Canine mammary neoplasms - evaluation of tumor microenvironment

Background: The tumor microenvironment is an important target of studies in different types of neoplasms. Understanding the role of general components such as immune, vascular and fibroblastic cells has the objective of contributing to prognosis and treatment. The aim of this study was to evaluate the relationship between mast cells and angiogenesis in benign and malignant mammary neoplasms by investigating the role of degranulation and microlocation of mast cells and neoformed vessels in canine mammary neoplasms. Materials, Methods & Results: Mammary glands (n = 122) from 50 female dogs submitted to mastectomy without chemotherapy were evaluated and categorized into 3 groups: control group (n = 46); malignant group (n = 57) and benign group (n = 19). Lymph nodes without changes (n = 59) and with metastases (n = 6) were also evaluated. To evaluate the MCD (mast cell density) and angiogenesis, Toluidine Blue (0.1%) and Gomori’s Trichrome techniques were performed and adapted from previous studies. Photomicrographs of 10 hotspot areas on a 40x objective lens of the mammary glands and lymph nodes were captured to assess MCD and angiogenesis. In the absence of these areas, random fields were captured. For the mammary glands of the malignant and benign groups, 20 fields were analyzed, as the analysis considered the microlocation (peritumoral and intratumoral). Counting was performed manually using ImageJ software version 1.42q by 2 observers. The statistical analysis were performed using SPSS software version 19.0. The most frequent histological type in the malignant group was carcinoma in mixed tumor (68.42%; 39/57) and in the benign group was benign mixed tumor (57.89%; 11/19). Female dogs without breed pattern were more frequently affected represented 70% of the animals and the mean age was 9 years and 8 months ± 3 years and 1 month...

Canine mammary neoplasms - evaluation of tumor microenvironment

Background: The tumor microenvironment is an important target of studies in different types of neoplasms. Understanding the role of general components such as immune, vascular and fibroblastic cells has the objective of contributing to prognosis and treatment. The aim of this study was to evaluate the relationship between mast cells and angiogenesis in benign and malignant mammary neoplasms by investigating the role of degranulation and microlocation of mast cells and neoformed vessels in canine mammary neoplasms. Materials, Methods & Results: Mammary glands (n = 122) from 50 female dogs submitted to mastectomy without chemotherapy were evaluated and categorized into 3 groups: control group (n = 46); malignant group (n = 57) and benign group (n = 19). Lymph nodes without changes (n = 59) and with metastases (n = 6) were also evaluated. To evaluate the MCD (mast cell density) and angiogenesis, Toluidine Blue (0.1%) and Gomoris Trichrome techniques were performed and adapted from previous studies. Photomicrographs of 10 hotspot areas on a 40x objective lens of the mammary glands and lymph nodes were captured to assess MCD and angiogenesis. In the absence of these areas, random fields were captured. For the mammary glands of the malignant and benign groups, 20 fields were analyzed, as the analysis considered the microlocation (peritumoral and intratumoral). Counting was performed manually using ImageJ software version 1.42q by 2 observers. The statistical analysis were performed using SPSS software version 19.0. The most frequent histological type in the malignant group was carcinoma in mixed tumor (68.42%; 39/57) and in the benign group was benign mixed tumor (57.89%; 11/19). Female dogs without breed pattern were more frequently affected represented 70% of the animals and the mean age was 9 years and 8 months ± 3 years and 1 month...(AU)

Use of omentum flap for induction of free skin graft healing / [Utilização de flap de omento para indução da cicatrização de enxertos cutâneos]

The objective of this study was to assess healing induction of free skin grafts following transposition of omental flap through a subcutaneous tunnel to the recipient bed. Macroscopic and microscopic evaluations were performed. Nineteen piglets were used. Two surgical wounds were created of each subject. The graft removed from the left side (LS) was placed on the right side (RS) without the omental flap in the graft-bed (control group-CG). On the LS, an omental flap was placed between the graft removed from the RS and the recipient bed (omentum group-OG). Macroscopic evaluations showed edema, which gradually decreased on both groups. Suture dehiscence was highest at day 10 compared to other days in both groups. The CG had a higher incidence of unvitalized tissue compared to OG, although no difference was found among days of postoperative evaluation. The presence of unvitalized tissue was seen on 32% on OG and 53% on CG. Microscopic evaluations revealed higher collagenization, reepithelization, keratinization and less swelling in the OG compared to CG. In conclusion, mesh skin grafts evolved satisfactorily in swine even in newly created bedding without granulation tissue, but with appropriate vascularization. The omentum flap provided better macroscopic and microscopic outcomes regarding graft integration.

