SURGICAL MODELS:

Surgical models have played a key role in diabetes research when Diabetes occurs in dogs. This procedure was first Discovered by Marcel Eugene Emile Gley in 1891 an early approach to creating diabetic models in research.²⁰Later on, such as pancreatic duct ligation in rabbits by Walpole and Innes in 1946, provided a model for type 2 diabetes. Partial pancreatectomy techniques allowed researchers to study early or prediabetic states. These models use not only for dogs but for other animal also —including rats, mice, cats, rabbits, pigs, and dogs, understanding of diabetes and supporting the development of targeted therapies.^21,22

· Pancreatectomy:

Models of surgery have been key for diabetes research for a century or more. Beginning with complete pancreas removal in dogs in 1891, these techniques have developed to allow partial removal and duct ligation, which has enabled investigators to study both type 1 and type 2 diabetes.²³ Widely used in species as diverse as rabbits, rats, mice, pigs and cats, these models have advanced understanding of how diabetes develops and progresses, and have been valuable for testing treatments and for studying the disease more generally.^24,25

· Nephrotomy-STZ Model:

This double-model was used to more closely mimic diabetic nephropathy in rats, to observe the main biochemical and histological variations in a reduced period of time. It recapitulates aspects of metabolic disturbances of human DN, including hyperglycemia, hypoinsulinemia, azotaemia, hypertriglyceridemia, hypercholesterolemia and glycosylated proteins-important indicators of diabetic complications.²⁶

· Unilateral Nephrectomy (UNx) Combined with Diabetes Induction:

UNx-diabetes model The UNx-diabetes model is most frequently used for expediting the onset of diabetic nephropathy in experimental animals, most often in rodents. It was produced by the surgical ablation of one kidney and the chemical induction of diabetes, typically with streptozotocin (STZ), in which the contralateral kidney is exposed to even more hemodynamic load and metabolic pressure. Similarly, animals exhibit histologic features of human diabetic nephropathy, including albuminuria, glomerular hypertrophy, mesangial expansion and tubulointerstitial fibrosis. This two-hit model bears strong resemblance of the chronic and progressive process of diabetic kidney.²⁷

· Combined Surgical and Chemical Induction:

The combined surgical and chemical induction model can be used for a more intense study of diabetic nephropathy in a more standardised model. In this model, operations—e.g., ureteral ligation or unilateral nephrectomy, are combined with chemical agents—e.g., streptozotocin (STZ) exposure, to induce diabetes. This 'Double-Hit' intervention causes a simultaneous attack and significantly shortens the time period and the severity of damage to Kidney. The remaining kidney is exposed not only to metabolic stress through diabetes but also to increased functional demand through nephrectomy, resulting in marked pathological features, including albuminuria, glomerular hypertrophy, mesangial matrix expansion, and interstitial fibrosis. This model more closely mimics late stages of human diabetic kidney disease and is particularly relevant for assessing intervention strategies designed to arrest or reverse disease development.²⁸

Pancreatectomy with Uninephrectomy:

The pancreatectomy with uninephrectomy protocol presents a potent model to investigate diabetic nephropathy, where two independent surgical effects partial pancreatectomy to induce diabetes, and removal of one kidney to cause renal stress are combined. This combination replicates the metabolic and hemodynamic elements of diabetic kidney disease. This model has similar characteristics to those reported for the development of T2D-associated nephropathy that include sustained hyperglycemia, albuminuria, glomerular hypertrophy, and progressive renal fibrosis. It is particularly useful for examining the crosstalk between insulin deficiency and kidney damage.²⁹

Chemical Models:

Animal studies use chemically induced diabetes to reproduce diabetic nephropathy using drug injections. Streptozotocin (STZ) or alloxan are injected to destroy insulin-secreting pancreatic cells which result high blood sugar levels. Initially, the kidneys do not show any damage, but as time passes this prolonged hyperglycemia cause damage to kidneys, mimicking the process of diabetic kidney disease in Human. These models are sufficient for research purposes because they are easy to use, inexpensive.³⁰

Alloxan:

Alloxan is one of the earliest and most commonly employed agents in the production of T1DM and acts via selective destruction of pancreatic β-cells causing insulin shortage. Injecting it into animals (parenterally), it causes diabetes both in rabbits, rats, monkeys, cats, and dogs. It acts selectively taken up into β-cells through GLUT2 transporter, resulting in production of oxygen-free radicals which lead to necrosis of the β-cell.^31,32

