Journal of Indian Society of Periodontology

: 2016  |  Volume : 20  |  Issue : 4  |  Page : 360--368

Ethical guidelines, animal profile, various animal models used in periodontal research with alternatives and future perspectives

Mohan Kumar Pasupuleti1, Subramanya Shetty Molahally2, Supraja Salwaji3,  
1 Department of Periodontics, St. Joseph Dental College, Duggirala, Eluru, West Godavari, Andhra Pradesh, India
2 Department of Orthodontics, Yenepoya Dental College, Mangalore, Karnataka, India
3 Department of Oral and Maxillofacial Pathology, St. Joseph Dental College, Duggirala, Eluru, West Godavari, Andhra Pradesh, India

Correspondence Address:
Mohan Kumar Pasupuleti
Flat No. 302, Sai Swarna Residency, Sriram Nagar - 9th Road, Eluru, West Godavari, Andhra Pradesh


Laboratory animal models serve as a facilitator to investigate the etiopathogenesis of periodontal disease, are used to know the efficacy of reconstructive and regenerative procedures, and are also helpful in evaluation of newer therapeutic techniques including laser and implant therapies prior to application in the human beings. The aim of this review is to know the different animal models used in various specialties of dental research and to know the ethical guidelines prior to the usage of experimental models with main emphasis on how to refine, replace, and reduce the number of animal models usage in the laboratory. An online search for experimental animal models used in dental research was performed using MEDLINE/PubMed database. Publications from 2009 to May 2013 in the specialty of periodontics were included in writing this review. A total of 652 references were published in PubMed/MEDLINE databases based on the search terms used. Out of 245 studies, 241 were related to the periodontal research published in English from 2009 to 2013. Relevant papers were chosen according to the inclusion and exclusion criteria. After extensive electronic and hand search on animal models, it has been observed that various animal models were used in dental research. Search on animal models used for dental research purpose revealed that various animals such as rats, mice, guinea pigs, rabbit, beagle dogs, goats, and nonhuman primates were extensively used. However, with the new advancement of ex vivo animal models, it has become easy to investigate disease pathogenesis and to test the efficacy of newer therapeutic modalities with the reduced usage of animal models. This review summarized the large amount of literature on animal models used in periodontal research with main emphasis on ethical guidelines and on reducing the animal model usage in future perspective.

How to cite this article:
Pasupuleti MK, Molahally SS, Salwaji S. Ethical guidelines, animal profile, various animal models used in periodontal research with alternatives and future perspectives.J Indian Soc Periodontol 2016;20:360-368

How to cite this URL:
Pasupuleti MK, Molahally SS, Salwaji S. Ethical guidelines, animal profile, various animal models used in periodontal research with alternatives and future perspectives. J Indian Soc Periodontol [serial online] 2016 [cited 2017 Mar 27 ];20:360-368
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Full Text


It is important to do an experimental research to find the origin and the pathology of the disease process on animal models prior to the clinical trials on humans. Hence, the animal models are used to test the efficacy and effects of restorative materials on dental pulp; to evaluate the etiopathogenesis, clinical characteristics, and histological and immunologic aspects of periodontal disease; and to test the success of newer surgical techniques prior to its application on humans.[1],[2] Gingival and periodontal diseases are a group of chronic inflammatory diseases of supporting structures of periodontium. These periodontal diseases are mainly caused by oral bacterial biofilm in the periodontium. To prevent these diseases, it is important to know the disease etiology, pathogenesis, use of biomaterials, tissue responses to the newer techniques in animal models prior to the application in human beings.[3],[4]

Collection of data

An online search on animals models used for dental research purpose was performed using MEDLINE/PubMed databases using the following search terms: “animal models,” “experimental models,” and “laboratory models.” The term “periodontal” was added to all the search terms. Relevant papers were chosen according to the inclusion and exclusion criteria. A flowchart showing the steps used for study selection is illustrated in [Chart 1].[INLINE:1]

Inclusion criteria

A thorough screening of the titles, abstracts, and full texts of the articles by both electronic and hand search was carried out by a single examiner.

All chosen studies detailed the following:

All the papers published in English language only were included in this review All the study articles irrespective of the specialty journals, including reviews which were related to the periodontal research, were searched and included to write a review from the journals published from 1980 to May 2013 Animal studies in the specialty of periodontics from 2009 to 2013 journals, indexed in PubMed, were included in writing the commonly used various animal models and the subject of interest in periodontal research Also included the various recent articles/handbooks and current guidelines provided by the governing bodies online till date on animal testing and welfare act.

Exclusion criteria

Animal studies other than periodontal research were not included in the study.

