Use of animal models in biomedical research in periodontics

Introduction

Animal models play a crucial role in biomedical research by providing insights into the complex biological processes that occur in humans. In fields like periodontics, where research focuses on understanding disease mechanisms, tissue regeneration, and the efficacy of treatments, animal models are invaluable. They bridge the gap between in vitro studies (experiments in controlled environments outside living organisms) and human clinical trials, helping researchers investigate diseases in a living organism’s context.

Rationale for using animal models in biomedical research

The use of animal models in biomedical research is driven by the need to understand complex biological processes, test hypotheses, and develop new therapies in ways that are not feasible or ethical in human subjects. One of the primary reasons for using animals in research is their biological similarity to humans. Many mammals, such as rodents, non-human primates, and pigs, share a significant portion of their genetic makeup, as well as similar anatomical, physiological, and biochemical characteristics. Mice, for example, share about 95-98% of their genome with humans, making them excellent models for genetic and molecular studies. These similarities allow researchers to study human disease mechanisms, drug responses, and treatment outcomes in a controlled environment that closely mimics the human body. Furthermore, non-human primates have a circulatory system, immune response, and organ structure highly similar to humans, making them invaluable for studying diseases like HIV or for organ transplantation research.

Conducting experimental research directly on humans, particularly in the early stages of scientific discovery or drug development, raises significant ethical issues. Animal models allow for the study of diseases, potential treatments, and side effects in a manner that would be unethical to test in humans, particularly when the safety and efficacy of the treatments are uncertain. Before drugs or medical procedures can be approved for human trials, regulatory agencies such as the FDA (U.S.) and EMA (Europe) require preclinical testing in animals to assess toxicity, safety, and biological activity. Animal studies help mitigate the risks associated with human experimentation by ensuring that treatments are as safe and effective as possible before being tested in humans. Along with this, animal models provide a controlled environment where researchers can manipulate variables such as diet, genetic makeup, exposure to pathogens, and environmental conditions. This level of control is often impossible in human studies due to variability in lifestyle, genetics, and uncontrollable environmental influences. Researchers can standardize animal populations (e.g., using genetically identical strains of mice) to ensure reproducibility and consistency in experimental outcomes. Animals can be subjected to a wide range of experimental conditions, from inducing diseases to testing new therapeutic approaches, in ways that would be difficult or impossible in human studies.

Animal models allow for in-depth investigation of the underlying mechanisms of diseases in a living system. They enable researchers to observe how diseases develop, progress, and respond to interventions over time. This is particularly important for studying complex diseases with multiple contributing factors, such as cancer, cardiovascular diseases, and neurodegenerative conditions. By inducing diseases in animals that closely resemble human conditions (e.g., diabetes in mice or cardiovascular diseases in pigs), researchers can investigate the progression of disease and identify molecular pathways that may serve as therapeutic targets. Different models allow for the study of both chronic conditions (e.g., chronic inflammatory responses in periodontal diseases) and acute responses (e.g., wound healing or immediate immune responses).

Animal models are indispensable in the development and testing of new drugs, surgical techniques, and medical devices. They provide a platform to assess the efficacy, safety, dosage, and side effects of novel treatments before human clinical trials. Animal studies are critical for testing new pharmaceuticals for various conditions, including periodontitis, cancer, and infectious diseases. Animal models help determine the therapeutic window (the dosage at which a drug is effective but not toxic) and identify potential adverse effects. Furthermore, in periodontics and other fields, animal models are used to test the efficacy of biomaterials (e.g., bone grafts, scaffolds) for tissue regeneration. For example, dogs or pigs may be used to test new approaches for regenerating bone and soft tissues around teeth and implants.

