Neonatal female donkey foal (Equus africanus asinus).A middle-aged donkey jenny was presented to the University of Melbourne for imminent parturition. The sire of the foal was unknown. Foaling was routine and the foal was bright and vigorous, but approximately one hour post-parturition the foal suffered a fracture of the right tibia during an attempt to stand. Radiographs indicated a displaced spiral fracture of the distal tibial diaphysis and metaphysis. The tibia and femur also had radiographic evidence of reduced medullary cavity size, thickened mid-diaphyseal cortices, and conical metaphyseal bone extending toward the mid-diaphysis. Due to the suspicion of underlying bone disease, the foal was euthanized with owner consent.

Gross Description:  

The animal displayed marked brachygnathism inferior and failure of dental eruption. All bones were brittle and easily cut with a knife. There was an acute spiral fracture extending along the left tibia, as well as a nondisplaced fracture of the right transverse process of the L2 vertebrae. Focal bone thickenings (callouses) and hemorrhages were present over the midpoint of the ribs bilaterally from T6-T13, and the orientation of the ribs was deviated. On sectioning, all long bones displayed thickened cortices and large cores of intramedullary trabecular bone that extended along the diaphysis from both the proximal and distal metaphysis, with severe reduction in medullary space.

Histopathologic Description:

Long bone, metaphysis: Extending from the growth plate through the medullary cavity of the metaphysis there is a marked increase in the extent of metaphyseal trabeculae. Trabeculae are disorganized and are composed of irregular cores of retained cartilage derived from the hypertrophic zone of the physis, overlayed and interspersed by mineralized bone. Towards the diaphysis there is a progressive decrease in trabecular size and number, and intertrabecular medullary spaces contain foci of hematopoietic cells within loose fibroadipose connective tissue. The cortex is markedly thickened and poorly compacted, with cortical bone arranged in longitudinal lamellae separated by loose fibrovascular and adipose tissue with occasional sparse bone marrow infiltration. Osteoclast numbers throughout the section are markedly decreased, and there is no evidence of reversal lines or Howship's lacunae within the section.

Morphologic Diagnosis:  

Bone: Osteopetrosis, diffuse, donkey (Equus africanus asinus).



Contributor Comment:  

Osteopetrosis (AKA marble bone disease) is a heterogeneous disease process characterized by defective osteoclastic bone resorption, resulting in failure of bone remodeling and dense, fragile bones that fracture readily. The condition has been reported in a number of species, including Angus, Hereford and Simmental cattle, Peruvian Paso and Appaloosa horses, white-tailed deer, rabbits and several dog breeds.(12) To the author's knowledge, this is the first case observed in a donkey. Brachygnathia inferior is a common finding in affected animals, reflecting failure of mandibular growth, and dental eruption is typically impaired, as this process requires bone resorption. Anemia has also been reported in some cases, as well as leukopenia and immune deficiencies such as hypoglobulinaemia.(1,6) Anemic animals often display hepatosplenomegaly due to prominent extramedullary hematopoiesis. Hemato-logical assessment was not performed in the present case, but the presence of abundant hematopoietic tissue within the medullary cavity, as well as a lack of extramedullary hematopoiesis in the liver and spleen, suggest that marrow function was adequate. In humans, failure of bone remodeling has also been associated with nerve entrapment and compression, most commonly manifesting as blindness and deafness, though these feature have not been noted in domestic species. Osteopetrosis occurs in various forms, but the disease typically reflects either functional osteoclast impairment (such as defects in carbonic anhydrase or the H+-ATPase proton pump)(13) or depletion of the osteoclast cell population. Many cases in domestic species are suspected to represent genetic defects with autosomal recessive heritability, but the specific mutation has so far only been identified in Angus cattle, which display a deletion mutation of the SLC4A2 anion exchanger.(8) The SLC42A defect results in failure of acidification at the sites of remodeling and defective bone demineralization. Osteoclasts are rare in the Angus form of disease, but animals with functional osteoclast defects typically have adequate or increased osteoclast numbers, though the osteoclasts may display morphological abnormalities such as hypertrophy, increased numbers of nuclei or absence of a ruffled border.