O objetivo deste estudo foi verificar a indução da cicatrização de enxertos cutâneos em malha após a utilização de flap de omento transposto através de túnel no subcutâneo até o leito receptor. Avaliações macroscópica e microscópica foram realizadas. Foram utilizados 19 suínos. Duas feridas cirúrgicas foram criadas em cada animal. O enxerto removido do lado esquerdo (LE) foi fixado do lado direito (LD), sem a presença do flap de omento entre o enxerto e o leito receptor (grupo controle - GC). No LE, foi fixado um flap de omento entre o enxerto cutâneo removido do LD e o leito receptor (grupo omento - GO). As avaliações macroscópicas mostraram que o edema diminuiu gradativamente em ambos os grupos. A deiscência foi maior no dia 10 em comparação aos demais dias, em ambos os grupos, entre os diferentes dias de avaliação. Foi verificada a presença de tecido desvitalizado em 32% do GO e em 53% no GC. Foi observada, na avaliação microscópica, maior colagenização, reepitelização, queratinização e menor edema no GO, quando comparado ao GC. Concluiu-se, com este estudo, que enxertos cutâneos em malha evoluíram satisfatoriamente em suínos, mesmo em leito receptor recém-criado e sem presença de tecido de granulação, desde que vascularizado, e que o flap de omento propiciou melhores resultados macro e microscópicos relativos à integração do enxerto.

BM-MSC-derived small extracellular vesicles (sEV) from trained animals presented nephroprotective potential in unilateralureteral obstruction model

Background: The efficacy of bone marrow mesenchymal stromal cells (BM-MSC) and its extracellular vesicles has been demonstrated for a broad spectrum of indications, including kidney diseases. However, BM-MSC donor characteristics and their potential are not usually considered. Therefore, the present work aims to evaluate the nephroprotective capacity of sEV secreted by BM-MSC from trained rats inunilateral ureteral obstruction (UUO) model. Methods: BM-MSC was characterized by their differentiation potential and immunophenotypic markers. The sEV were isolated by ultracentrifugation and characterized by nanoparticle tracking analysis and western blot. Its miRNA cargo was examined by quantitative PCR analysis for miR-26a, 126a, and 296. Wistar rats were submitted to UUO procedure and concomitantly treated with sEV secreted by BM-MSC from the untrained andtrained rats. The kidney tissue from all groups was evaluated for fibrosis mediators (transforming growth factor beta1 and collagen), CD34-angiogenesis marker, and hypoxia-inducible factor 1 alpha (HIF-1α). Results: Treadmill training stimulated in BM-MSC the production of sEV loaded with pro-angiogenic miR-296. The treatment with this sEVin UUO-rats was able to attenuate collagen accumulation and increase CD34 and HIF-1α in the kidney tissue when compared to untrained ones. Tubular proximal cells under hypoxia and exposed to BM-MSC sEV demonstrate accumulation in HIF-1α and NFR-2 (nuclear factor erythroid 2-related factor 2), possibly to mediate the response to hypoxia and oxidative stress, under these conditions. Conclusion: The BM-MSC sEV from trained animals presented an increased nephroprotective potential compared to untrained vesicles by carrying 296-angiomiR and contributing to angiogenesis in UUO model.(AU)

Skin graft associated with platelet-rich plasma in correcting extensive injuries resulting from the resection of skin cancer chemically induced in rats