In vitro experiments (e.g., cell cultures) and computer models are useful for understanding basic biological processes, but they cannot replicate the complexity of whole organisms. Animal models provide a bridge between in vitro studies and human clinical trials by allowing researchers to observe how cells, tissues, and organs interact in a living system. Animals also offer insights into how localized treatments (e.g., periodontal therapies) affect the entire body. For instance, periodontal inflammation in animals can help researchers understand its links to systemic conditions like cardiovascular disease or diabetes. Animal models allow us to study the immune response, healing processes, and interactions between different systems (e.g., endocrine and immune systems) which can provide a more holistic understanding of disease processes. Also, animal models allow researchers to observe disease progression and treatment effects over extended periods, which would be difficult to achieve in human studies. This is particularly useful for studying chronic diseases or long-term effects of treatments. In periodontics, long-term animal studies have helped elucidate the progression of periodontal disease, bone loss, and the effects of regenerative therapies over time. Animals with shorter lifespans allow researchers to study aging processes, which is crucial for understanding diseases that develop over many years in humans, such as Alzheimer’s disease or osteoporosis.

With advancements in genetic engineering, animals can be genetically modified to mimic human diseases. This has led to the creation of transgenic animals that develop specific conditions, allowing researchers to study the genetic basis of diseases and test potential gene therapies. Genetically modified mice are used to study the role of specific genes in disease. For example, “knockout” mice that lack certain immune system genes can help researchers understand the role of these genes in inflammatory responses associated with periodontal disease. Some animals are engineered to express human genes, making them even more accurate models for studying human diseases and testing therapies.

Types of animal models in biomedical research in periodontics

Various animal models are used depending on the research goal, whether it is to study a specific disease, evaluate therapeutic efficacy, or explore biological processes. These models are chosen based on their relevance to the condition being studied, their genetic and physiological similarities to humans, and practical considerations such as availability and cost. Following is the detailed description of these animal models,

Rodent models

Rodent models, particularly mice and rats, are among the most widely used animals in biomedical research due to their physiological, genetic, and anatomical similarities to humans. They are versatile, easily bred, cost-effective, and can be genetically manipulated, making them invaluable in a wide range of scientific fields including genetics, immunology, neuroscience, pharmacology, and disease modeling. There are multiple advantages of using rodent models. Mice and rats share approximately 95-98% of their genes with humans, making them excellent models for studying human diseases. Many of the biological pathways that lead to disease are conserved between rodents and humans, allowing researchers to extrapolate findings to human conditions. Rodents can develop many human-like diseases either spontaneously or through genetic manipulation, including diabetes, cancer, cardiovascular disease, and neurological disorders. Furthermore, genetic manipulation can be easily done in rodent models, such as,

• Knockout mice: Mice and rats can be genetically modified to either express or suppress specific genes, allowing researchers to study the role of these genes in disease progression. In knockout mice, specific genes are “knocked out” or deactivated, allowing researchers to study the function of those genes. This has been essential in studying gene-specific diseases, including cystic fibrosis, cancer, and heart disease.
• Transgenic models: Transgenic mice carry foreign genes that have been deliberately inserted into their genome. These models are used to study gene function, mimic human diseases, and test gene therapies. For example, transgenic mice have been used to model neurodegenerative diseases such as Alzheimer’s, where researchers introduce human genes associated with the condition.
• Immunodeficient mice (SCID or Nude Mice): The immunodeficient mice lack a functional immune system and are used to study cancer, immunology, and for testing human cell or tissue transplantation (e.g., in cancer therapy research).
• Humanized Mice: These are genetically engineered to carry human genes, cells, or tissues, making them particularly useful in studying human diseases such as infectious diseases (HIV, hepatitis) and immune responses.

Mice and rats have short gestation periods (about 20 days for mice), allowing researchers to observe multiple generations in a relatively short amount of time. This is particularly valuable in studies involving genetics, aging, or long-term disease progression. Rodents are extensively used to study cancer biology, from identifying oncogenes and tumor suppressor genes to testing novel cancer therapies. Genetically modified mice, such as P53 knockout mice, are used to study tumor development and metastasis.

Limitations of Rodent Models

Despite their widespread use, rodent models have limitations:

• Genetic differences: Although rodents share many genes with humans, the differences in gene expression and regulation can lead to discrepancies in how diseases manifest or respond to treatments.
• Physiological differences: Certain aspects of rodent biology, such as metabolic rates, immune responses, and lifespans, differ significantly from humans, which can limit the direct applicability of findings to human health.
• Ethical considerations: The use of rodents in research raises ethical concerns regarding animal welfare. Researchers are required to follow strict guidelines to ensure humane treatment and minimize suffering.

