Pathology in Practice

Caterina Maestrale Istituto Zooprofilattico Sperimentale della Sardegna G. Pegreffi, 07100 Sassari, Italy.

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Mariangela Masia Istituto Zooprofilattico Sperimentale della Sardegna G. Pegreffi, 07100 Sassari, Italy.

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Giuseppe Marruchella Department of Comparative Biomedical Sciences, Faculty of Veterinary Medicine, University of Teramo, 64100 Teramo, Italy.

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Giovanni Di Guardo Department of Comparative Biomedical Sciences, Faculty of Veterinary Medicine, University of Teramo, 64100 Teramo, Italy.

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Alberto Mancosu Regional Breeders' Association, Viale Porto Torres 32, 07100 Sassari, Italy.

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Maria G. Cancedda Istituto Zooprofilattico Sperimentale della Sardegna G. Pegreffi, 07100 Sassari, Italy.

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Ciriaco Ligios Istituto Zooprofilattico Sperimentale della Sardegna G. Pegreffi, 07100 Sassari, Italy.

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History

During 3 consecutive breeding seasons, a number of lambs had died shortly after birth in a flock of Sarda sheep in Sardinia. Affected lambs had hairless areas of skin on the limbs and pinnae; the hooves were absent in 1 lamb. The flock included approximately 300 ewes and 4 rams, and records indicated that 10 lambs were affected with this congenital skin disorder within the preceding year. Flock feeding relied on the exploitation of natural pastures; the diet was supplemented with concentrate and hay during the milking period. The owner reported that no new introduction of sheep had occurred before the onset of the first clinical case. A dead female lamb was transported to the diagnostic laboratories of the Istituto Zooprofilattico Sperimentale della Sardegna for postmortem examination and subsequent histologic and immunohistochemical (IHC) investigations.

Clinical and Gross Findings

The lamb had been found dead at the farm 2 hours after birth. Macroscopically, there were symmetric, reddish, translucent, and well-demarcated alopecic skin areas on the pinnae and the more distal portions of the forelimbs (Figure 1). No additional lesions involving other organs were identified during necropsy. On the basis of morphologic characteristics, the fetal development of the lamb appeared normal.

Figure 1—
Figure 1—

Photograph of a Sarda breed lamb with bilateral hairless, reddish skin lesions on the pinnae and lower regions of the forelimbs. The lamb had been found dead at the farm 2 hours after birth. In the farm's flock, a number of lambs had died shortly after birth during 3 consecutive breeding seasons.

Citation: Journal of the American Veterinary Medical Association 242, 2; 10.2460/javma.242.2.179

Formulate differential diagnoses from the history, clinical findings, and Figure 1then turn the page

Histopathologic Findings

Samples of both macroscopically affected and apparently normal skin were collected from several body regions of the lamb. Additional tissue samples of major organs such as heart, lungs, liver, kidneys, and brain were also collected. These samples were subsequently processed for histologic examination by means of paraffin embedding, microtome sectioning at a thickness of 5 μm, and final staining with H&E or periodic acid-Schiff stain. Finally, DNA was isolated from liver tissue for genetic analyses of the LAMA3 gene, which encodes for 2 laminin subunits.

Microscopically, the damaged skin areas lacked epidermis, including basement membrane, as well as adnexal structures (hair follicles, sebaceous glands, and sweat glands). At the junction between unaffected skin and affected tissue, the epidermis ended abruptly, with its margins remaining strongly adherent to (Figure 2) or, rarely, detached from the underlying dermis. The exposed dermis had variably extensive and superficial necrotic foci, with a prominent inflammatory infiltration by neutrophils and lymphocytes. In the skin adjacent to the damaged areas as well as in other body regions, the epidermis was morphologically normal and adherent to the dermis, even after injury was inflicted to test the resistance. Bullae or dermoepidermal clefting was not evident in any cutaneous area.

Figure 2—
Figure 2—

Photomicrographs of sections of skin of the Sarda breed lamb in Figure 1 that underwent H&E staining (A, B, and C) for histologic examination or immunohistochemical staining (D, E, and F) to detect cytokeratin (by use of a routine biotin-streptavidin method, which included monoclonal primary antibody against antigenic determinants of most of the subfamily A and B cytokeratins, use of 3,3'-diaminobenzidine tetrahydrochloride as the chromogen, and Meyer hematoxylin counterstaining). A—Section of an auricle at the junction between unaffected and affected skin. Unaffected skin ends abruptly (arrow), with the epidermis remaining strongly adherent to the underlying dermal component. B— Another view of the section of skin from the auricle in panel A at the junction between unaffected and affected areas. The terminal portion of the epidermis appears to be detached from the underlying dermal component (arrow). C—Section of skin obtained from an affected forelimb. All epidermal layers are missing, and an abundant inflammatory infiltrate composed of neutrophils and lymphocytes is present throughout the dermis. D—Section of skin at the junction of unaffected and affected skin of an auricle. The cytokeratin immunolabeling is restricted to normal skin. E—Section of skin obtained from an affected forelimb. No cytokeratin immunolabeling is evident demonstrating the absence of keratinocytes from affected skin areas. F—Section of skin obtained from an unaffected hind limb. Keratinocytes in apparently normal skin are strongly and uniformly labeled for cytokeratin. Bar = 100 m (applies to all panels).