Streptozotocin (STZ):

This method used for T1DM induction in rats and mice, STZ essentially breaks β‐cell DNA, resulting in their destruction. It is absorbed via GLUT2 and has species-specific effects, such as permanent b-cell damage in guinea pigs.^32,33

Dithizone:

Dithizone is one of the chemicals that scientists have used to induce diabetes in animals such as mice, rabbits, cats and hamsters by targeting the insulin-storing vesicles in the pancreatic β-cells, causing high blood sugar. Even though it now mostly serves to verify the purity of islet samples prior to transplant, it continues to provide an important contribution to diabetes research as a model to understand the development of the disease.

Nicotinamide and STZ Combination:

The stimulant injection of STZ and nicotinamide, usually combined with HFD, brings about T2DM in animals, and therefore, this approach provides significant information on the mechanism and treatment of T2DM.³⁴

Dexamethasone:

Rodents treated with high-level glucocorticoid, such as dexamethasone, also showed high levels of glucocorticoid-induced hyperglycemia and insulin resistance; however, they are less commonly used than those treated with alloxan and STZ.³⁵

Virus Models:

Coxsackie Virus B4 Model:

Coxsackie Virus B4 (CVB4) model is one of the most well-known animal models to simulate virus-induced diabetes, especially T1D. CVB4 infects pancreatic β-cells and elicits inflammatory and autoimmune processes resembling those occurring in human disease. This model facilitates the investigation of how infection with virus may trigger or accelerate autoimmune diabetes and provide insights into environmental triggers and immune mechanisms preceding β cell destruction and the appearance diabetic complications such as nephropathy.³⁶

Reovirus Models:

Models of reovirus infection are employed to investigate virus factors associated with type 1 diabetes development and its complications. In these models, reovirus‐induced infection in genetically susceptible models causes immune responses which mediate pancreatic β‐cell apoptosis and subsequent hyperglycemia. Although not directly applicable to research on diabetic nephropathy, these models provide useful information on the role that viral infections may play in the initiation or intensification of autoimmune diabetes and its renal complication.^37,38

Encephalomyocarditis (EMC) Virus Model:

The Encephalomyocarditis (EMC) virus model represents a widely used model to study virus-induced diabetes and its complications. Notably, EMC virus was demonstrated to incite acute-onset diabetes and nephropathy in sensitive mouse strains, as described for DBA mice, by the D-variant (EMC-D). These mice develop severe hyperglycemia and renal changes, including mesangial thickening and nodular glomerular lesions, within 2–3 months following infection. This model closely resembles certain aspects of human diabetic nephropathy.^39,40

Congenital Rubella Infection Model:

The congenital rubella infection model is a model in neonatal golden Syrian hamsters inoculated with rubella virus to mimic characteristics of human congenital rubella syndrome. These animals become chronically hyperglycemic and hypoinsulinemic, recapitulating the diabetic state in affected individuals. Such model is useful in investigating the mechanisms associated with virus-mediated β-cell injury and the autoimmune mechanisms in type 1 diabetes. It adds useful information about mechanisms by which prenatal viral infections may contribute to the development of autoimmune diabetes and its complications.⁴¹

Strain and Age Variability:

In models of diabetic nephropathy in animals, strain and age are strong factors for the development and severity of the disease. For example, mouse strain susceptibility to diabetic kidney injury is dependent with DBA/2 and 129/SvEv strains being more susceptible than C57BL/6 strain widely used in diabetes experimentation that are resistant to the development of nephropathy. Moreover, the age at which diabetes is induced becomes important when the onset and the degree of renal lesions are considered: in the older animals, albuminuria and the glomerular lesions are more severe. Understanding.^42,43

Host Susceptibility:

Host susceptibility contributes to the development and progression of diabetic nephropathy (DN) in animals. The genetic background plays a critical role in disease severity, such as 129 being more susceptible to DN, whereas C57BL/6 mice are resistant. This variation has been associated to differences in immune and inflammatory responses where susceptible strains have upregulated pro-inflammatory pathways. Knowledge of these genetic and immunologic (immunological) factors is essential for selection of suitable animal models and for the design of targeted therapies in DM.⁴⁴

Genetic Models:

Spontaneous Models:

Chinese Hamsters:

