Synergistic effects of follicle-stimulating hormone and epidermal growth factor on in vitro maturation and parthenogenetic development of red-rumped agouti oocytes.

Although oocyte in vitro maturation (IVM) is routinely used for in vitro embryo production in mice and rats, its use in wild rodents remains unexplored. Evidence suggests that hormone and growth factor supplementation influence oocyte meiotic resumption. This study evaluated the synergistic effects of follicle-stimulating hormone (FSH) and epidermal growth factor (EGF) on the IVM and parthenogenetic development of red-rumped agouti oocytes. Initially, we evaluated the IVM rates, mature oocyte quality, oocyte morphometry, and early embryonic development during IVM in the presence of 10, 50, and 75 mIU/mL FSH. No differences among the FSH concentrations were observed for IVM rates, oocyte morphometry, cumulus cell expansion, and viability. Although oocytes matured with 50 mIU/mL FSH showed a higher rate of cumulus expansion index (CEI), only oocytes matured with 10 mIU/mL FSH resulted in morulae after chemical activation (7.9% ± 4.2%). Thus, 10 mIU/mL FSH was used for further experiments. We subsequently evaluated the synergistic effects of 10, 50, and 100 ng/mL EGF and 10 mIU/mL FSH on the same parameters. No differences among the groups were observed in IVM rates, oocyte morphometry, and cumulus viability. Nevertheless, FSH with 10 ng/mL EGF showed a CEI superior to that of the other groups. Furthermore, oocytes matured with FSH alone or with both FSH and 10 or 50 ng/mL EGF developed morulae after activation (5.8%-8.3%). In conclusion, oocytes matured with 10 mIU/mL FSH and 10 ng/mL EGF are recommended for use in red-rumped agouti oocyte IVM, as they positively influence embryonic development.

Different Methods for Seminal Plasma Removal and Sperm Selection on the Quality and Fertility of Collared Peccary Sperm.

Methods for seminal plasma (SP) removal and the selection of collared peccary sperm for fertilization were compared. The experiments evaluated the following: the (I) impact of centrifugation for SP removal before swim-up for sperm selection and (II) a comparison of different Percoll® gradient densities (PG 45-90% and PG 35-70%). Non-selected sperm served as the control. Sperm quality was assessed based on motility patterns, morphology, membrane functional integrity, viability, reactive oxygen species (ROS), glutathione (GSH), and DNA integrity. Subsequently, the most successful group in the previous experiment and washing by centrifugation (WC) were compared for motility patterns and fertilization using pig oocytes. Swim-up decreased motility and enhanced ROS compared to the control. Centrifugation before swim-up harmed integrity and viability compared to the control. PG 45-90% (96.8 vs. 69.7 vs. 40.7 µm/s) allowed for a better velocity average pathway (VAP), a better velocity straight line, and better linearity (LIN) than those of the control and PG 35-70% (88.4 vs. 56.0 vs. 27.3 µm/s). Thus, PG 45-90% was used for fertilization. PG 45-90% obtained a higher VAP, a higher amplitude of the lateral head, straightness, and higher LIN than those of the control and WC. Cleavage (25.2-26.3%) and morula (8.1-10.5%) rates did not differ between the groups. Therefore, PG 45-90% and WC were efficient in isolating collared peccary sperm capable of fertilizing pig oocytes.

Changes in Sperm Morphology, Morphometry, and Motility from the Epididymis to the Vas Deferens in Rheas (Rhea americana, Linnaeus, 1758).

The objective was to characterize morphological, morphometric, and ultrastructural changes in rhea spermatozoa between the epididymis and the vas deferens. Sperm samples were collected from the reproductive tracts of seven adult individuals and evaluated for sperm characteristics using brightfield microscopy as well as ultrastructural features using scanning electron microscopy (SM). Mean sperm count tended to increase in the vas deferens (378.0 ± 135.0 × 106) compared to the epididymis (201.0 ± 77.4 × 106). Percentages of motile sperm grew from 37.0 ± 4.9% in the epididymis to 58.5 ± 7.7% in the vas deferens. The proportion of normal spermatozoa was 75.6 ± 1.8% and most common defects were bent tails (9.7 ± 0.9%). However, these proportions were not different between epididymis and vas deferens. SM analysis revealed further features of rhea spermatozoa. Normal rhea spermatozoa were threadlike with an acrosome (0.95 ± 0.0 µm), head (7.53 ± 0.01 µm), midpiece (2.08 ± 0.01 µm), and tail (30.7 ± 0.06 µm). Lengths of sperm acrosome, head, midpiece, and tail were longer in the vas deferens compared to the epididymis. Our findings suggest that rhea spermatozoa undergo a maturation process during the passage from the epididymis to the vas deferens.