Citation: Journal of the American Veterinary Medical Association 242, 2; 10.2460/javma.242.2.179

These skin lesions were further characterized by means of IHC staining to detect cytokeratin. The IHC staining was performed with a routine biotin-streptavidin method, which included a mouse monoclonal primary antibodya (final dilution, 1:50) against antigenic determinants of most cytokeratins in subfamilies A and B (which are both expressed in normal ovine skin), use of 3,39'-diaminobenzidine tetrahydrochloride as the chromogen, and Meyer hematoxylin counterstaining. Results of the IHC analysis confirmed the complete absence of epidermis and related adnexal structures from all affected skin areas (Figure 2). In contrast, all regions of unaffected skin had an IHC staining pattern expected for normal skin. The preliminary data from the genetic investigations indicated that there were no point mutations or deletions in the LAMA3 gene that could be considered to be associated with the dermal changes in the lamb.

Morphologic Diagnosis and Case Summary

Morphologic diagnosis: congenital, focal aplasia of the epidermis and adnexal structures, associated with superficial necrosis and extensive dermal inflammation.

Case summary: epitheliogenesis imperfecta (EI) in a lamb.

Comments

Sarda is the sole breed comprising the large sheep population of Sardinia, Italy. In this breed of medium-sized dairy sheep, the mean body weight of ewes is 50 kg (110 lb) and that of rams is 70 kg (154 lb). The lambing period extends from November to February.

On the basis of history, clinical signs, histopathologic changes, and IHC analysis results, a final diagnosis of EI was made for the lamb of this report. Epitheliogenesis imperfecta is a rare congenital pathological condition, which has been described in several animal species, such as cattle, swine, sheep, and horses1 as well as in humans2,3; in humans, the condition is known as aplasia cutis. Moderately to severely affected domestic animals usually die within the first days after birth as a result of septicemia or concomitant anomalies.1 Epitheliogenesis imperfecta–associated lesions are characterized by the absence of squamous epithelia in variably sized areas of the skin and mucous membranes. This defect has been reported for lambs from Swiss4 and Australian breeds5; in those affected lambs, the absence of the epidermis was microscopically evident in some cutaneous areas as well as in the oral mucosa. Malformations of the hooves, nails, and teeth accompanied the histopathologic changes. In the case described in the present report, the lack of lesions in both mucous membranes and teeth illustrates the fact that the extension and distribution of skin lesions may vary among animals with EI. However, given that the etiopathogenesis of EI is currently unknown, the cause of the variability in EI-associated pathological changes remains to be elucidated. Despite the absence of lesions in mucous membranes and teeth, the microscopic findings for the lamb of this report largely overlapped those reported for other EI-affected individuals.1,4–6

A detailed histopathologic description of 70 cases of EI in piglets has been published.7 Of the 70 affected piglets, 65 (93%) were males, suggesting that EI could be a sex-related condition in swine. Nevertheless, currently available data for sheep are scarce and do not support any influence of sex on EI occurrence in lambs. Additionally, in the swine study,7 the morphologic features of the hair follicles at the junction of normal and affected skin were variable, allowing definition of different histopathologic patterns, namely areas in which follicles were apparently normal, multifocally increased in number or size, or dysplastic. Although similar changes have not been categorized in lambs because of the low number of cases available for investigation, the histopathologic pattern in the affected lamb of the present report can be considered normal on the basis of the pattern definitions for affected swine.

Interestingly, a partial absence of dermis was further observed in piglets7; thus, aplasia cutis (the term used in human medicine) may be a more appropriate definition of this condition. In this respect, it should be also underscored that EI, aplasia cutis, and epidermolysis bullosa (a neonatal genodermatosis also affecting lambs1,8,9) have been commonly used as synonyms, thereby generating some confusion. In the lamb of the present report, considering that the congenital disorder affected only the epidermis, it may be correctly defined as EI.1

Absence of adnexal structures that is restricted to the diseased skin regions is another interesting morphologic hallmark of EI. On the basis of histologic observations in the case described in this report, we believe that the absence of the adnexal structures was not due to an inflammatory process affecting the damaged skin; conversely, the absence of the adnexal structures may have represented an expected consequence of deficient epithelial development and morphogenesis during embryogenesis. Indeed, results of a study10 of mice homozygous for a disrupted p63 gene (a homologue of the tumor suppressor p53) indicated that congenital defects of squamosus epithelia development in the skin may be associated with absence of the epithelial appendages.