Non-human primate models

Non-human primates including monkeys, baboons, and macaques, are among the most valuable and closely related animal models for studying human diseases. They are used in a range of biomedical research fields because of their close genetic, anatomical, and physiological similarities to humans. Their use is especially important for investigating complex diseases, drug testing, and understanding human biology where smaller mammals, like rodents, are less applicable. The non-human primates share about 90-98% of their DNA with humans, making them particularly useful for studying human-specific diseases. For example, rhesus macaques have a genetic makeup that is approximately 93% similar to humans, and chimpanzees, although used less frequently due to ethical concerns, share over 98% of their genes with humans. Along with this, they exhibit complex behaviors, social structures, and cognitive abilities that are remarkably similar to humans, making them highly relevant models for studying neurological, psychological, and behavioral disorders. The immune system of NHPs closely mirrors that of humans, which is critical for studying infectious diseases, vaccines, and immunotherapy.

Unlike rodents, which have short lifespans, non-human primates live longer, making them ideal for studying chronic diseases and conditions that develop over time, such as neurodegenerative diseases, cardiovascular diseases, and cancer. Their longer life span allows for longitudinal studies that track disease progression, therapeutic response, and aging. The non-human primates play a key role in preclinical drug testing, particularly for drugs that require testing in species with physiological systems very close to humans. Their use is often a regulatory requirement for drug approval processes, especially for testing the safety and efficacy of biologics (drugs derived from living organisms), monoclonal antibodies, and vaccines. However, it should be remembered here that use of non-human primates in research is tightly regulated due to ethical concerns, given their high level of cognitive abilities and social behaviors. Research using NHPs is typically only permitted when no other animal model can be used, and the potential benefits of the research are considered to outweigh the ethical costs. Guidelines for non-human primate research emphasize the 3Rs (Reduction, Refinement, and Replacement) to minimize harm.

Some of the non-human primates used in research include,

Rhesus Macaques (Macaca mulatta)

Rhesus macaques are the most widely used non-human primate species in biomedical research due to their relatively easy handling, availability, and close genetic similarity to humans. These are extensively used to study HIV/AIDS, using the simian immunodeficiency virus (SIV) model, which mirrors the progression of HIV in humans. They are also vital in testing antiretroviral therapies and potential vaccines. They are a key model for testing vaccines for diseases such as influenza, Zika, and COVID-19 due to their similar immune response to humans. Rhesus macaques are used in cognitive and neurological studies, including Alzheimer’s disease, Parkinson’s disease, and stroke research.

Cynomolgus Monkeys (Macaca fascicularis)

Cynomolgus monkeys are commonly used in pharmacological and toxicological studies due to their genetic similarity to humans and their smaller size compared to rhesus macaques. These monkeys are often used in studies that investigate how drugs are absorbed, distributed, metabolized, and excreted in the body. Cynomolgus monkeys are used in brain research, particularly in models of neurodegenerative diseases, stroke, and drug addiction.

Baboons (Papio spp.)

Baboons are larger primates with similar cardiovascular systems and reproductive biology to humans, making them ideal for studying certain human conditions. These are used to study hypertension, atherosclerosis, and other cardiovascular diseases. They are often used in research involving reproductive technologies and pregnancy, due to their similar gestational physiology to humans. Baboons are also used in research on infectious diseases like hepatitis and tuberculosis.

Marmosets and Tamarins (Callithrix and Saguinus)

Marmosets and tamarins are smaller primates that are increasingly being used for neuroscience and aging research. Marmosets are becoming a popular model for studying neurodegenerative diseases like Parkinson’s and Alzheimer’s, as their small size allows for easier handling, and they exhibit age-related cognitive decline similar to humans. They are also used to study aging and longevity due to their relatively short lifespan compared to larger primates.

Limitations of non-human primate models

While non-human primate models provide unparalleled value in biomedical research, they have some limitations:

• Ethical concerns: The use of NHPs raises significant ethical issues due to their cognitive abilities and close relationship to humans. The high ethical burden limits their use to only the most critical studies.
• Cost and logistics: NHP research is expensive and requires specialized facilities and personnel for proper care and handling.
• Species differences: Despite their genetic and physiological similarities to humans, NHPs are not identical, and species-specific differences can still affect the translation of research findings to humans.