Influence of Intracellular Cryoprotectants on the Conservation of Dermal Somatic Tissues Derived from Antillean Manatees (Trichechus manatus manatus Linnaeus, 1758).

Cryopreservation of somatic tissue has been studied as a tool for the knowledge and conservation of endangered species, such as Antillean manatees. The use of vitrification protocols is an important step in the establishment of biological banks. To decrease the damage caused by this technique, a reduction in the concentration of cryoprotectants has been proposed. Therefore, we aimed to evaluate combinations and concentrations of intracellular cryoprotectants for the conservation of somatic tissues derived from Antillean manatees. Dulbecco's modified Eagle's medium, F-12 composed of 10% fetal bovine serum and 0.25 M sucrose, was supplemented with 3.0 M ethylene glycol (EG) plus 3.0 M dimethyl sulfoxide (DMSO), or 1.5 M EG plus 1.5 M DMSO or 3.0 M EG or 3.0 M DMSO, to produce four solutions for solid-surface vitrification. Noncryopreserved tissues were used as the controls. After warming, tissues derived from four Antillean manatees were evaluated for ultrastructure, histology, and in vitro culture. No differences were observed among the cryopreserved and noncryopreserved tissues in terms of ultrastructure. The dermis thickness of the cryopreserved fragments in solutions containing 3.0 M EG plus 3.0 M DMSO, 3.0 M EG, and 3.0 DMSO was similar to that of the control. Moreover, cryopreservation with 3.0 M EG plus 3.0 M DMSO maintained tissue proliferative capacity potential evaluated by quantification of nucleolar organizing regions. Nevertheless, none of the cryopreserved fragments were able to maintain the number of fibroblasts and the collagen percentage as compared with that of the noncryopreserved fragments. Also, none of the cryopreserved fragments in the different solutions were able to produce cells in vitro. In summary, even reducing the concentration of intracellular cryoprotectants as well as their association did not guarantee the maintenance of cells after in vitro culture. Further studies are needed to optimize the cryopreservation protocols in Antillean manatee somatic tissues.

Morphological, morphometric, ultrastructural, and functional evaluation of red-rumped agouti (Dasyprocta leporina) sperm during epididymal transit.

The red-rumped agouti (Dasyprocta leporina) is a hystricognath rodent with reproductive anatomical peculiarities presenting as an intra-abdominal testes-epididymis complex. This study was carried out to describe, for the first time, details related to the morphological and functional changes in sperm along the epididymal transit in agoutis. The testes-epididymal complexes were sampled from seven sexually mature agoutis. Sperm from different epididymal regions (caput, corpus, and cauda) were collected using the floating technique, and their morphology, morphometry, ultrastructure, mitochondrial activity, membrane structural integrity, and kinetic parameters were determined. The number of sperm collected (823.5 ×106 sperm) was higher in the epididymis cauda. No significant differences in normal sperm morphology among the different epididymal regions (caput, 82.42%; corpus, 86.71%; and cauda, 88.86 %) were observed. The mean head length, head width, and tail length were highest in the caput (5.15 µm, 3.44 µm, and 32.04 µm, respectively), decreasing along the epididymal transit. Ultrastructure by scanning electron microscopy (SEM) revealed agglomeration of spermatozoa from caput and corpus, thus, enabling analysis of the gametes from only the epididymal cauda with clarity. Sperm from epididymis cauda showed the greatest proportion of membrane integrity and mitochondrial activity, followed by those from corpus and caput (79.71 %, 58.9 %, 47.7 %, respectively). Significant increase in total motility, progressive motility, velocity average pathway -VAP, velocity straightline - VSL, velocity curvilinear - VCL, and rapid sperm in the caput-corpus-cauda direction were observed. These novel data contribute to the knowledge of sperm maturation in the red-rumped agouti.