Infection, vascular malformations, defective amniogenesis, and various teratogens have been proposed as causative factors of aplasia cutis in humans, but no unifying theory has been identified.3 Epitheliogenesis imperfecta probably represents a simple autosomal recessive trait in cattle, horses, swine, and sheep.11

The occurrence of EI in the flock of the present report was not associated with the introduction of new animals. In this flock, however, these data were not sufficient to rule out the presence of a genetic causative factor. Further investigations are needed to better characterize the possible association of this birth defect with 1 or more unknown genetic factors. Indeed, EI appears to be a recessively inherited trait in piglets.7

Epidermolysis bullosa is often misdiagnosed as EI. Epidermolysis bullosa includes a group of hereditary mechanobullous diseases characterized by cutaneous blisters in response to trauma. This disease in sheep has been repeatedly reported, and definitive diagnosis is made on the basis of history and physical examination and histopathologic findings.8,9 The histopathologic findings are characterized by subepidermal vesicles, resulting from dermoepidermal separation below the basement membrane, that are present at birth or develop within a few days or weeks after birth.1

Furthermore, epidermolysis bullosa has been associated, both in horses and sheep, with a partial deletion of LAMA3 and LAMC212,13 genes that encode for 2 laminin subunits. Laminin is one of the most important components of the basement membrane of the epidermis and acts as a pathway for the migration, differentiation, and organization of cutaneus epithelial cells. Nevertheless, results of LAMA3 sequencing for the lamb of the present report did not suggest an involvement of this gene in the pathogenesis of EI.

a.

Clone AE1/AE3, Dako, Carpinteria, Calif.

References

  • 1. Scott DW. Congenital and hereditary diseases. In:Scott DW, ed. Large animal dermatology. Philadelphia: WB Saunders Co, 1988;334357.

    • Search Google Scholar
    • Export Citation
  • 2. Moros Peña M, Labay MM, Valle Sánchez F, et al. Aplasia cutis congenita in a newborn: etiopathogenic review and diagnostic approach [in Spanish]. An Esp Pediatr 2000; 52:453456.

    • Search Google Scholar
    • Export Citation
  • 3. Fagan LL, Harris PA, Coran AG, et al. Sporadic aplasia cutis congenita. Pediatr Surg Int 2002; 18:545547.

  • 4. Tontis A, Hofstetter H. Epitheliogenesis imperfecta in lambs. Schweiz Arch Tierheilkd 1991; 133:287289.

  • 5. Munday BL. Epitheliogenesis imperfecta in lambs and kittens. Br Vet J 1970; 126:47.

  • 6. Saperstein G, Leipold HV, Dennis SM. Congenital defects of sheep. J Am Vet Med Assoc 1975; 167:314322.

  • 7. Benoit-Biancamano MO, Drolet R, D'Allaire S. Aplasia cutis congenita (epitheliogenesis imperfecta) in swine: observations from a large breeding herd. J Vet Diagn Invest 2006; 18:573579.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 8. Alley MR, O'Hara PJ, Middelberg A. An epidermolysis bullosa of sheep. N Z Vet J 1974; 22:5559.

  • 9. McTaggart HS, Ritchie JS, Copland AN. Red foot disease of lambs. Vet Rec 1974; 94:153159.

  • 10. Yang A, Schweitzer R, Sun D, et al. p63 is essential for regenerative proliferation in limb, craniofacial and epithelial development. Nature 1999; 398:714718.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 11. Ginn PE, Mansell JEKL, Rakich PM. Skin and appendages. In:Maxie MG, ed. Jubb, Kennedy and Palmer's pathology of domestic animals. 5th ed. Philadelphia: Elsevier Saunders, 2007;575576.

    • Search Google Scholar
    • Export Citation
  • 12. Graves KT, Henney PJ, Ennis RB. Partial deletion of LAMA3 gene is responsible for hereditary junctional epidermolysis bullosa in the American Saddlebred Horse. Anim Genet 2008; 40:3541.

    • Search Google Scholar
    • Export Citation
  • 13. Mömke S, Kerkmann A, Wöhlke A, et al. A frameshift mutation within LAMC2 is responsible for Herlitz type junctional epidermiolysis bullosa (HJEB) in black headed mutton sheep. PLoS One 2011; 6:e18943.

    • Crossref
    • Search Google Scholar
    • Export Citation
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