ABSTRACT Purpose: To evaluate whether using platelet-rich plasma (PRP) in the graft recipient bed after the resection of a neoplasia can influence its recurrence because this product stimulates angiogenesis, mitogenesis and chemotaxis. Methods: A study with 30 rats Wistar (Rattus norvegicus albinus), which were separated into group A (induction of carcinogenesis, PRP in the postoperative period) and group B (induction of carcinogenesis, absence of PRP in the postoperative period), with 15 animals in each. Carcinogenesis was induced on the skin of the animals' chest by the topical application of 0.5% dimethylbenzantracene (DMBA) diluted in acetone. After surgical resection of the induced neoplasia, PRP was used to stimulate angiogenesis before surgical wound synthesis. Data on the control and experimental groups and macroscopic and microscopic variables were evaluated using analysis of variance and the Tukey's test (5%). Results: It was possible to determine that the use of PRP is good in reconstructive surgeries, but it is contraindicated in patients during tumor resection, as it can cause changes in the surgical bed, in addition to stimulating recurrences and metastases. Conclusions: PRP may interact with tumour cells that were in the recipient site of the surgical wound during the resection of a neoplasia, and a local recurrence process can be triggered by applying this product.

BM-MSC-derived small extracellular vesicles (sEV) from trained animals presented nephroprotective potential in unilateralureteral obstruction model

Abstract Background: The efficacy of bone marrow mesenchymal stromal cells (BM-MSC) and its extracellular vesicles has been demonstrated for a broad spectrum of indications, including kidney diseases. However, BM-MSC donor characteristics and their potential are not usually considered. Therefore, the present work aims to evaluate the nephroprotective capacity of sEV secreted by BM-MSC from trained rats inunilateral ureteral obstruction (UUO) model. Methods: BM-MSC was characterized by their differentiation potential and immunophenotypic markers. The sEV were isolated by ultracentrifugation and characterized by nanoparticle tracking analysis and western blot. Its miRNA cargo was examined by quantitative PCR analysis for miR-26a, 126a, and 296. Wistar rats were submitted to UUO procedure and concomitantly treated with sEV secreted by BM-MSC from the untrained andtrained rats. The kidney tissue from all groups was evaluated for fibrosis mediators (transforming growth factor beta1 and collagen), CD34-angiogenesis marker, and hypoxia-inducible factor 1 alpha (HIF-1). Results: Treadmill training stimulated in BM-MSC the production of sEV loaded with pro-angiogenic miR-296. The treatment with this sEVin UUO-rats was able to attenuate collagen accumulation and increase CD34 and HIF-1 in the kidney tissue when compared to untrained ones. Tubular proximal cells under hypoxia and exposed to BM-MSC sEV demonstrate accumulation in HIF-1 and NFR-2 (nuclear factor erythroid 2-related factor 2), possibly to mediate the response to hypoxia and oxidative stress, under these conditions. Conclusion: The BM-MSC sEV from trained animals presented an increased nephroprotective potential compared to untrained vesicles by carrying 296-angiomiR and contributing to angiogenesis in UUO model.

Clínica cirúrgica de pequenos animais: controle microbiológico na obtenção do plasma rico em plaquetas (PRP) / Microbiological control in obtaining platelet-rich plasma (PRP)

O plasma rico em plaquetas é um produto obtido por meio da centrifugação do sangue, e devido as suas propriedades de estimular a angiogênese, este é utilizado em cirurgias reconstrutivas a fim de evitar complicações pós-cirúrgicas. A coleta do sangue é feita de forma asséptica para obtenção do PRP, porém há possibilidade de contaminação bacteriana no momento da sua confecção podendo assim ocasionar complicações ao paciente. Objetivou-se ressaltar a importância do rigor no controle microbiológico do produto obtido, bem como avaliar a possibilidade de contaminação bacteriana no momento da confecção (PRP), antes do seu emprego no leito de enxertos cutâneos em coelhos. No presente estudo, apenas quatro amostras das sessenta confeccionadas apresentaram colônias bacterianas, indicando que o plasma rico em plaquetas obtido de maneira asséptica é propício e seguro em cirurgias reconstrutivas. Conclui-se que o rigor no controle microbiológico aliado a adequada desinfecção do centro cirúrgico e antissepsia do campo operatório, bem como o preparo asséptico do PRP, seu uso é indicado e seguro, não provocando riscos de contaminação da ferida cirúrgica.