Cryopreservation of Testicular Tissue from Adult Red-Rumped Agoutis (Dasyprocta leporina Linnaeus, 1758).

This study measured the effects of different freezing techniques and permeating cryoprotectants on the preservation of testicular tissues from adult red-rumped agoutis. Tissue biopsies (3.0 mm3) from five individuals were allocated to different experimental groups: control (non-cryopreserved); slow freezing (SF), solid-surface vitrification (SSV), and conventional vitrification (CV). Each method used dimethyl sulfoxide (DMSO), ethylene glycol (EG), or a DMSO + EG combination. Morphology, viability, mitochondrial activity, and proliferative potential were assessed in fresh and frozen tissue samples. Testicular morphology was better using SSV with a combination of DMSO and EG. Across the different cryopreservation approaches, as well as cryoprotectant combinations, cell viability was comparable. Regarding mitochondrial activity, DMSO + EG/SSV or CV, and DMSO + EG/CV were similar to the EG/SF group, which was the best group that provided values similar to fresh control groups. Adequate preservation of the proliferative potential of spermatogonia, Leydig cells, and Sertoli cells was obtained using SSV with DMSO + EG. Overall, the use of SSV with DMSO + EG was the best protocol for the preservation of testicular tissues from adult red-rumped agoutis.

Diseases of the digestive system of agoutis (Dasyprocta leporina) raised in captivity in the Brazilian semiarid region / Doenças do sistema digestivo de cutias (Dasyprocta leporina) criadas em cativeiro na região semiárida brasileira

The objective of this study was to describe the clinical and pathological aspects of diseases of the digestive system in agoutis (Dasyprocta leporina Linnaeus, 1758) diagnosed by the "Laboratório de Patologia Veterinária" (Veterinary Pathology Laboratory) of the "Universidade Federal Rural do Semi-Árido" (UFERSA), from January 2018 to February 2020. During the study period, necropsy and a survey of the clinical history of 27 agoutis were performed, 25.93% (7/27) of which were diagnosed with digestive system diseases. The percentages of digestive tract diseases among the diagnosed were: acute carbohydrate overload (11.12%), gastric ulcer (7.41%), gastric volvulus (3.70%), and intestinal volvulus (3.70%). Studies on the occurrence rate of these diseases, as well as the description of their clinical and anatomopathological aspects, may serve as a basis for guiding the appropriate management in the breeding of these animals.(AU)

O objetivo deste estudo foi descrever os aspectos clínicos e patológicos das doenças do aparelho digestivo em cutias (Dasyprocta leporina Linnaeus, 1758) diagnosticadas pelo Laboratório de Patologia Veterinária da Universidade Federal Rural do Semiárido (UFERSA), de janeiro 2018 a fevereiro de 2020. Durante o período do estudo, foram realizadas necropsias e levantamento da história clínica de 27 cutias, sendo 25,93% (7/27) diagnosticadas com doenças do aparelho digestivo. Os percentuais de doenças do aparelho digestivo foram: sobrecarga aguda de carboidratos (11,12%), úlcera gástrica (7,41%), vólvulo gástrico (3,70%) e vólvulo intestinal (3,70%). Estudos sobre a taxa de ocorrência dessas doenças, bem como a descrição de seus aspectos clínicos e anatomopatológicos, podem servir de base para orientar o manejo adequado na criação dessa espécie.(AU)

Diseases of the digestive system of agoutis (Dasyprocta leporina) raised in captivity in the Brazilian semiarid region / Doenças do sistema digestivo de cutias (Dasyprocta leporina) criadas em cativeiro na região semiárida brasileira

The objective of this study was to describe the clinical and pathological aspects of diseases of the digestive system in agoutis (Dasyprocta leporina Linnaeus, 1758) diagnosed by the "Laboratório de Patologia Veterinária" (Veterinary Pathology Laboratory) of the "Universidade Federal Rural do Semi-Árido" (UFERSA), from January 2018 to February 2020. During the study period, necropsy and a survey of the clinical history of 27 agoutis were performed, 25.93% (7/27) of which were diagnosed with digestive system diseases. The percentages of digestive tract diseases among the diagnosed were: acute carbohydrate overload (11.12%), gastric ulcer (7.41%), gastric volvulus (3.70%), and intestinal volvulus (3.70%). Studies on the occurrence rate of these diseases, as well as the description of their clinical and anatomopathological aspects, may serve as a basis for guiding the appropriate management in the breeding of these animals.

