Humane endpoints
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A broad definition of Humane endpoint is a concept for continuous refinement of animal studies (Hendriksen et al., 2010).
This broad definition is aligned with the 3R principles (Balls et al., 1995, Russell and Burch, 1959).
Continuous improvements or refinement is embedded in the directive 2010/63 (European Commission, 2010)
- Article 4.3 Member States shall ensure refinement of breeding, accommodation and care, and of methods used in procedures, eliminating or reducing to the minimum any possible pain, suffering, distress or lasting harm to the animals.
Refinement of study specific endpoints and alternatives to death/moribund as endpoint
| Focus of the Study | Proposed Endpoint | Reference | |
| INFECTIONS, IMMUNE SYSTEM | |||
| Infectious diseases | Acute phase responses | Olfert, E.D. and D.L. Godson, Humane endpoints for infectious disease animal models. ILAR J, 2000. 41(2): p. 99-104. | |
| Body Temperature | Olfert, E.D. and D.L. Godson, Humane endpoints for infectious disease animal models. ILAR J, 2000. 41(2): p. 99-104. | ||
| Weight loss | Olfert, E.D. and D.L. Godson, Humane endpoints for infectious disease animal models. ILAR J, 2000. 41(2): p. 99-104. | ||
| Shock by cecal ligations and puncture (CLP) | Observation of Loss of ability to ambulate.
Body weight and temperature |
Nemzek, J. A., Xiao, H. Y., Minard, A. E., BOLGOS, G. L. & REMICK, D. G. 2004. Humane endpoints in shock research. Shock, 21, 17-25. | |
| Rheumatoid arthritis in rodents | Swollen digits, knuckle, midfoot and ancle/wrist scoring, Ulceration, gait and posture analysis | Hawkins, P., et al., Applying refinement to the use of mice and rats in rheumatoid arthritis research. Inflammopharmacology, 2015. 23(4): p. 131-50. | |
| Tuberculosis vaccine studies | Bodyweight AND Breathing pattern AND activity during handling combined predict need for euthanasia 1 week ahead | Collymore, C., Kent, et al. 2018. Humane Endpoints for Guinea Pigs Used for Mycobacterium tuberculosis Vaccine Research. Comparative Medicine, 68, 41-47 | |
| Inoculation with infectious organisms | Temperature AND Body weight | Trammell, R. A. & Toth, L. A. 2011. Markers for predicting death as an outcome for mice used in infectious disease research. Comp Med, 61, 492-8. | |
| Vaccine potency testing | Reduced temperature AND bodyweight | Hendriksen. (2011). Humane endpoints in vaccine potency testing. Procedia in Vaccinology, 5, 221–226. https://doi.org/10.1016/j.provac.2011.10.022 | |
| CANCER | |||
| Early Cancer detection | Ultrasound | Wang, L. V. (2003). Ultrasound-mediated biophotonic imaging: a review of acousto-optical tomography and photo-acoustic tomography. Disease Markers, 19(2-3), 123–138. https://doi.org/10.1155/2004/478079 | |
| Intracranial Glioma in rat | Body weight algorithm | Helgers, S.O.A., et al., Body weight algorithm predicts humane endpoint in an intracranial rat glioma model. Sci Rep, 2020. 10(1): p. 9020. | |
| Severe irradiations studies in NHP | Rapid decrease of body temperature for 3 consecutive days combined with general behaviour score | Bertho, J.M., et al., Humane endpoints in severe irradiation experiments using non-human primates: a retrospective analysis. Scandinavian Journal of Laboratory Animal Science, 2020. 46(1): p. 1-15 | |
| Total-Body Irradiation in Mouse | Body weight, temperature by telemetry | Koch, A., et al., Establishment of Early Endpoints in Mouse Total-Body Irradiation Model. PLoS One, 2016. 11(8): p. e0161079. | |
| Colon cancer in rats | black feces or presence of fresh blood
|
Kobaek-Larsen, Morten, Fenger, Claus, & Ritskes-Hoitinga, Jelmera. (2004). Secondary effects induced by the colon carcinogen azoxymethane in BDIX rats. APMIS : Acta Pathologica, Microbiologica et Immunologica Scandinavica, 112(6), 319–329. https://doi.org/10.1111/j.1600-0463.2004.apm1120601.x | |