The Chinese hamster harbors spontaneously occurring diabetes, which closely resembles human T2DM. The infected animals experience hyperglycemia, polyuria, glycosuria, ketonuria and proteinuria. Histology shows decreased appearance of pancreatic islets and abnormal β-cell feature coupled with hepatic and renal alterations. The genetic pattern is believed to be autosomal recessive with the involvement of more than one gene suggested by genetic studies. This model is useful for investigating the genetic and metabolic factors of death of T2DM.^45,46

Non-Obese Diabetic (NOD) Mice:

Non-obese diabetic (NOD) mice are a commonly used spontaneous model of type 1 diabetes (T1D). The NOD mouse The NOD mouse is an autoimmune diabetic mouse derived from the Cataract Shionogi strain and is considered to be a natural occurrence model with autoimmune diabetes, which closely resemble human T1D in terms of genetics and immunology. In models of spontaneous diabetes, such as NOD, animals progress to the clinical syndrome and display insulitis and β-cell destruction, depending on sex, genetic, and environmental factors. T1D pathogenesis, and physiology have been dependent upon this model in order to test new potential therapies.⁴⁷

Targeted Mutations:

Targeted mutation models are indispensable for the investigation of diabetic nephropathy, since they enable researchers to examine the involvement of genetic defects in disease progression. Mice models such as Akita mouse, db/db mouse (leptin receptor mutation) and OVE26 mouse (β-cell-specific calmodulin overexpression) reproduce different types of diabetes and present important nephropathy features, like albuminuria and glomerular lesions. These models serve as tools for the discovery of molecular mechanisms and evaluation of potential therapies for diabetic kidney disease.^48,49

Autoimmune Models:

Autoimmune models, in particular the Non-Obese Diabetic (NOD) mouse and the Bio Breeding (BB) rat have played an important role in the study of DN. These models are predisposed to take up type 1 diabetes (T1D) naturally caused by immune attacks on pancreatic β-cells. NOD mice display multiple early indications of DN such as albuminuria and mesangial expansion and BB rats have thicker glomerular basement membranes and elevated glomerular filtration rate. These models closely resemble disease process in humans, and are useful for studying immunological mechanisms associated with DN.^50,51,52

Obesity Models:

Models of obesity, including the ob/ob and db/db mouse and Zucker diabetic fatty (ZDF) rat, are commonly utilized to assess type 2 diabetes and its renal complications. These animals have genetic defects causing obesity, insulin resistance, and hyperglycemia, which are major factors in the development of diabetic nephropathy. Subsequently, albuminuria, glomerular hypertrophy and interstitial fibrosis develop over time, mimicking the human disease. These models have been particularly useful for the investigation of metabolic pathways and the testing of strategies to treat obesity-associated kidney disease.⁵³

Polygenic Models:

Polygenic models, including the BTBR ob/ob mouse, are useful for investigating mechanisms of DN, as they contain multiple genetic factors associated with disease susceptibility. The BTBR ob/ob mouse, which carries genetic leptin deficiency combined with insulin resistance, manifests major morphological aspects of human DN such as early podocyte loss and mesangiolysis at onset of hyperglycemia. These models closely recapitulate the complex genetic and metabolic interactions of human DN, making these an important tool for dissecting disease mechanisms and assessing.^54,55

Transgenic Models:

Transgenic models are genetically modified animals for studying diabetic nephropathy with changing certain genes of kidney disease. Such models can be classified as overexpressing, knockout, knock-in, inducible, and tissue-specific, among others. Each permits investigators to study different parts of the evolution of disease, including inflammation, fibrosis, and glomerular injury. With models such as the OVE26 and eNOS−/− mice that can recapitulate human-like DKI, it thus becomes necessary to utilize transgenic strategies to investigate the pathogenesis of disease and examine targeted therapies.⁵⁶

Diet-Induced Models:

Diet-induced diabetes was first reported in 1947 by Houssay and Martinez, which involved diet as a manipulation in T2DM models.

Key Studies and Models:

· Sand Rats: Commercial Rat Chow caused obesity and diabetes; vegetable diet prevented these changes.

· Sprague-Dawley Rats: A 30% fat diet led to hyperphagia, obesity and hyperglycemia.

· Rabbits and Minipigs a/highsugar fat diet was administered to produce diabetes and insulin resistance.

· C57BL/6J Mice: on high-fat diets showed modest hyperglycemia and marked hyperinsulinemia.

· Spiny Mice and Nile Grass Rats: Demonstrated species-specific responses to diet-generated diabetes.