O objetivo deste estudo foi descrever os aspectos clínicos e patológicos das doenças do aparelho digestivo em cutias (Dasyprocta leporina Linnaeus, 1758) diagnosticadas pelo Laboratório de Patologia Veterinária da Universidade Federal Rural do Semiárido (UFERSA), de janeiro 2018 a fevereiro de 2020. Durante o período do estudo, foram realizadas necropsias e levantamento da história clínica de 27 cutias, sendo 25,93% (7/27) diagnosticadas com doenças do aparelho digestivo. Os percentuais de doenças do aparelho digestivo foram: sobrecarga aguda de carboidratos (11,12%), úlcera gástrica (7,41%), vólvulo gástrico (3,70%) e vólvulo intestinal (3,70%). Estudos sobre a taxa de ocorrência dessas doenças, bem como a descrição de seus aspectos clínicos e anatomopatológicos, podem servir de base para orientar o manejo adequado na criação dessa espécie.

Diseases of the digestive system of agoutis (Dasyprocta leporina) raised in captivity in the Brazilian semiarid region

ABSTRACT: The objective of this study was to describe the clinical and pathological aspects of diseases of the digestive system in agoutis (Dasyprocta leporina Linnaeus, 1758) diagnosed by the Laboratório de Patologia Veterinária (Veterinary Pathology Laboratory) of the Universidade Federal Rural do Semi-Árido (UFERSA), from January 2018 to February 2020. During the study period, necropsy and a survey of the clinical history of 27 agoutis were performed, 25.93% (7/27) of which were diagnosed with digestive system diseases. The percentages of digestive tract diseases among the diagnosed were: acute carbohydrate overload (11.12%), gastric ulcer (7.41%), gastric volvulus (3.70%), and intestinal volvulus (3.70%). Studies on the occurrence rate of these diseases, as well as the description of their clinical and anatomopathological aspects, may serve as a basis for guiding the appropriate management in the breeding of these animals.

RESUMO: O objetivo deste estudo foi descrever os aspectos clínicos e patológicos das doenças do aparelho digestivo em cutias (Dasyprocta leporina Linnaeus, 1758) diagnosticadas pelo Laboratório de Patologia Veterinária da Universidade Federal Rural do Semiárido (UFERSA), de janeiro 2018 a fevereiro de 2020. Durante o período do estudo, foram realizadas necropsias e levantamento da história clínica de 27 cutias, sendo 25,93% (7/27) diagnosticadas com doenças do aparelho digestivo. Os percentuais de doenças do aparelho digestivo foram: sobrecarga aguda de carboidratos (11,12%), úlcera gástrica (7,41%), vólvulo gástrico (3,70%) e vólvulo intestinal (3,70%). Estudos sobre a taxa de ocorrência dessas doenças, bem como a descrição de seus aspectos clínicos e anatomopatológicos, podem servir de base para orientar o manejo adequado na criação dessa espécie.

Evaluation of different skin regions derived from a postmortem jaguar, Panthera onca (Linnaeus, 1758), after vitrification for development of cryobanks from captive animals.

Biological resource banks represent valuable tools for the conservation of species vulnerable to extinction, such as the jaguar. Cryobanks of skins have the potential to safeguard rare genotypes, allowing the potential exploitation of biological samples in animal multiplication technologies and the study of genetic variability. Determination of the most suitable skin regions for tissue conservation can help increase the efficiency of cryobanks and the storage of biological samples. To this end, we evaluated the effects of vitrification of skin tissues from the ear, caudal, and femoral regions of a post-mortem jaguar belonging to a zoo in Brazil. Non-vitrified and vitrified samples were evaluated and compared using quantitative methods, focusing on skin thickness, cell quantification, number of perinuclear halos, collagen and elastic density, and proliferative activity. No differences were observed in skin thickness, number of perinuclear halos, elastic density, and proliferative activity between non-vitrified and vitrified tissues in skin from any region. However, vitrified tissues derived from femoral skin showed a reduction in the number of fibroblasts, epidermal cells and collagen density compared to non-vitrified tissues. In summary, the ear and caudal regions provided the best conservation of somatic tissues derived from jaguars, and skin samples from these regions are therefore the most suitable for the formation of cryobanks.