| Experimental liver metastases | Laparoscopy | Kobaek-Larsen, M., et al., Laparoscopy of rats with experimental liver metastases: a method to assess new humane endpoints. Lab Anim, 2004. 38(2): p. 162-8. | |
| Lung Cancer | Iincreases in respiratory rate | Winn, Hwang, S.-K., Morin, J., Bluette, C. T., Manickam, B., Jiang, Z. K., Giddabasappa, A., Liu, C.-N., & Matthews, K. (2021). Automated monitoring of respiratory rate as a novel humane endpoint: A refinement in mouse metastatic lung cancer models. PloS One, 16(9), e0257694. https://doi.org/10.1371/journal.pone.0257694 | |
| Urinary Bladder Carcinogenesis | Hematuria, Reduced hematocrit, lower final body weight, increase mean drink intake, increased urinary output | Oliveira, Nascimento-Gonçalves, E., Silva, J., Oliveira, P. A., Ferreira, R., Antunes, L., Arantes-Rodrigues, R., & Faustino-Rocha, A. I. (2017). Implementation of Humane Endpoints in a Urinary Bladder Carcinogenesis Study in Rats. In Vivo (Athens), 31(6), 1073–1080. | |
| CNS | |||
| Amyotrophic Lateral Sclerosis (ALS) | Decline in motoric Function | Mead, R.J., et al., Optimised and rapid pre-clinical screening in the SOD1(G93A) transgenic mouse model of amyotrophic lateral sclerosis (ALS). PLoS One, 2011. 6(8): p. e23244. | |
| Home cage running wheel | Bennett, E.J., et al., Early detection of motor dysfunction in the SOD1G93A mouse model of Amyotrophic Lateral Sclerosis (ALS) using home cage running wheels. PLoS One, 2014. 9(9): p. e107918. | ||
| Stroke | Weight change on the 1st and 3rd day after treatment.
Core temperature change on the 3rd day after treatment (male, stroke model), or with Neuroscore, weight change, and core temperature change on the 2nd day after treatment (female, stroke model) by machine learning. |
Mei, J., Banneke, S., et al. Refining humane endpoints in mouse models of disease by systematic review and machine learning-based endpoint definition. ALTEX, 36, 555-571. | |
| TOX testing | |||
| Alternatives to LD 50 | Hypotermiadose50 (HD50) replaces LD50 | Soothill, J. S., Morton, D. B. & Ahmad, A. 1992. The HID50 (hypothermia-inducing dose 50): an alternative to the LD50 for measurement of bacterial virulence. Int J Exp Pathol, 73, 95-8. | |
| SURGERY | |||
| Post laparotomy | Telemetric recording of heart rate | Arras, M., et al., Assessment of post-laparotomy pain in laboratory mice by telemetric recording of heart rate and heart rate variability. BMC Vet Res, 2007. 3: p. 16.
| |
| OTHER ORGAN SYSTEMS | |||
| Murine model for cholestasis | Burrowing activity | Zhang, X., et al., A rational approach of early humane endpoint determination in a murine model for cholestasis. ALTEX, 2020. 37(2): p. 197-207. | |
| Anti-urolithiasis activity of test compounds by zinc disc implantation | Imaging (X-ray radiographs) of the bladder deposits | Singh, P.K., et al., Zinc disc implantation model of urinary bladder calculi and humane endpoints. Lab Anim, 2010. 44(3): p. 226-30. | |
| Ocular Herpes virus Infection | Combined Body weight reduction (>0.05 g/day) AND Body Temperature less than 34,5°C measured by s.c implanted temperature probe. - Predicted death in 98% of mice. | Hankenson, F. C., Ruskoski et al , 2013. Weight Loss and Reduced Body Temperature Determine Humane Endpoints in a Mouse Model of Ocular Herpesvirus Infection. Journal of the American Association for Laboratory Animal Science, 52, 277-285. | |
| Non-alcoholic fatty liver | Computed tomography in | Lintrup K, Ipsen DH, Skat-Rørdam J, Lykkesfeldt J, Tveden-Nyborg P, Buelund LE. Validation of computed tomography as a diagnostic tool in guinea pigs with non-alcoholic fatty liver disease. Lab Anim. 2023 Nov 24:236772231182511. doi: 10.1177/00236772231182511. Epub ahead of print. PMID: 37999627. | |
| AGING STUDIES | |||