Platelet-rich plasma is a product obtained by centrifuging blood, and due to its properties of stimulating angiogenesis, it is used in reconstructive surgery in order to avoid post-surgical complications. Blood collection is done aseptically to obtain platelet-rich plasma, but there is a possibility of bacterial contamination at the time of making it, thus causing complications to the patient. The objective was to emphasize the importance of rigor in the microbiological control of the product obtained, as well as to evaluate the possibility of bacterial contamination when making plateletrich plasma (PRP), before its use in the bed of skin grafts in rabbits. In the present study, only four samples of the sixty made had bacterial colonies, indicating that the platelet-rich plasma obtained aseptically is conducive and safe in reconstructive surgery. It is concluded that the rigor in microbiological control combined with adequate disinfection of the operating room and antisepsis of the operating field, as well as the aseptic preparation of the PRP, its use is indicated and safe, without causing risks of contamination of the surgical wound.

Avaliação morfo-histológica e morfo-histométrica de feridas cutâneas tratadas com Sphagneticola trilobata (L) Pruski em ratos / [Morphohistological and morphohistometric evaluation of skin wounds treated with Sphagneticola trilobata (L) Pruski in rats]

O objetivo deste trabalho foi avaliar macro e microscopicamente a atividade cicatrizante da Sphagneticola trilobata em feridas cutâneas induzidas em ratos, a partir da aplicação de creme contendo extrato hidroalcoólico bruto de folhas da planta. A análise fitoquímica apresentou terpenos e flavonoides como compostos majoritários. Sessenta ratos foram divididos em três grupos experimentais (n=20): grupo tratado (GT), grupo controle (GC) e grupo controle absoluto (GCA). Quatro feridas excisionais de 0,8cm de diâmetro foram realizadas no dorso dos animais, tratadas diariamente e avaliadas nos tempos três, sete, 14 e 21 dias de pós-operatório (PO) quanto à contração e à avaliação macroscópica, morfo-histológica e morfo-histométrica. Macroscopicamente, não houve diferença estatística na contração das feridas entre os grupos testados. Na avaliação morfológica e na morfométrica, o GT apresentou menor concentração de células inflamatórias, maior e melhor preenchimento do tecido de granulação pelas fibras colágenas e melhor vascularização das feridas. Não houve diferença entre o GC e o GCA. Conclui-se que o creme à base do extrato hidroalcoólico bruto das folhas de Sphagneticola trilobata contribui positivamente para o processo de cicatrização das feridas em pele de ratos.(AU)

The objective of this work was to macro and microscopically evaluate the healing activity of Sphagneticola trilobata in rat-induced skin wounds by applying cream containing crude hydroalcoholic extract from plant leaves. The phytochemical analysis showed terpenes and flavonoids as major compounds. Sixty rats were divided into three experimental groups (n=20): treated group (GT), control group (CG) and absolute control group (GCA). Four 0.8cm diameter excision wounds were performed on the back of the animals, treated daily and evaluated at the three, seven, 14 and 21 postoperative days (PO) for contraction, macroscopic, morphologic and morphologic evaluation. The TG presented smaller scar area at 21 postoperative days (P<0.05). In the morphological and morphometric evaluation, the WG presented lower inflammation, greater and better filling of granulation tissue by collagen fibers and better wound vascularization. There was no difference between GC and GCA. It was concluded that the cream based on the crude hydroalcoholic extract of Sphagneticola trilobata leaves contribute positively to the healing process of the skin wounds of rats.(AU)

Clínica cirúrgica de pequenos animais: controle microbiológico na obtenção do plasma rico em plaquetas (PRP) / Microbiological control in obtaining platelet-rich plasma (PRP)