Osteopetrosis caused by failure of osteoclast development is rare, but osteoclast-poor disease with autosomal recessive heritability exists in humans. Osteoclast differentiation is dependent on RANK/RANKL (Receptor Activator of Nuclear Factor-KB/ Receptor Activator of Nuclear Factor-KB Ligand) signaling, and mutations in genes responsible for this receptor/ligand pair ( TNFRSF11A and TNFRSF11, respectively) lead to the profound osteoclast deficiency.(11,5) In mouse models, osteopetrosis is also observed with defects in a range of other genes associated with osteoclast differentiation, but these have not been described in natural disease.(3) The depletion of osteoclasts in the present case suggests it may be a different form of osteopetrosis to that described in most other equine cases, which typically display normal to increased osteoclast numbers. However, a single case of osteoclast-poor osteopetrosis has been reported in a Peruvian Paso foal.(8)

Acquired osteoporosis like disease has been reported secondary to a number of viral infections, presumably reflecting viral osteoclast tropism and cell depletion. Zonal lesions consistent with osteopetrosis have been reported with bovine viral diarrhoea virus in cattle,(9) and similar features have been identified in the metaphyseal region of dogs with distemper and cats infected with feline leukaemia virus.(12) Osteosclerotic disease similar to osteopetrosis has also been reported associated with hypervitaminosis,(6) lead poisoning,(12) and exogenous oestrogen(5) and glucocorticoid administration.(4) Cases of rickets may display retention of cartilaginous metaphyseal trabeculae overlayed by osteoid, similar to those observed in osteopetrosis. In rickets, however, the trabeculae are poorly mineralized, while the bone in cases of osteopetrosis undergoes normal mineralization.

JPC Diagnosis:  

Long bone: Physeal dysplasia, diffuse, severe with osteoclast depletion, failure of chondroclasis, cortical osteopenia and diffuse medullary osteosclerosis (osteopetrosis).

Conference Comment:  

The histologic description discussed in conference was very similar to the contributors. The noted primary features included increased bony trabeculae arranged in disorganized transverse and longitudinal arrays with Tibial physis, donkey. There is marked persistence of the physeal zone of hypertrophy. Cartilage cores bridge in a horizontal fashion (arrows) as well as laterally, and there are large areas of degenerate cartilage with minimal osteoid deposition or mineralization. (HE, 35X) persistent cartilage cores, absence of osteoclasts, decreased bone marrow elements, absence of a cutback zone and cortical osteopenia. The physeal zones involved in endochondral ossification appear histologically normal until subjacent to the zone of hypertrophy, where osteoclasts would normally be present remodeling mineralized cartilage, but in this case are absent. Cortical osteopenia, including multiple thin, widely separated longitudinal bands of bone, was also described and discussed as a feature not commonly associated with osteopetrosis, and there was speculation it may reflect a secondary process. The gross images were viewed during conference, and there was discussion regarding the multiple callouses and hemorrhages on the ribs being secondary to in utero fracture.

There are two main forms of osteopetrosis described in humans: a recessively inherited lethal form in which lesions are present at birth, and a dominant form which manifests in adults.(2) Osteopetrosis in Red Angus calves with a deletion mutation in the gene SLC4A2, which encodes the anion exchanger for carbonate and chloride, has recently been compared to the recessive form which occurs in children, and aspects of the craniofacial lesions were found to be similar. Craniofacial lesions in affected calves include dorsoventrally compressed brains with depressions of the parietal cortex due to thickening of the parietal bone; compression of the cerebral hemispheres with vermis herniation through the foramen magnum; chromatolysis in multiple cranial nerve nuclei; optic nerve atrophy; loss of retinal ganglion cells; and dysplastic changes in the molar and premolar teeth. There was also corpora amylacea in the thalamus, basal nuclei and midbrain, and mineralization in vessels of the thalamus. One important difference noted between the condition in Red Angus calves and the recessive lethal form in humans is the presence of increased numbers of osteoclasts in people, while decreased numbers of osteoclasts were present in the long bones of affected calves. Osteoclast numbers in calves, however, were more normal in bones of the head. Affected calves died in utero or shortly after birth and were either homozygous or heterozygous for the SLC4A2 mutation, respectively.(9) Long bone lesions in affected calves included dense unresorbed bony trabeculae from the metaphysis to the central diaphysis, similar to what is present in other species.(2)