Canine (dog) models

The dog models share many anatomical and physiological features with humans, particularly in areas like the cardiovascular system, respiratory system, musculoskeletal structure, and gastrointestinal tract. This makes them ideal models for studying heart diseases, orthopedic conditions, and metabolic diseases. Unlike rodent models where diseases must be artificially induced or genetically engineered, dogs naturally develop many of the same diseases as humans, including cancers, diabetes, and arthritis. This provides a more natural model for studying disease progression and therapeutic interventions. Along with this, domestic dogs have a high degree of genetic diversity due to selective breeding practices. This makes them valuable for studying genetic diseases and understanding how specific genetic traits contribute to disease. Compared to rodents, dogs are closer in size to humans, which allows for more accurate testing of medical devices, surgical techniques, and prosthetics. This is particularly important in cardiology, orthopedics, and dentistry, where device testing requires a model that can replicate human-scale anatomy.

Canine models are valuable for periodontics research due to their similarities to humans in oral anatomy, periodontal structure, and immune response. Dogs, like humans, naturally develop periodontal diseases, such as gingivitis and periodontitis, making them a practical and relevant model for studying the pathogenesis of periodontal diseases, treatment approaches, and tissue regeneration techniques. The use of canine models has provided insights into periodontal disease mechanisms, therapeutic interventions, and dental implant technologies. Canine models have an immune response to periodontal pathogens that is comparable to humans. The inflammatory process involving cytokine release, immune cell recruitment, and tissue destruction follows similar patterns, making it easier to translate research findings into human treatments. The bacterial communities found in the canine oral cavity, while not identical, share many similarities with human periodontal pathogens. Species like Porphyromonas gingivalis are found in both human and canine oral microbiomes, which is useful for studying microbial interactions and host responses.

Applications of canine models in periodontics research

Pathogenesis of Periodontal Disease

Canine models are used to study how dental plaque forms and induces periodontal inflammation, leading to tissue destruction and bone loss. Research often focuses on how bacterial biofilms trigger immune responses and the role of specific bacteria in periodontitis progression. Studies using canine models explore the interactions between oral microbiota and the host’s immune response. Understanding these interactions is crucial for developing therapies that target the microbial communities involved in periodontal diseases.

Periodontal Regeneration

Canine models are widely used to test the efficacy of guided tissue regeneration (GTR) techniques. These involve the use of barrier membranes to promote the regrowth of periodontal tissues, including bone, cementum, and the periodontal ligament. Research in dogs has demonstrated the effectiveness of various barrier materials, such as resorbable and non-resorbable membranes, in promoting bone regeneration and restoring periodontal structures. Bone grafts and biomaterials are frequently tested in canine models to assess their ability to regenerate lost alveolar bone in periodontal defects. This research helps develop new biomaterials for clinical use in human periodontal therapy. Autografts, allografts, and xenografts have been evaluated in canine models to determine their effectiveness in periodontal defect repair. Synthetic biomaterials, such as bioactive glass and calcium phosphate-based scaffolds, are also tested in canine models for their potential to support bone regeneration.

Testing of dental implants and osseointegration

Canine models are valuable for testing the placement of dental implants and studying the process of osseointegration. The bone density and structure in dogs are similar to those of humans, making them an appropriate model for implant dentistry. Research in canine models has focused on the effects of implant surface modifications, such as roughened or bioactive surfaces, on the rate and quality of osseointegration. This has contributed to the development of advanced implant technologies used in human dentistry. Dogs are also used to model peri-implantitis, a condition characterized by inflammation and bone loss around dental implants. Studies in canine models help understand the etiology of peri-implantitis and evaluate the effectiveness of different treatment strategies.