O plasma rico em plaquetas é um produto obtido por meio da centrifugação do sangue, e devido as suas propriedades de estimular a angiogênese, este é utilizado em cirurgias reconstrutivas a fim de evitar complicações pós-cirúrgicas. A coleta do sangue é feita de forma asséptica para obtenção do PRP, porém há possibilidade de contaminação bacteriana no momento da sua confecção podendo assim ocasionar complicações ao paciente. Objetivou-se ressaltar a importância do rigor no controle microbiológico do produto obtido, bem como avaliar a possibilidade de contaminação bacteriana no momento da confecção (PRP), antes do seu emprego no leito de enxertos cutâneos em coelhos. No presente estudo, apenas quatro amostras das sessenta confeccionadas apresentaram colônias bacterianas, indicando que o plasma rico em plaquetas obtido de maneira asséptica é propício e seguro em cirurgias reconstrutivas. Conclui-se que o rigor no controle microbiológico aliado a adequada desinfecção do centro cirúrgico e antissepsia do campo operatório, bem como o preparo asséptico do PRP, seu uso é indicado e seguro, não provocando riscos de contaminação da ferida cirúrgica.(AU)

Platelet-rich plasma is a product obtained by centrifuging blood, and due to its properties of stimulating angiogenesis, it is used in reconstructive surgery in order to avoid post-surgical complications. Blood collection is done aseptically to obtain platelet-rich plasma, but there is a possibility of bacterial contamination at the time of making it, thus causing complications to the patient. The objective was to emphasize the importance of rigor in the microbiological control of the product obtained, as well as to evaluate the possibility of bacterial contamination when making plateletrich plasma (PRP), before its use in the bed of skin grafts in rabbits. In the present study, only four samples of the sixty made had bacterial colonies, indicating that the platelet-rich plasma obtained aseptically is conducive and safe in reconstructive surgery. It is concluded that the rigor in microbiological control combined with adequate disinfection of the operating room and antisepsis of the operating field, as well as the aseptic preparation of the PRP, its use is indicated and safe, without causing risks of contamination of the surgical wound.(AU)

Evaluation of platelet-rich plasma gel as an angiogenesis-inducing agent in canine advancement skin flaps / Avaliação do gel de plasma rico em plaquetas como agente indutor da angiogênese em flapes cutâneos de avanço em cães

This work aimed to evaluate the effect of platelet-rich plasma (PRP) on advancement skin flaps in dogs regarding improvement of vascularization, with focus on increasing its viable area, since there are reports that it is a potential angiogenesis stimulator. The experimental group was composed of eight adult bitches, in which two advancement skin flaps were made in the ventral abdominal region. No product was applied in the control flap (CF), while PRP was used in the contralateral flap, called treated flap (TF). The areas were clinically evaluated every two days until the 7th postoperative day regarding skin color and presence of necrosis. At 10 days, both flaps were removed and submitted to histological examination and blood vessel morphometry. The vessels counted in each group were statistically analyzed by the F-test at 1% probability. Results showed no significant difference in macroscopic changes in the wound, or CF and TF vascularization, thus suggesting that PRP gel did not improve advancement skin flap angiogenesis in bitches under the experimental conditions in which this research was developed.(AU)

Objetivou-se com o presente artigo avaliar a ação angiogênica do gel de plasma rico em plaquetas (PRP) em flapes cutâneos de avanço em animais da espécie canina, visando aumentar a viabilidade da pele após o procedimento, uma vez que existem relatos de que o produto é um potente estimulador da angiogênese. O grupo experimental foi composto por oito cadelas adultas, onde foram confeccionados dois flapes de avanço (de padrão subdérmico) na região abdominal ventral. Em um dos flapes, considerado controle (FC) não foi aplicado nenhum produto, enquanto que no flape contralateral, denominado tratado (FT), foi usado o gel de PRP. As áreas foram macroscopicamente avaliadas a cada dois dias até o 7º dia de pós-operatório em relação à coloração da pele e presença de área de necrose, e com 10 dias ambos os flapes foram coletados por biópsia e submetidos ao exame histológico e morfometria dos vasos sanguíneos. Os vasos contados em cada grupo foram estatisticamente analisados pelo teste de F ao nível de 1% de probabilidades. Os resultados demonstraram que não houve diferença significativa nas alterações macroscópicas das feridas e na morfometria vascular dos FC e FT, sugerindo dessa maneira que dentro das condições experimentais nas quais a pesquisa foi executada, que o gel de PRP não incrementou a angiogênese de flapes de avanço em cadelas.(AU)