Responses of matrix metalloproteinases to hyperbaric oxygen treatment: changing for good or ill?

Background: Hyperbaric oxygen (HBO2) is currently emerging as a promising therapeutic option for diseases involving impaired tissue repair and remodeling. In this regard, HBO2 has been shown to modulate signaling pathways responsible for matrix metalloproteinases (MMPs) regulation, which makes the MMPs interesting targets for investigation. However, the understanding regarding how HBO2 treatment affects the expression and activity of the MMP family members in different tissues and diseases needs to be clarified. The precise roles of MMPs in the physiopathology of various tissue repair disorders also remain unclear. Because of potential off-target systemic effects of the HBO2 on MMPs, researchers and physicians should carefully consider whether their patients could be affected adversely by HBO2 exposure. Aims: This narrative review provides an overview of MMP biology (structure, function, and regulation) and summarizes available data showing how MMPs respond to HBO2 in different tissues and pathologies, also highlighting possible mechanisms.

Ovarian gene expression in collared peccary (Pecari tajacu Linnaeus, 1758) subjected to gonadotropin stimulation protocols.

Ovarian response of collared peccaries (Pecari tajacu), after hormonal stimulation with gonadotropin association (eCG/hCG), was accessed by both gene expression and follicular development. Thus, collared peccaries (n = 8) were treated with the dose used for sows (swine dose, SWD) or with dose adjusted for peccary's weight (allometric dose, ALD). The gene expression of receptors was evaluated for both gonadotropins (FSHR and LHCGR) and growth factors (proteins codified by TGFßR-1, BMPR1-A and BMPR2 genes) in antral follicles, cortex and corpora haemorrhagica (CH). Five days after gonadotropin injection, all females presented CH. The ovulation rate was similar (p > .05) between SWD (4.00 ± 1.17) and ALD (2.50 ± 0.43) group. The total number of follicles per animal and amounts of small (<3 mm), medium (3-5 mm) and large (>5 mm) follicles was similar among groups. However, SWD produced large follicles heavier than ALD group, as accessed by weight of follicular wall biopsies. Ovarian follicles expressed both gonadotropin and growth factor receptors at levels which are independent from gonadotropin dose. In conclusion, the two gonadotropin doses (SWD and ALD) can be used for ovarian stimulation of collared peccary. Additionally, FSH and growth factors (TGFßR-1, BMPR1-A and BMPR2) receptors are more expressed in the early follicle development, while LH receptor seems to be more important in the final of follicular growth.

Spatial clustering of orientation preference in primary visual cortex of the large rodent agouti.

All rodents investigated so far possess orientation-selective neurons in the primary visual cortex (V1) but - in contrast to carnivores and primates - no evidence of periodic maps with pinwheel-like structures. Theoretical studies debating whether phylogeny or universal principles determine development of pinwheels point to V1 size as a critical constraint. Thus, we set out to study maps of agouti, a big diurnal rodent with a V1 size comparable to cats'. In electrophysiology, we detected interspersed orientation and direction-selective neurons with a bias for horizontal contours, corroborated by homogeneous activation in optical imaging. Compatible with spatial clustering at short distance, nearby neurons tended to exhibit similar orientation preference. Our results argue against V1 size as a key parameter in determining the presence of periodic orientation maps. They are consistent with a phylogenetic influence on the map layout and development, potentially reflecting distinct retinal traits or interspecies differences in cortical circuitry.