The specific type of osteoclast defect present influences the number of osteoclasts, whether increased, decreased, or absent, in various types of osteopetrosis. For example, in mutations involving the chloride channel (ClCN7) or the proton pump (ATP6i), osteoclasts may be present in increased numbers but are nonfunctional; however, in mutations involving the RANKL gene, necessary for proper osteoclast differentiation, osteoclasts are decreased or absent. Osteopetrosis in Hereford and Simmental breeds of cattle is similar to the condition in Angus calves. One difference is the presence of thickened frontal bones with cystic spaces, creating a domed forehead that can be mistaken for hydrocephalus. Osteopetrosis is described in Belgian Blue cattle in Europe in combination with abnormal skull formation and mandibular gingival hamartomas. A mutation was identified in the chloride / proton exchanger lysosomal anion transporter ClCN7 in affected calves. Osteopetrosis is also described in inbred Polypay sheep and white-tailed deer, both of which also have brachygnathia inferior, a common finding in many affected species. The white-tailed deer also have calluses on several ribs, similar to the disease in horses, suggesting in utero rib fractures. Osteopetrosis has been documented in dogs but is poorly characterized.(2)


1. Berry CR, House JK, Poulos PP, et al. Radiographic and pathologic features of osteopetrosis in two Peruvian Paso foals. Veterinary Radiology & Ultrasound. 1994;35:355-361.

2. Craig LE, Dittmer KE, Thompson KG. Bones and Joints. In: Maxie MG, ed. Jubb, Kennedy, and Palmer's Pathology of Domestic Animals. 6th ed. Vol1. St. Louis, MO: Elsevier; 2015:49-53.

3. Del Fattore A, Cappariello A, Teti A. Genetics, pathogenesis and complications of osteopetrosis. Bone. 2008;42:19-29.

4. Glade MJ, Krock L. Glucocorticoid-lnduced inhibition of osteolysis and the development of osteopetrosis, osteonecrosis and osteoporosis. Cornell Vet. 1982;72:76-91.

5. Guerrini MM, Sobacchi C, Cassani B, et al. Human osteoclast-poor osteopetrosis with hypogammaglobulinemia due to TNFRSF11A (RANK) mutations. Am J Hum Genet. 2008;83:64-76.

6. Kramers P, Fluckiger MA, Rahn BA, et al. Osteopetrosis in Cats. Journal of Small Animal Practice. 1988;29:153-164.

7. Meyers SN, McDaneld TG, Swist SL, et al. A deletion mutation in bovine SLC4A2 is associated with osteopetrosis in Red Angus cattle. BMC Genomics. 2010;11:337.

8. Nation PN, Klavano GG. Osteopetrosis in two foals. Can Vet J. 1986;27:74-7.

9. OToole D, Swist S, Steadman L, Johnson GC. Neuropathology and craniofacial lesions of osteopetrotic Red Angus calves. Vet Pathol. 2012;49(5):746-754.

10. Scruggs OW, Fleming SA, Maslin WR, et al. Osteopetrosis, anemia, thrombocytopenia, and marrow necrosis in beef calves naturally infected with bovine virus diarrhea virus. J Vet Diagn Invest. 1995;7:555-9.

11. Sobacchi C, Frattini A, Guerrini MM, et al. Osteoclast-poor human osteopetrosis due to mutations in the gene encoding RANKL. Nat Genet. 2007;39:960-2.

12. Thompson K. Diseases of Bones and Joints. In: Maxie MG, ed. Jubb, Kennedy and Palmer 's Pathology of Domestic Animals. 5th ed. Vol1. Philadelphia, PA: Elsevier Ltd; 2007:24-40, 75-80.

13. Tolar J, Teitelbaum SL, Orchard PJ. Osteopetrosis. N Engl J Med. 2004;351:2839-49.

Click the slide to view.

2-1. Mandible

2-2. Mandible

2-3. Tibia

2-4. Ribs

2-5. Long bones

2-6. Tibia

2-7. Tibia physis

2-8. Tibial diaphysis

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