Effects of canine adipose-derived mesenchymal stem cells on the epithelialization of rabbits' skin autograft (Oryctolagus cuniculus) / Efeitos das células-tronco mesenquimais derivadas do tecido adiposo de cães na epitelização de autoenxertos de pele em coelhos (Oryctolagus cuniculus)

The present study aimed to evaluate the effects of mesenchymal stem cells derived from canine adipose tissue in the healing process of full-thickness mesh skin grafts in rabbits. The stem cells were collected from young dogs; and, after characterization, remained in cryopreservation, in independent doses containing 2 x 106 cells. The mesh distal limb graft technique was performed in 60 rabbits, divided into three groups, CG (Control Group), GT1 (Intralesional Stem Cell Treated Group), and GT2 (Intravenous Stem Cell Treated Group), containing 20 animals each. After grafting, each group was randomly divided into four subgroups according to euthanasia time 3, 7, 14, and 30 days, containing five animals in each group. Animals of GT1_14, GT1_30, and GT2_14, GT2_30 subgroups received a second dose of xenogeneic cells on the seventh day. Meanwhile, animals from GT1_30 and GT2_30 received the third dose of xenogeneic cells on day 14. The groups treated with xenogeneic stem cells positively affected type III collagen re-epithelialization and deposition, and possibly GT1 had a controlled inflammatory response. However, no effect on angiogenesis. Thus, it was possible to demonstrate tolerance and therapeutic action of mesenchymal stem cells from canine adipose tissue in skin grafts in rabbits.(AU)

O presente estudo teve como principal objetivo avaliar os efeitos das células-tronco mesenquimais derivadas do tecido adiposo de cães no processo de cicatrização de autoenxertos de pele de espessura total em malha em coelhos. As células-tronco foram coletadas de cães jovens, após a caracterização estas permaneceram em criopreservação, em doses individuais contendo 2 x 106 células. A técnica de enxerto em malha na região distal do membro foi realizada em 60 coelhos, divididos em três grupos, GC (Grupo Controle), GT1 (Grupo tratado com células-tronco intralesional) e GT2 (Grupo tratado com células-tronco via endovenosa), contendo 20 animais cada. Imediatamente após a enxertia, cada grupo foi dividido aleatoriamente em quatro subgrupos, de acordo com o tempo de eutanásia 3, 7, 14 e 30 dias contendo cinco animais cada. Animais dos subgrupos GT1_14, GT1_30 e GT2_14, GT2_30 receberam uma segunda dose de células xenógenas no sétimo dia. Ademais, animais do GT1_30 e do GT2_30 receberam a terceira dose de células xenógenas no dia 14. Os grupos tratados com células-tronco xenógenas tiveram um efeito positivo na reepitelização e deposição de colágeno tipo III, e possivelmente, o GT1 teve uma resposta inflamatória controlada, entretanto o efeito na angiogênese não foi observado. Dessa forma, foi possível demonstrar que houve tolerância e ação terapêutica das células-tronco mesenquimais derivadas do tecido adiposo de cães em enxertos de pele em coelhos.(AU)

Effects of canine adipose-derived mesenchymal stem cells on the epithelialization of rabbits' skin autograft (Oryctolagus cuniculus) / Efeitos das células-tronco mesenquimais derivadas do tecido adiposo de cães na epitelização de autoenxertos de pele em coelhos (Oryctolagus cuniculus)