Outcome measures and search strategy

Although the outcome measures of clinical, radiographical, histopathological, immunological, and regenerative studies have shown positive correlation in animal studies, it is the time to reduce, refine, and replace the animal models used for experimental studies due to ethical issues. Due to the recent extensive research on ex vivo animal models, this is the right time to switch on to the most advanced technology which helps in restricting the animal models usage participating in dental research.

 Various Animal Models Used

A wide range of animals is used in dental research including nonhuman primates, dogs, rats, hamsters, ferrets, and other species with definite advantages and disadvantages [Table 1].[5],[6],[7],[8],[9],[10]{Table 1}

In the last two decades, majority of the experimental animal models were used by various departments of dentistry. Most of the animals used for research purposes in dentistry were purely for teaching or educational training purposes.[11],[12],[13],[14],[15],[16] In many of the dental research institutes, small animal models were used. Larger animals were reserved as the last choice for validation of newer surgical techniques prior to the use in human beings.[17],[18],[19],[20],[21],[22]

To understand the origin and the pathology of the periodontal disease process, it is important to do an experimental research.[23],[24],[25],[26] Experimental animal models are used to evaluate the etiopathogenesis, clinical characteristics, and histological and immunologic aspects of periodontal disease.[27],[28],[29],[30],[31]

Animal models are also used to assess the advanced surgical procedures such as lasers and implant research and to evaluate newer drug efficacy before applying onto clinical trials.[32],[33],[34],[35],[36] It is very important in selecting an animal model which has the anatomical structures similar to the humans.[37],[38],[39],[40],[41] Thus, animal models are used in almost all the specialties of dentistry to increase the knowledge on disease pathology and newer treatment approaches to prevent the dental diseases at an earlier stage.[42],[43],[44],[45],[46]

All the clinical studies and systematic reviews from 2009 to May 2013 in the specialty journals of periodontics included in this review summarized that various types of animal models were used in periodontal research.[47],[48],[49],[50],[51] Various studies in chronological order, listing the various types of animal models used in periodontal research with their respective research topic, are enumerated in [Table 2].[52],[53],[54],[55],[56]{Table 2}

Benefits of animal models in research

The following are the benefits of animal models use in research.[57],[58],[59],[60],[61],[62]

The use of animals in research has made a substantial contribution to the understanding of biological processes It has been responsible for many important biomedical discoveries It used in the development of a great number of therapies and preventative treatments, such as antibiotics, insulin, vaccines, and organ transplantation.

 Guidelines to Use Animals in Research

The Indian National Science Academy developed the updated guidelines to use of animals in scientific research.[63],[64],[65] By knowing the guidelines to use of animals in research, it is easy to follow the ethical guidelines.

Depending on the need, one should allow and provide facilities to use animal models for the research purpose It is recommended to the researchers that one should not use animals unless until it is an unavoidable situation. They should also ensure that unnecessary pain or injury is avoided It should provide adequate care, housing, and make sure that the animal models used for research purpose are physically comfort and in good health Sources of experimental animals for research should be from recognized animal facilities where there will be availability of genetic, health, and nutritional status of each animal model It should provide training facilities to the scientific researcher and the supporting staff those who take care of the animals during experiment Alternative animal models should be used to replace experimental animals wherever possible Procedures which are painful should ensure appropriate use of anesthesia Forming ethical committees will ensure the minimal usage of animal models.

 Ethical Guidelines

All the scientists who are working with experimental animal models should follow the ethical guidelines at institutional or national level before starting their research work. Every individual should strictly adhere to the animal ethics committee.[66]

An animal ethics committee should include a senior biological scientist at the institute, two scientists from different biological disciplines, a veterinarian, the scientist-in-charge of animal facility, a scientist from outside the institute, a nonscientific socially aware member, a member or nominee of appropriate regulatory authority of Government of India, and a specialist may be needed to review special projects involving radiation exposure and pathogenic microorganisms.[67]

This committee should involve in examining the animals, scientists, and the technicians handling the animals before starting the experiment. The following are some of the ethical guidelines to be followed for use of animals in dental research.[68]

Animal studies should be started only when they are relevant to increase the human or animal health and when they increase the knowledge in promotion of health Make sure that minimum number of animal models should be selected Treat animals with kindness and make sure that adequate living conditions are provided Experiments that cause discomfort or pain should be performed under anesthesia Animals that cannot be relieved or repaired at the end of the study should be killed under anesthesia Every individual should be qualified or should have experience for conducting animal research work In vitro methods and ex vivo animal models can be used to reduce the number of animal use wherever possible The most important guidelines to follow while doing research on experimental animal models are the principle of three Rs. The Rs introduced by Russel and Burch is replacement, reduction, and refinement. The objective of three Rs when the experimental animals used during the research should include that the animals are used only when it is absolutely necessary, the number of animals used for study purpose should be kept minimum, and the suffering of the animals during study is also minimized In addition, the Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA) in India inculcated the credo of 4 Rs – replacement, reduction, refinement, and rehabilitation of animals used in experimentation. CPCSEA has made it a national policy that rehabilitation of animals postexperimentation be a part of the research. By adding the 4th Rs to the research field, it guides the researcher to pay additional responsibility in using animal models postoperatively.[69]