| Identify objective criteria that predict death in aged mice | Combined Body weight reduction (>0.05 g/day) AND Body Temperature | Trammell, Cox, L., & Toth, L. A. (2012). Markers for Heightened Monitoring, Imminent Death, and Euthanasia in Aged Inbred Mice. Comparative Medicine, 62(3), 172–178. | |
| Improve care, husbandry and management of aging mice | Enhanced clinical monitoring of age related parameters | Wilkinson, Selman, C., McLaughlin, L., Horan, L., Hamilton, L., Gilbert, C., Chadwick, C., & Flynn, J. N. (2019). Progressing the care, husbandry and management of ageing mice used in scientific studies. Laboratory Animals (London), 54(3), 23677219865291–238. https://doi.org/10.1177/0023677219865291 | |
| Literature surveys | |||
| Lung injury | Temperature AND Body weight | McGinn, Fergusson, D. A., Stewart, D. J., Kristof, A. S., Barron, C. C., Thebaud, B., McIntyre, L., Stacey, D., Liepmann, M., Dodelet-Devillers, A., Zhang, H., Renlund, R., Lilley, E., Downey, G. P., Brown, E. G., Côté, L., Dos Santos, C. C., Fox-Robichaud, A. E., Hussain, S. N. A., … Lalu, M. M. (2021). Surrogate Humane Endpoints in Small Animal Models of Acute Lung Injury: A Modified Delphi Consensus Study of Researchers and Laboratory Animal Veterinarians. Critical Care Medicine, 49(2), 311–323. https://doi.org/10.1097/CCM.0000000000004734 | |
| Acute lung injury | Temperature AND Body weight | Zingarelli. (2021). First Do No Harm: A Proposal of an Expert-Guided Framework of Surrogate Humane Endpoints in Preclinical Models of Acute Lung Injury. Critical Care Medicine, 49(2), 373–375. https://doi.org/10.1097/CCM.0000000000004758 | |
OTHER OBSERVATION STRATEGIES WITH POTENTIAL TO BE USED FOR DEFINING ENDPOINTS
| In-Tank Underwater Cameras can Refine Monitoring of Laboratory Fish | Ellis, T; Rimmer, GSE; Parker, S-J; Joiner, Claire; Sebire, Marion, Verner-Jeffreys, David, Lines, Jeffrey
2019/05/01, 191, 203, In-Tank Underwater Cameras can Refine Monitoring of Laboratory Fish DO - 10.7120/09627286.28.2.191, Animal Welfare |
|---|---|
Many or few endpoint criteria
Some authors stress that endpoints can rarely be generalized. Using too many and too general endpoints for scoring is labor intensive. Researchers might be required to evaluate variables that might appear arbitrary or unrelated to the animal’s condition. Compliance may be increased if the measurement of the variables is related to research objectives. Other authors underline that humane endpoint cannot rely on a single variable, suggesting to use a combination of variables and recommend a combination of assessment parameters for a robust welfare assessment in laboratory animals and early predictor of death. For example, the combined reduction of body weight and body temperature, has shown to be a reliable predictor of death in studies including:
- Nemzek JA, Xiao HY, Minard AE, Bolgos GL, Remick DG. Humane endpoints in shock research. Shock. 2004;21(1):17-25.
- McGinn R, Fergusson DA, Stewart DJ, Kristof AS, Barron CC, Thebaud B, et al. Surrogate Humane Endpoints in Small Animal Models of Acute Lung Injury: A Modified Delphi Consensus Study of Researchers and Laboratory Animal Veterinarians. Crit Care Med. 2021;49(2):311-23.
- Zingarelli B. First Do No Harm: A Proposal of an Expert-Guided Framework of Surrogate Humane Endpoints in Preclinical Models of Acute Lung Injury. Crit Care Med. 2021;49(2):373-5.
- Trammell RA, Toth LA. Markers for predicting death as an outcome for mice used in infectious disease research. Comp Med. 2011;61(6):492-8.
- Hendriksen C. Humane endpoints in vaccine potency testing. Procedia in Vaccinology 5 (2011) 2011;5:221 – 6.
- Hankenson FC, Ruskoski N, van Saun M, Ying GS, Oh J, Fraser NW. Weight Loss and Reduced Body Temperature Determine Humane Endpoints in a Mouse Model of Ocular Herpesvirus Infection. J Am Assoc Lab Anim. 2013;52(3):277-85.
- Trammell, Cox, L., & Toth, L. A. (2012). Markers for Heightened Monitoring, Imminent Death, and Euthanasia in Aged Inbred Mice. Comparative Medicine, 62(3), 172–178.