Periodontal pharmacotherapy

Canine models are used to test the efficacy of systemic and local antimicrobial therapies for treating periodontal infections. These studies help determine the most effective antibiotic regimens for managing periodontal pathogens and preventing disease progression. Research has focused on locally delivered antimicrobials, such as minocycline microspheres and chlorhexidine chips, which are placed directly into periodontal pockets. Canine models help evaluate the effectiveness and safety of these treatments. Therapies aimed at modulating the host’s immune response to periodontal disease have been tested in canine models. These therapies include drugs that inhibit matrix metalloproteinases (MMPs), which contribute to tissue breakdown, and anti-inflammatory agents that reduce the destructive immune response.

Wound healing and soft tissue management

Canine models are used to study the healing of soft tissue grafts in the management of gingival recession and other periodontal defects. The success of different graft materials and techniques, such as autogenous grafts or allogeneic grafts, can be evaluated in terms of tissue integration and aesthetic outcomes. Advances in tissue engineering, such as the use of growth factors (e.g., platelet-derived growth factor, bone morphogenetic proteins) and stem cells, are often tested in canine models. These studies explore ways to enhance the healing and regeneration of both hard and soft tissues in periodontics.

Ethical Considerations in Canine Periodontics Research

The use of canine models in periodontics research is subject to strict ethical regulations to ensure the humane treatment of animals. Researchers must follow guidelines such as the 3Rs principle (Reduction, Refinement, and Replacement), which emphasize minimizing animal use, refining experimental techniques to reduce suffering, and replacing animal models with alternative methods where possible. Institutional Animal Care and Use Committees (IACUCs) oversee the ethical use of animals in research, ensuring that all experiments involving dogs are necessary and that their welfare is protected.

Limitations of Canine Models in Periodontics

While canine models are valuable for periodontics research, there are limitations to their use:

• Species differences: Despite their similarities, dogs are not identical to humans in terms of their oral microbiome and immune response. These differences can affect the translation of findings to human clinical practice.
• Ethical considerations: The use of companion animals like dogs raises ethical concerns, particularly when alternative methods, such as in vitro models or computer simulations, might be available for certain studies.
• Cost and logistical challenges: Canine models are more expensive and require more complex housing and care than smaller animal models like rodents, limiting their widespread use in research.

Swine (Pig) models

Swine, particularly domestic pigs (Sus scrofa), are increasingly recognized as valuable animal models in biomedical research due to their anatomical, physiological, and genetic similarities to humans. Pigs are often used in various fields, including cardiovascular, metabolic, orthopedic, and periodontal research, making them suitable for studying complex diseases and testing new therapies. Their size, ease of handling, and availability of specific breeds that develop human-like diseases have made them an essential model in translational research. Pigs are larger than traditional rodent models, allowing for procedures and interventions that closely resemble human applications. Their anatomical structures, such as the cardiovascular system, respiratory system, and digestive tract, have significant similarities to those of humans, making them relevant models for studying human diseases. Pigs possess a skin structure similar to humans, making them valuable for dermatological research. Their musculoskeletal system also allows researchers to study orthopedic conditions and treatments. Swine share a considerable degree of genetic homology with humans, with approximately 80% of human genes having a counterpart in pigs. This genetic similarity enhances the relevance of findings in swine studies to human health. Research using swine has elucidated the role of specific bacteria in periodontal disease progression, contributing to the understanding of the microbiome’s impact on oral health. Canine-derived regenerative materials and biomaterials have been tested in swine models, leading to advances in periodontal surgery and tissue regeneration techniques.

Limitations of Swine Models in Research

While swine models offer numerous advantages, there are limitations to their use:

• Cost and logistics: Swine are more expensive to house and care for than smaller models like rodents. This can limit their widespread use in research, particularly in preliminary studies.
• Species-specific differences: Despite their similarities, swine are not identical to humans in all aspects of physiology and disease. These differences can affect the generalizability of research findings.
• Ethical concerns: The use of swine in research raises ethical questions regarding animal welfare, particularly given their cognitive abilities and social behaviors. Researchers must carefully justify the use of swine and ensure compliance with ethical standards.

Conclusion

In summary, animal models are essential for advancing biomedical research, particularly in periodontics. Their use helps to bridge the gap between theoretical research and practical applications, offering a living context for understanding complex biological interactions and testing new treatments. Despite their contributions, ongoing efforts focus on refining and replacing these models to ensure ethical responsibility in research.