 Regulations of Animal Testing With Animal Welfare Act

Best animal welfare results in the best science. Animal welfare refers to the state of the animal; the treatment that an animal receives is covered by other terms such as animal care, animal husbandry, and humane treatment. At present, animal welfare became an important concern while doing experimental studies on animals. Many of the regulatory bodies are showing concern about the animal welfare which in turn provides guidelines for proper animal care and usage during research.[68]

The regulatory bodies came into existence only after the pressure from the animal protection groups and public. These regulatory groups enacted laws to regulate the care and usage of laboratory animal models.

Many regulatory bodies are established to guide the care and use of laboratory animal models during research. Responsible regulatory decisions should be taken prior to the animal testing based on the ethical requirement of animal for research purpose. Regulation of animal testing will not only reduce the usage of animals but also help in assessing the data of already done research or helping in harmonizing the test methods. It is also important that regulations of animal testing should allow the advanced research tests by clearly defining the scope and limitations to the regulatory bodies.

The regulatory bodies formed a consultative group and framed five major principles for the utilization and care of animals used in research and testing.[68],[69],[70],[71]


Experiments on animals may be carried out for the purposes of advancement by new discovery of physiological knowledge and which will gain in significant well-being of the people The lowest animals on the phylogenetic tree which gives significant results in animal testing should be considered Avoid or minimize pain on distress when animals are used for research procedures. All the procedures should perform with adequate sedation, analgesia, or anesthesia Investigators are responsible for the postoperative care of the experimental animals Proper care should be taken to provide living conditions of animals.

 Animal Profile

It is important to know the animal profile prior to the use in research. Animal profile includes physiological norms of commonly used laboratory models such as mouse, rat, hamster, guinea pig, rabbit, cat, dog, and monkey. Age at maturity, adult weight, respiratory rate, rectal temperature, pulse rate per minute, and life span are some of the physiologic norms of the healthy animal models to follow during selection for research.[72]


Age at maturity: 6–8 weeks, adult weight: 25–30 g, respiratory rate: 90–180 per min, rectal temperature: 37.4°C, pulse rate per min: 600 on average, and life span: 1.5–2 years.


Age at maturity: 10–12 weeks, adult weight: 200–300 g, respiratory rate: 80–150 per min, rectal temperature: 37.5°C, pulse rate per min: 300 on average, and life span: 2.5–3 years.


Age at maturity: 6–8 weeks, adult weight: 80–100 g, respiratory rate: 40–120 per min, rectal temperature: 37.6°C, pulse rate per min: 450 on average, and life span: 1.5–2 years.

Guinea pig

Age at maturity: 16–20 weeks, adult weight: 400–500 g, respiratory rate: 60–110 per min, rectal temperature: 38.6°C, pulse rate per min: 150 on average, and life span: 4–5 years.


Age at maturity: 24–32 weeks, adult weight: 2–2.5 kg, respiratory rate: 35–56 per min, rectal temperature: 38.7°C, pulse rate per min: 133 on average, and life span: 4–5 years.


Age at maturity: 30–35 weeks, adult weight: 3–5 kg, respiratory rate: 20–30 per min, rectal temperature: 39.5°C, pulse rate per min: 110 on average, and life span: 8–12 years.


Age at maturity: 1–1.2 years, adult weight: 12–15 kg, respiratory rate: 14–28 per min, rectal temperature: 38.6°C, pulse rate per min: 95 on average, and life span: 10–15 years.


Age at maturity: 4–5 years, adult weight: 10–12 kg, respiratory rate: 30–54 per min, rectal temperature: 38.4°C, pulse rate per min: 200 on average, and life span: 15–20 years.

 Alternatives to Experimental Animal Models

With the order of prevention of cruelty to animals act, 1960, many of in vitro systems can be used to reduce or replace animals. Both the living and nonliving systems could be used in dental research to reduce number of animals in experimentation.[73]

To overcome the disadvantages and unethical procedure of using experimental animal models, alternative models have been introduced. The advantages of alternative methods are time efficiency, less manpower, and cost-effectiveness. Anything from absolute to partial replacement of live animals in biomedical research and testing is considered as alternative or substitute to the animal testing.