Ultrastructural and morphometric description of the ear skin and cartilage of two South American wild histricognate rodents (Dasyprocta leporina and Galea spixii) / Descrição ultraestrutural e morfométrica da pele e cartilagem auricular de dois roedores histricognatos silvestres da América do Sul (Dasyprocta leporina e Galea spixii)

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 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 cavy (Galea spixii). Ultrastructural and quantitative methods were used to measure skin and cartilage thickness, density of collagen and elastic fibers, cell type number and distribution, and proliferative activity. Although ultrastructural analysis revealed a similar pattern between species, morphometric analysis of the skin and cartilage showed 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. 29.8±7.6), and fibroblasts (103.6±24.7 vs. 112.2±11.3). Moreover, a higher percentage of collagen fibers and proliferative activity was observed in the skin of cavies, when compared to the skin of agoutis. Therefore, there are differences between agouti and cavy for ear skin and cartilage, requiring the establishment of species-specific cryopreservation protocols.(AU)

A pele e cartilagem têm sido uma importante fonte de recuperação de células somáticas a serem utilizadas em estratégias de conservação em mamíferos silvestres. Nesse contexto, uma importante etapa para criopreservação é conhecer, inicialmente, as propriedades que compõem a pele e cartilagem. Sabendo, então, que a pele pode diferir-se entre espécies e com o objetivo de contribuir para o estabelecimento de criobancos, o estudo evidenciou as diferenças da pele e da cartilagem do pavilhão auricular apical de cutias (Dasyprocta leporina) e preás (Galea spixii) que são roedores silvestres presentes na América do Sul. Para tanto, métodos ultraestruturais e quantitativos foram utilizados para mensurar a espessura da pele e da cartilagem, densidade de fibras colágenas e elásticas, número e distribuição dos tipos celulares e atividade proliferativa. Embora as propriedades ultraestruturais em cutias e preás tenham se mostrado semelhantes, avaliações acerca da morfometria da pele e da cartilagem demonstrou diferenças, especialmente nas 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 vs. 4,8±0,5μm), derme (183,1±44,0μm vs. 258,2±22,9μm), pele total (211,8±46,0μm vs. 290,3±23,7μm) e pericôndrio (27,6±6,1μm vs. 10,5±1,8μm). Além disso, um número maior de células epidérmicas (61,7±15,2 vs. 24,8±7,6) e condrócitos (32,7±9,0 vs. 27,5±4,7) foram observados em cutias, enquanto em preás um maior número de melanócitos (12,6±4,7 vs. 29,9±6,2), queratinócitos (14,7±4,2 vs. 29,8±7,6) e fibroblastos (103,6±24,7 vs. 112,2±11,3) foram evidenciados. Ainda, em preás, uma maior porcentagem de fibras colágenas e da atividade proliferativa foram observadas quando comparadas a pele de cutias. Portanto, existem diferenças entre cutias e preás para pele e cartilagem do pavilhão auricular, exigindo desta forma um estabelecimento de protocolos de criopreservação específica para cada uma destas espécies.(AU)

Live endothelial cells on plasma-nitrided and oxidized titanium: An approach for evaluating biocompatibility.

We evaluated the effects of titanium plasma nitriding and oxidation on live endothelial cell viscoelasticity. For this, mechanically polished titanium surfaces and two surfaces treated by planar cathode discharge in nitriding (36N2 and 24H2) and oxidant (36O2 and 24H2). Surfaces were characterized regarding wettability, roughness and chemical composition. Rabbit aortic endothelial cells (RAECs) were cultured on the titanium surfaces. Cell morphology, viability and viscoelasticity were evaluated by scanning electron microscopy (SEM), methyl thiazolyl tetrazolium (MTT) assay and atomic force microscopy (AFM), respectively. Grazing Incidence X-ray Diffraction confirmed the presence of TiN0,26 on the surface (grazing angle theta 1°) of the nitrided samples, decreasing with depth. On the oxidized surface had the formation of TiO3 on the material surface (Theta 1°) and in the deeper layers was noted, with a marked presence of Ti (Theta 3°). Both plasma treatments increased surface roughness and they are hydrophilic (angle <90°). However, oxidation led to a more hydrophilic titanium surface (66.59° ± 3.65 vs. 76.88° ± 2.68; p = 0.001) due to titanium oxide films in their stoichiometric varieties (Ti3O, TiO2, Ti6O), especially Ti3O. Despite focal adhesion on the surfaces, viability was different after 24 h, as cell viability on the oxidized surface was higher than on the nitrided surface (9.1 × 103 vs. 4.5 × 103cells; p < 0.05). This can be explained by analyzing the viscoelastic property of the cellular cytoskeleton (nuclear and peripheral) by AFM. Surface oxidation significantly increased RAECs viscoelasticity at cell periphery, in comparison to the nucleus (2.36 ± 0.3 vs. 1.5 ± 0.4; p < 0.05), and to the RAECs periphery in contact with nitrided surfaces (1.36 ± 0.7; p < 0.05) and polished surfaces (1.55 ± 0.6; p < 0.05). Taken together, our results have shown that titanium plasma treatment directly increased cell viscoelasticity via surface oxidation, and this mechanobiological property subsequently increased biocompatibility.

