the operation, blood sugar increased, endothelial cells decreased, and acellular capillaries increased. After 21 months, the lesions included hemangioma and thickening of the retina. Although retinopathy develops longer in these mice, they live longer than other models, allowing them to be observed for a longer period of time, up to 26 months. Similarly, the rats maintained a high-galactose diet for more than 2 years. Continuous high-sugar diet observed the phenotype of rodents with acellular capillaries and capillary basement membrane thickening. In contrast, larger animals usually take longer to develop DR, whether through drug induction or diet. Rabbits on a high sucrose diet established strong fluorescence spots at 24 weeks and microaneurysms appeared at the 12th week. Within 1 year, dogs fed with a 30% increase in galactose will produce more complex disease manifestations, including DR and cataracts. The hyperglycemic zebrafish has developed into a model. The zebrafish are placed in fresh water and water with alternating concentrations of 0 and 2% glucose. After 28 days, hyperglycemia and IPL refinement appear. Since this model has only been maintained for 28 days so far, the characteristics of the retina structure, vascular structure visualization and fluorescent expression are similar. The short life span and large reproduction scale shorten the experimental time, making zebrafish a powerful model for studying DR.

　　 Hypoxic injury causes retinopathy: In recent years, retinal neovascularization models and vascular leakage models lacking hyperglycemia have been used for research. It was later discovered that retinal damage was caused by the release of angiogenic factors. Several different injury models of retinal neovascularization using mice, rats, primates and zebrafish. Hyperoxia-exposed mouse models, usually 7–12 days after birth, once they return to normal air and the growth of retinal blood vessels under hyperoxia conditions will cause hypoxia in the retina. Oxygen-induced retinopathy (OIR) in these models exhibits neovascularization and non-perfusion areas, accompanied by the appearance of microvessels, and usually occurs 5 days after exposure.

　　 Cytokine induction: Alkaline burn model leads to increased cytokine activity and DR-like neovascularization. This model is not commonly used, and a painful method is needed to induce retinal neovascularization in mice. This technique has been applied to inbred mouse strains, such as BALB/c mice. Including the placement of 2 mm filter discs with 1 N NaOH for the ocular surface of adult mice. Neovascularization is usually observed within 2 weeks, and mice also show increased levels of neovascularized retinal inflammatory cytokines. Although cytokines may be a secondary factor in the phenotype of neovascular disease, they may play an important role in maintaining the disease state.

　　 Genetic model: There are many ways of inheriting DR in mice, rats and zebrafish. These models include spontaneous, strain-specific, and genetic mutations. For example, several inbred mice, including non-obese diabetes (NOD) and db/db, exhibit hyperglycemia, which is the main feature of diabetes, and thus maintain and serve as a diabetes model for research. Rodents are often used as genetic models of DR because they are easy to maintain, have a good genetic background, and are easy to manipulate to generate gene knockout or transgenic models.

　　 Conclusion: Animal models play a vital role in understanding the etiology and pathophysiology of DR and developing feasible treatment methods to prevent and alleviate the disease. DR is a complex disease involving a variety of genetic and environmental factors, so it is a challenging disease model. Combining genetic and/or induced models can provide more accurate DR models. Most of the existing models only better simulate the early stages of the disease, which limits the usability of the model, and treatment is usually limited to the early progression of the disease. In addition, an animal model of retinal neovascularization without hyperglycemia has been developed. These models can be used to understand the pathogenesis of advanced DR disease and formulate appropriate treatment plans. A correct understanding of the pathophysiology and limitations of each available model is critical to determine the best model for a study.