Many of the organizations dedicated for the development of test methods that replace the live animal models. Research methods, when superior to the use of animals to learn about human disease or predict the safety of new drugs, are termed as alternatives to animal models. Stem cells, microdosing, DNA chips, microfluidics chips, human tissue, new imaging technologies, and postmarketing drug surveillance are few examples of alternatives to animal testing.[74]

Computer models, cell and tissue cultures, and various alternative organisms such as invertebrates, lower vertebrates, and some microorganisms such as prokaryotes, protists, and fungi also serve as alternatives to the most of the experimental animal models. Computer models with the specialized software are used to study the biologic effectiveness of the drugs, and the other alternatives such as organisms are mostly used as model for molecular and genetic studies.[75]

No doubt that these alternatives will minimize the usage of animal models, but they cannot completely eliminate their usage in preclinical studies.

Computer models

The use of computers in predicting the various biological and toxic effects of a chemical, one can reduce the usage of animals. For example, computer-aided drug design helps in identifying the receptor binding site for the potential drug and thus avoids testing of unwanted drugs which have no biological activity on animals.


It is less expensive than using experimental animal models and also the total number of animal usage for experimentation is lowered.

Cells and tissue cultures

Cell and tissue cultures in a laboratory are an important alternative for various research purposes. Toxicity and efficacy of every drug can be tested by these methods.


It is less time-consuming and less expensive.

Alternative organisms

Different model organisms are used instead of higher vertebrates. For example, zebrafish, which is a lower vertebrate, can be used as an attractive alternate to study molecular, genetic, and cancer research.

Various invertebrates such as fruit fly and eukaryotic nematode are used to study various diseases such as Parkinson's disease, Alzheimer's disease, diabetes, and cancer.


It has less ethical issues, less laboratory expenses.


Brewing yeast is the most important organism used as an alternative to experimental animal models. Brewing yeast is used to understand programmed cell death, cell death regulators in humans and is very useful in cancer research.


It is less ethical issues, less expensive.

 Experimental Models Versus Ex Vivo Models

Experimental animal models are used as an adjunct or supplement by the researcher or scientist to decide whether an experiment or test on an animal is likely to produce a useful result in humans.

Besides the major concern of ethics, there are a few more disadvantages of animal experimentation such as requirement of skilled manpower, time-consuming protocols, and high cost. Moreover, very high cost involved in breeding, housing, and lengthy protocols of animal experiments is another drawback of experimental animal models.[76]

With the development of ex vivo culture model, with the support of different culture medium, it has become easy to investigate the inflammatory cell behavior and bone metabolism in periodontal disease type. Thus, the ex vivo animal models not only allows in studying the different molecular pathogenic mechanisms of periodontal disease but also applies to the development of novel therapies in which disease can be identified at an earlier stage and treated with well-advanced techniques. Hence, it is clear that development of ex vivo model definitely reduces the usage of experimental animal models and also acts as a promising alternative to the in vivo studies.[76]

As the modern biomedical research is facing multifactorial, infectious, and complex diseases, it is difficult to use more number of experimental animal models for the research purpose. The two main objectives for the future research on animal models are to develop more ex vivo culture models which are relevant and more informative for the periodontal research and also to follow all the protection requirements of animal models by the investigator before and after the research.

To refine, reduce, and replace the animal models, it is suggested to use any other advanced technique such as ex vivo models. A new three-dimensional ex vivo mouse model is used by Dr. Alastair Sloan, Cardiff University, to study inflammation and bone metabolism in gingival and periodontal diseases. This three-dimensional model uses a slice culture of the mouse mandible to study the progression of gingival and periodontal disease pathology and to investigate bone repair at cellular and molecular level. Hence, there will be 80–96% reduction of animal usage with the new three-dimensional ex vivo mouse model in the research studies of periodontal diseases.[77]

 Future Perspectives

As the basics of biology have reached the molecular level and various phenomena of life are now dependent on genes, the studies done on these animal models are referred as biomedical research. For example, animal models for human diseases are improved to the present level through the integrated application of genetic, cellular engineering and embryonic manipulation principles according to the requirements or objectives of the study. Following are the objectives to follow during the usage of animal models.[78],[79]

Replace the animal models in nonanimal-based experimental researches The number of animals used must be adjusted to the minimum by following in vitro methods or using ex vivo models Animal research is conducted once all the requirements are followed which are provided by regulatory bodies Animal models must be constantly improved to the current level of advancements in biomedical research, and the studies should be more reliable and informative Protection of animals required for research is a permanent consideration

According to this review, it could be stated that all the scientific researchers should follow the ethical guidelines while selecting the animal models for their research or training purpose in educational institutions. This review not only provides information regarding the specific uses of each animal model in various specialties of dentistry but also emphasizes on how to reduce, refine, and replace the animal models in future research purpose.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.


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