BMP-15 activity on in vitro development of collared peccary (Pecari tajacu Linnaeus, 1758) preantral follicles.

This study investigated the effects of BMP-15 on the in vitro development of preantral follicles of collared peccaries. Ovarian fragments were cultured for 1 or 6 days in Tissue Culture Medium 199 (TCM199+ ) supplemented with BMP-15 at rates of 0, 1, 25 or 50 ng/ml. The fragments were analysed histologically by evaluating follicular morphology, activation and growth as well as the potential for proliferation of granulosa cells. Our results show the addition of 25 ng/ml BMP-15 in the medium provided the greatest percentage of normal follicles (79.67% ± 0.69) when compared to other treatments (p < .05); however, this result is similar to 1 ng/ml BMP-15 (74.00% ± 1.90, p > .05). Moreover, 25 and 50 ng/ml of BMP-15 promoted follicular activation. BMP-15 supplements did not affect oocyte and follicular growth. All concentrations of BMP-15 increased the number of nucleolus organizer regions (NORs) after 1 day of culture when compared to fresh fragments or the control samples (p < .05). However, at the end of the experiment, the number of NORs in follicles cultured in all treatments was higher than that observed in the fresh control (sample taken prior to culturing) (p > .05). In summary, the addition of 25 ng/ml BMP-15 to the culture medium of collared peccary preantral follicles maintained a high number of morphologically healthy follicles and stimulated the activation of primordial follicles after 6 days in culture.

Effects of Vitrification Techniques on the Somatic Tissue Preservation of Agouti (Dasyprocta leporina Linnaeus, 1758).

The cryobanks of agouti somatic tissues represent a promising tool for the conservation of this species and of those that are phylogenetically related and endangered. For these purposes, one strategy to guarantee the quality of samples after warming would be to choose the appropriate tissue vitrification technique. Therefore, we evaluated the effects of two different techniques, direct vitrification in cryovials (DVC) and solid-surface vitrification (SSV), on the preservation of ear somatic tissues derived from agoutis kept in a scientific center of creation. Noncryopreserved somatic tissues were used as controls. Although SSV reduced the thickness of the dermis and cartilage (p < 0.05), the epidermal thickness of these samples was observed to be similar to controls (p > 0.05). Notably, the number of fibroblasts was not altered with either technique. However, both vitrification methods led to an increase in the number of perinuclear halos, with a particularly strong increase observed in DVC-derived fragments (p < 0.05). Compared with the DVC group, SSV showed a larger number of normal chondrocytes and smaller number of degenerate chondrocytes. Furthermore, the number of empty lacunae in SSV-derived fragments remained similar to controls (p > 0.05). In summary, SSV was found to be a more efficient method for vitrifying agouti somatic tissues compared with DVC. These results are important for the proper formation of agouti somatic banks, an essential step in the study of biological resources in this species.

Vertebromedullary Topography of the Red-Rumped Agouti (Dasyprocta leporina Linnaeus, 1758).