· Combination Models-High fat diets in combination with low doses of STZ, representing a more cost-effective and faster model for T2DM induction.^57,58,59

Gestational Diabetes Model:

There are a few strategies used to model GDM, none of which accurately replicate human disease some of the Methods are

· Late pregnancy total pancreatectomy.

· Chemical induction of pregnancy with alloxan and STZ.

· Diets Diet induction with high-CHO/high-fat.

· Genetic manipulation.

Organ-on-Chip Models:

Organ-on-chip (OoC) is a concept for microdevices that replicate the physiological or pathological conditions of designated organs, an integration of cell biology, engineering, and biomaterials for highly accurate human organ Modeling. Microscale organ-on-chip (OoC) models that mimic human organ function are driving the transformation of the study of diabetes. Glomerulus-on-a-chip models the kidney filtration primitive unit that can be used to investigate hyperglycemia induced kidney damage or diabetic nephropathy.⁶⁰ Pancreas-on-a-chip devices are engineered so as to study islet function and insulin secretion dynamics, which in turn can aid researchers to investigate β-cell dysfunction in diabetes. Diabetic foot ulcer-on-a-chip to investigate impaired wound healing in diabetic patients in a chronic wound-like microenvironment. Models of pancreas-on-a-chip emphasize islet function and hormone secretion kinetics and can be used to study dysfunction of β-cells in diabetes.⁶¹ Diabetic foot ulcer-on-a-chip is designed to replicate chronic wound conditions to study defective healing in individuals with diabetes. The use of patient-derived cells in OoC platforms represents a personalized disease model for patients, and significantly promotes the study on genetic and individual variability as they related to diabetic complications.⁶²

Strengths of Animal Models:

Animal models are still invaluable for understanding DN pathogenesis, for the identification of therapeutic targets, and for the preclinical evaluation of treatment effects but still animal model useful.

· Preclinical evidence: These models generate invaluable information on the natural history of DN and permit the investigation of chronically evolving renal injury, cellular alteration and metabolic effects of chronic hyperglycemia.

· Genetic Engineering: Recent progress in genetic tools like zebrafish and rodent models allows fine-tuned gene studies. It is worth mentioning that zebrafish and Drosophila have other advantages–low cost, clear embryos and easy genetic manipulation, that make them good tools of early screening.

· Physiological and genetic homology with human beings-There are many animals whose physiology and genetics are similar to humans: mammals such as rodents and even primates. This likeness enables scientists to investigate disease processes and treatments in an environment which closely mimics human biology.

· Experimental conditions Under control: Animal models allow researchers to manipulate variables in genetics, environment and diet to study individual components in the development and formation of disease. This control is essential to the setting of cause-and-effect relationships.^63,64

Limitations of Animal Models:

· Imperfect Disease Replication: The chronic and multifactorial nature of human DN is not completely reproduced often missing hallmarks of disease such as progressive glomerulosclerosis and significant proteinuria.⁶³

· Validation Limitations: Few models are validated against the strict standards set out by the Diabetic Complications Consortium, reducing their translational utility.⁶⁴

· Species-Specific Differences: Intrinsic physiological differences, especially renal structure and function exist between humans and animals that preclude the translation of findings.

· Augmented Induction Strategies: The mild kidney pathologies in T1D-DKD models are enhanced by dietary modifications, by combining them with other nephrotoxic insults, or by genetic alteration. Nonetheless, the potential nephrotoxicity involved in the application of a high dose of STZ may confound interpretable results through nonspecific cytotoxicity as it would directly damage the kidney independently of hyperglycemia.⁶⁵

· Resource requirements: The upkeep of animal colonies is costly, the ongoing care of animals is time-consuming, and ethical, humane use of animal’s dictates minimizing use in line with the 3Rs.

· Translation Gap: While pre clinical research has produced encouraging results, there is difficulty in translating into the clinic, therefore highlighting the need of human-relevant models for further study.⁶⁶

Recent Advances in Diabetic Nephropathy Research:

SUCNR1 Inhibitor (Cpd3): The effects of this molecule are the suppression of the NF-κB pathway, the attenuation of EMT induction and a decrease in ECM accumulation in high glucose–treated renal tubular epithelial cells—showing potential as a new anti-fibrotic therapy.⁶⁷

Glycated Urea: Monoglycated and diglycated urea in plasma were introduced also as possible sensitive glycemic control markers (blood [22/4] and/or urine) and released in most of diabetes publication that show correlation with similar methods.⁶⁸

Protein Glycation: Glycation end-products (AGEs) are the silver bullet of diabetic complications. More recent studies indicate that glycated amines may represent new diagnostic markers of diabetes and its renal sequelae.

