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Ultrastructural and Morphometric Description of the Ear Skin

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Ultrastructural and Morphometric Description of the Ear Skin Pesq. Vet. Bras. 41:e06775, 2021 DOI: 10.1590/1678-5150-PVB-6775 Original Article Wildlife Medicine ISSN 0100-736X (Print) ISSN 1678-5150 (Online) Ultrastructural and morphometric description of the ear skin and cartilage of two South American wild histricognate rodents (Dasyprocta leporina and Galea spixii)1 Alexsandra F. Pereira2*, Leonardo V.C Aquino2, Matheus B. Nascimento2, Ferdinando V.F. Bezerra3, Alana A. Borges2, Érika A. Praxedes2 and Moacir F. Oliveira3 ABSTRACT.- Pereira A.F., Aquino L.V.C., Nascimento M.B., Bezerra F.V.F., Borges A.A., Praxedes E.A. & Oliveira M.F. 2021. Ultrastructural and morphometric description of the ear skin and cartilage of two South American wild histricognate rodents (Dasyprocta leporina and Galea spixii). Pesquisa Veterinária Brasileira 41:e06775, 2021. Universidade Federal Rural do Semi-Árido, Av. Francisco Mota 572, Presidente Costa e Silva, Mossoró, RN 59625-900, Título Original Brazil. E-mail: [email protected] Skin and cartilage have been the main source for the recovery of somatic cells to be used in conservation strategies in wild mammals. In this sense, an important step for the [Título traduzido]. cryopreservation of these samples is to recognize the properties of the skin and cartilage. Thus, knowing that the skin may differ among species and aiming to contribute to the establishment of cryobanks, the study examined the differences in the ear skin and cartilage of wild rodents from South America, agouti (Dasyprocta leporina) and spix’s yellow-toothed Autores cavy (Galea spixii). Ultrastructural and quantitative methods were used to measure skin and and proliferative activity. Although ultrastructural analysis revealed a similar pattern cartilagebetween thickness,species, morphometric density of collagen analysis and elastic of the fibers, skin and cell cartilagetype number showed and distribution, differences between agoutis and cavies regarding thickness of epidermis layers (corneum: 5.3±2.5µm vs. 3.9±0.6µm; intermediate: 16.4±6.2µm vs. 23.4±8.1µm; basal: 9.9±2.1µm vs. 4.8±0.5µm), dermis (183.1±44.0µm vs. 258.2±22.9µm), total skin (211.8±46.0µm vs. 290.3±23.7µm) and perichondrium (27.6±6.1µm vs. 10.5±1.8µm). A greater number of epidermal cells (61.7±15.2 vs. 24.8±7.6) and chondrocytes (32.7±9.0 vs. 27.5±4.7) were observed in agouti, while the cavy presented a greater number of melanocytes (12.6±4.7 vs. 29.9±6.2), keratinocytes (14.7±4.2 vs.to the 29.8±7.6), skin of agoutis.and fibroblasts Therefore, (103.6±24.7 there are vs. differences 112.2±11.3). between Moreover, agouti a andhigher cavy percentage for ear skin of collagen fibers and proliferative activity was observed in the skin of cavies, when compared INDEX TERMS: Wild rodents, , , ultrastructure, morphology, morphometry, and cartilage, requiring theDasyprocta establishment leporina ofGalea species-specific spixii cryopreservation protocols. ear skin, cartilage, cryobanks, wildlife animals. RESUMO.- [Descrição ultraestrutural e morfométrica da de recuperação de células somáticas a serem utilizadas em pele e cartilagem auricular de dois roedores histricognatos estratégias de conservação em mamíferos silvestres. Nesse silvestres da América do Sul (Dasyprocta leporina e Galea contexto, uma importante etapa para criopreservação é spixii)]. A pele e cartilagem têm sido uma importante fonte conhecer, inicialmente, as propriedades que compõem a pele e cartilagem. Sabendo, então, que a pele pode diferir- 1 Received on December 6, 2020. se entre espécies e com o objetivo de contribuir para o Accepted for publication on December 26, 2020. estabelecimento de criobancos, o estudo evidenciou as 2 Laboratório de Biotecnologia Animal, Universidade Federal Rural do diferenças da pele e da cartilagem do pavilhão auricular apical Semi-Árido (UFERSA), BR-110 Km 47, Presidente Costa e Silva, Mossoró, RN de cutias ( ) e preás ( ) que são 599625-900, Brazil. *Corresponding author: [email protected] Dasyprocta leporina Galea spixii 3 roedores silvestres presentes na América do Sul. Para tanto, Rural do Semi-Árido (UFERSA), BR-110 Km 47, Presidente Costa e Silva, métodos ultraestruturais e quantitativos foram utilizados para Mossoró,Laboratório RN 599625-900, de Morfofisiologia Brazil. Animal Aplicada, Universidade Federal mensurar a espessura da pele e da cartilagem, densidade de 1 2 Alexsandra F. Pereira et al. celulares e atividade proliferativa. Embora as propriedades ultrastructure and composition of the skin and cartilage of fibrasultraestruturais colágenas e emelásticas, cutias número e preás e tenham distribuição se mostrado dos tipos cartilagesthe South American of these two rodents’ species. agouti We specificallyand