The agouti is a wild rodent located almost throughout the Brazilian territory, with great scientific importance and high reproductive potential when farmed. In the clinical routine, clinical and surgical procedures are performed on the nervous system of these animals, such as epidural anesthesia and cerebrospinal fluid (CSF) collection; however, data available are limited in literature on the main points of these procedures in this species. In this aspect, the objective was to describe the vertebromedullary topography of the agouti in order to identify and quantify it, with the intention of expanding the information regarding the clinical-surgical procedures in this system. Ten animals were incised longitudinally in the dorsal median line, removing the skin, the musculature, and dorsal fascia of the vertebral column; then, removed the vertebral arches and consequent exposure of the spinal cord and meninges. The morphometry of the animals was done by obtaining the total length, body length, and crown-rump length, as well as the total length of the spinal cord and its segments. The cervical intumescence in the agouti is located between C4-T1 and measures 2.45 cm. The lumbar intumescence is between L5-L7 and is 2.52 cm long. The cauda equina arises from S2 and the filum terminale from S3. As for the medullary cone, it is located between L7-S1 and is 2.52 cm long. The suggested localization for epidural anesthesia or CSF puncture for laboratory tests in agoutis is the lumbosacral space, with the insertion of the needle to be carried out at angle of 90° in the sphinx position. Anat Rec, 303:1472-1477, 2020. © 2019 American Association for Anatomy.

Comparison of different intracellular cryoprotectants on the solid surface vitrification of red-rumped agouti (Dasyprocta Leporina Lichtenstein, 1823) ovarian tissue.

In contributing to the conservation of wild rodents, the aim of this study was to evaluate the use of distinct cryoprotectants, separately or in combination, for solid surface vitrification (SSV) of red-rumped agouti ovarian tissue. Ovarian cortex from nine females was recovered and fragmented. Fresh fragments (control) were used to analyse the pre-antral follicle (PF) morphology using a histologic procedure, viability using the Trypan blue test, cell proliferation by counting the argyrophilic nucleolar organizing regions (Ag-NORs technique) and DNA integrity using the TUNEL assay. The remaining fragments were vitrified using SSV method with 3 M or 6 M ethylene glycol (EG) or dimethyl sulfoxide (DMSO), or in combination (3 M EG/3 M DMSO), and further evaluated as reported for the fresh samples. All cryoprotectants were effective at preserving PFs morphology compared to the control group (80.7 ± 5.21%), except 6 M EG and 3 M DMSO that provoked a significant (p < .05) decrease on the values of morphologically normal primary (60.0 ± 19.0%) and primordial (44 ± 4.5%) follicles, respectively. Regarding viability, all cryoprotectants provided values similar to that verified for the control group (79.0%), but a significant decrease (p < .05) was observed with EG/DMSO combination (59%). Using Ag-NORs technique, the highest (p < .05) cell proliferative capacity was detected when using EG at each tested concentration. The TUNEL proved the preservation of DNA integrity regardless of the cryoprotectant. In summary, we suggest the use of 3 M EG for the solid surface vitrification of red-rumped agouti ovarian tissue.

Single injection of eCG/hCG leads to successful estrous synchronization in the collared peccary (Pecari tajacu Linnaeus, 1758).

The establishment of protocols for the control of the ovarian function of collared peccaries is recommended for the development of assisted reproductive techniques. The goals were to (1) compare a gonadotropin combination with prostaglandin analogue to synchronize timing of onset of estrus among animals, and (2) elucidate the effects of the most desirable protocol for performing an artificial insemination study and macroscopic evaluation of the ovaries. Three of five females treated with a double administration of 120 µg prostaglandin (cloprostenol) at a 9-day interval expressed symptoms of estrus 9 days after the second injection. One female presented estrus after 6 days, whereas other did not respond to the treatment. All females (5/5) treated with a single dose containing 400 IU eCG and 200 IU hCG manifested estrus 6 days after the hormone injection. In a second experiment, ten females that were estrous synchronized using eCG/hCG, were artificially inseminated with fresh semen and monitored for pregnancy every 30 days. Although there was no detection of fetuses by ultrasonic examination, seven females (7/10) had greater than basal progesterone values for 60 days after the treatments were imposed. Ovaries from two females treated with eCG/hCG were collected 6 days post-injection. There was confirmation of an ovarian stimulation as a result of the presence of 88 and 25 antral follicles, as well as three and eight hemorrhagic structures in ovaries of each female, respectively. It, therefore, is proposed that eCG/hCG can be used as an effective treatment for estrous synchronization in collared peccaries.