Multi-Omics Biomarkers: It was possible to identify five leading multi-Dimensional Omics biomarkers—AFM, DUSP1, KRT19, TGFBI, and ZFP36—associated with the DN start and development, with ability to improve the diagnostic accuracy and be useful for setting targeted therapies through machine learning-assisted bioinformatics.⁶⁹

Inflammatory Markers: Elevated serum levels of TNF-α and surrogate markers (CRP, ferritin, urea, creatinine, uric acid) in type 2 diabetic patients further strengthen the crucial role of systemic inflammation in the patho and progression of DN.⁷⁰

DISCUSSION:

The use of model organisms is fundamental in scientific investigation to study human biology and disease and investigate the effects of new drugs and treatments. This is because many animal species, in particular mammals, have very similar genes, physiology and anatomic features, enabling scientists to replicate human conditions in a controlled setting. The Experiments on animal have several advantages to using animals versus humans including the possibility of performing experiments that could not be done for ethical or practical reasons on humans, availability of well-established disease systems for animals and relatively rapid life cycles for the study of several generations in much less time than is possible with humans. But there are disadvantages too. Ethical concerns regarding the welfare of the animals are another source of concern, since a lot of people are uneasy about the idea of screening drugs on animals. The use of animal model is necessary since there are currently no satisfactory models for duplicate the intricate pathogenesis of DN including hyperglycemia, inflammation, and fibrosis. Ethical use is defensible only to the extent that the expected benefits to human health outweigh the ethical costs.

The 3Rs must be used by researchers to reduce the ethical burden:

· Replacement: Use alternatives to animals (e.g., cell cultures, organoids) when available.

· Reduction: Use number of animals according to statistical considerations.

· Refinement: Optimize procedures to reduce pain and distress in animal.^71,72

· Pain management: Ensure the correct application of aesthesia and analgesics.

· Housing and Enrichment: Ensure high-quality housing conditions are possible to minimise stress and experimental variance.

· Humane Endpoints: Establish and use humane endpoints to minimize pain and distress. Any study of DN with animals requires prior approval from an IACUC or equivalent. Protocols must detail: Scientific objectives, Justification for animal use, Steps taken to minimize suffering

· Transparency and Integrity in Publication: Comprehensive reporting of methods and results, including negative results, are essential to prevent redundant experimentation in animals.

· Choice of Suitable Models: Selecting appropriate animal models (e.g., genetically modified mice with DN) provides meaningful data. The use of models that most closely resemble human DN can decrease the requirement of large numbers of repetitions

Challenges in Diabetes Nephropathy Modeling:

· Incomplete DN Pathology Recapitulation: The majority of rodent models, including Akita, db/db, and streptozotocin (STZ)-induced mice, do not completely demonstrate late-stage features of human diabetic nephropathy (DN). Although these models frequently show early DN features such as moderate albuminuria and glomerular hypertrophy, they usually do not present with established advanced DN features like nodular glomerulosclerosis, prominent tubulointerstitial fibrosis, and ESRD.⁷³

· Genetic and strain variability: Susceptibility to DN is highly strain-dependent among mice. For example, strains such as FVB or DBA/2J, show more severe nephropathic changes, while C57BL/6J mice are resistant. This genetic variation makes standardization and reproducibility in experimental DN studies difficult.

· Drawbacks of STZ-Induced Models: In addition to causing hyperglycemia by damaging pancreatic β-cells, STZ promotes direct nephrotoxicity. These dual effects can mask whether observed kidney pathology results from hyperglycemia or direct STZ Toxicity.⁷⁴

· Non-Heterogeneity of Disease in the Models: Various factors such as genetic predisposition, dyslipidemia, and hypertension contribute to human DNs. This heterogeneity is not fully reproduced in most animal models and thus reduces their translational potential.⁷⁴

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Received on 23.06.2025 Revised on 13.09.2025

Accepted on 10.11.2025 Published on 13.04.2026

Available online from April 15, 2026

Asian J. Pharm. Tech. 2026; 16(2):161-170.

DOI: 10.52711/2231-5713.2026.00023

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