spix’s yellow-toothed examined the semelhantes, avaliações acerca da morfometria da pele e cavy, for the establishment of suitable biological resource da cartilagem demonstrou diferenças, especialmente nas banks for these species. camadas epidérmicas (córnea: 5,3±2,5µm vs. 3,9±0,6µm; espinhosa: 16,4±6,2µm vs. 23,4±8,1µm; basal: 9,9±2,1µm MATERIALS AND METHODS vs. 4,8±0,5µm), derme (183,1±44,0µm vs. 258,2±22,9µm), Except where indicated, all chemicals, media, and solutions were pele total (211,8±46,0µm vs. 290,3±23,7µm) e pericôndrio purchased from Sigma Chemical Co. (St. Louis/MO, USA), Gibco- (27,6±6,1µm vs. BRL (Carlsbad/CA, USA), and Labimpex (São Paulo/SP, Brazil). de células epidérmicas (61,7±15,2 vs. 24,8±7,6) e condrócitos All experimental protocols and animal care were approved by (32,7±9,0 vs. 27,5±4,7) 10,5±1,8µm). foram observados Além disso, em um cutias, número enquanto maior the Animal Ethics Committee of the “Universidade Federal Rural do Semi-Árido” (CEUA-UFERSA, no. 23091.001072/2015-92 and no. 23091.010566/2017-20), and the “Instituto Chico Mendes de em(103,6±24,7 preás um vs. maior 112,2±11,3) número foram de melanócitos evidenciados. (12,6±4,7 Ainda, emvs. Conservação da Biodiversidade” (ICMBio; no. 48633-2 and no. 604281). 29,9±6,2), queratinócitos (14,7±4,2 vs. 29,8±7,6) e fibroblastos Eight male agoutis proliferativa foram observadas quando comparadas a pele Animals, ear cartilage, and skin collection. (Dasyprocta leporina; 8 months old) and eight male spix’s yellow- preás,de cutias. uma Portanto,maior porcentagem existem diferençasde fibras colágenas entre cutias e da atividade e preás toothed cavy ( ; 8 months old) obtained from the Center para pele e cartilagem do pavilhão auricular, exigindo desta Galea spixii for Multiplication of Wild Animals (no. 1478912) were used in this forma um estabelecimento de protocolos de criopreservação study. The animals were previously anesthetized by intramuscular TERMOS DE INDEXAÇÃO: Roedores silvestres, , São Paulo/SP, Brazil) and 1mg/kg xylazine hydrochloride (Rompun; específica para cada uma destas espécies.Dasyprocta leporina Galea spixii, ultraestrutura, morfologia, morfometria, pele auricular, Bayer,administration São Paulo/SP, of 15mg/kg Brazil), ketamine according hydrochloride to Castelo et (Ketalar; al. (2015). Pfizer, Ear cartilagem, criobancos, animais selvagens. margin tissues (1-2cm2) were obtained using pliers and transported supplemented with 2% antibiotic-antimycotic solution (penicillin INTRODUCTION G,to streptomycin,the laboratory and in Dulbecco’s amphotericin modified B) at 37°CEagle’s for medium 1 h. (DMEM) Agouti (Dasyprocta leporina) and spix’s yellow-toothed In the laboratory, tissues were washed in 70% ethanol, and by using cavy (Galea spixii) are histricognate rodents belonging to a scalpel blade and a cutting mold, eight fragments (9.0mm3; 3 × 3 × the Dasyproctidae and Caviidae family, respectively, and 1mm) per individual per species were obtained and were equally and distributed throughout the Brazilian territory (Silva et al. randomly allocated for ultrastructural and morphometric analyses. 2017). These animals play different roles in the functionality of Processing and ultrastructural analysis. Initially, two fragments the environment that they inhabit and are models for rodents under the threat of extinction (Praxedes et al. 2018b), since from each individual were fixed in 2.5% glutaraldehyde solution in Reid 2016). In other species, the use of an experimental model andphosphate-buffered dehydrated with saline increasing (PBS) for concentrations five days. After of this ethanol period, (Ciena tissues et theirwith populationsa stable population are stable has (Catzeflis already et been al. 2016, and interestingEmmons & al.were 2017). post-fixed For scanning in 1% osmiumelectron microscopytetroxide diluted analysis, in distilled samples water were alternative and an important tool used in animal conservation conditioned in the drying equipment to the critical point of carbon research programs (Cardoso et al. 2020, Echeverry et al. 2020). dioxide (K850 Critical Point Dryer; Quorum Technologies, Lewes, Thus, in order to respond to this demand, for the study East Sussex, United Kingdom), placed in a stub, and metallized with of these species, skin and cartilage cryobanks have been gold. Finally, tissue ultrastructure was visualized using a scanning established in recent years (Praxedes et al. 2018a, Pereira et electron microscope (TESCAN VEGA3; Tescan Analytics, Fuveau, al. 2019), since skin and cartilage somatic cells can be used Bouches-du-Rhône, France). in the multiplication of individuals using cloning (Azuma Processing and morphometric analysis using histological et al. 2018), in regenerative medicine (Aliborzi et al. 2016), staining. and genetic (Comizzoli 2017) and morphological research
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