The present study aimed to evaluate the effects of mesenchymal stem cells derived from canine adipose tissue in the healing process of full-thickness mesh skin grafts in rabbits. The stem cells were collected from young dogs; and, after characterization, remained in cryopreservation, in independent doses containing 2 x 106 cells. The mesh distal limb graft technique was performed in 60 rabbits, divided into three groups, CG (Control Group), GT1 (Intralesional Stem Cell Treated Group), and GT2 (Intravenous Stem Cell Treated Group), containing 20 animals each. After grafting, each group was randomly divided into four subgroups according to euthanasia time 3, 7, 14, and 30 days, containing five animals in each group. Animals of GT1_14, GT1_30, and GT2_14, GT2_30 subgroups received a second dose of xenogeneic cells on the seventh day. Meanwhile, animals from GT1_30 and GT2_30 received the third dose of xenogeneic cells on day 14. The groups treated with xenogeneic stem cells positively affected type III collagen re-epithelialization and deposition, and possibly GT1 had a controlled inflammatory response. However, no effect on angiogenesis. Thus, it was possible to demonstrate tolerance and therapeutic action of mesenchymal stem cells from canine adipose tissue in skin grafts in rabbits.(AU)

O presente estudo teve como principal objetivo avaliar os efeitos das células-tronco mesenquimais derivadas do tecido adiposo de cães no processo de cicatrização de autoenxertos de pele de espessura total em malha em coelhos. As células-tronco foram coletadas de cães jovens, após a caracterização estas permaneceram em criopreservação, em doses individuais contendo 2 x 106 células. A técnica de enxerto em malha na região distal do membro foi realizada em 60 coelhos, divididos em três grupos, GC (Grupo Controle), GT1 (Grupo tratado com células-tronco intralesional) e GT2 (Grupo tratado com células-tronco via endovenosa), contendo 20 animais cada. Imediatamente após a enxertia, cada grupo foi dividido aleatoriamente em quatro subgrupos, de acordo com o tempo de eutanásia 3, 7, 14 e 30 dias contendo cinco animais cada. Animais dos subgrupos GT1_14, GT1_30 e GT2_14, GT2_30 receberam uma segunda dose de células xenógenas no sétimo dia. Ademais, animais do GT1_30 e do GT2_30 receberam a terceira dose de células xenógenas no dia 14. Os grupos tratados com células-tronco xenógenas tiveram um efeito positivo na reepitelização e deposição de colágeno tipo III, e possivelmente, o GT1 teve uma resposta inflamatória controlada, entretanto o efeito na angiogênese não foi observado. Dessa forma, foi possível demonstrar que houve tolerância e ação terapêutica das células-tronco mesenquimais derivadas do tecido adiposo de cães em enxertos de pele em coelhos.(AU)

Evaluation of platelet-rich plasma gel as an angiogenesis-inducing agent in canine advancement skin flaps / Avaliação do gel de plasma rico em plaquetas como agente indutor da angiogênese em flapes cutâneos de avanço em cães

This work aimed to evaluate the effect of platelet-rich plasma (PRP) on advancement skin flaps in dogs regarding improvement of vascularization, with focus on increasing its viable area, since there are reports that it is a potential angiogenesis stimulator. The experimental group was composed of eight adult bitches, in which two advancement skin flaps were made in the ventral abdominal region. No product was applied in the control flap (CF), while PRP was used in the contralateral flap, called treated flap (TF). The areas were clinically evaluated every two days until the 7th postoperative day regarding skin color and presence of necrosis. At 10 days, both flaps were removed and submitted to histological examination and blood vessel morphometry. The vessels counted in each group were statistically analyzed by the F-test at 1% probability. Results showed no significant difference in macroscopic changes in the wound, or CF and TF vascularization, thus suggesting that PRP gel did not improve advancement skin flap angiogenesis in bitches under the experimental conditions in which this research was developed.(AU)

Objetivou-se com o presente artigo avaliar a ação angiogênica do gel de plasma rico em plaquetas (PRP) em flapes cutâneos de avanço em animais da espécie canina, visando aumentar a viabilidade da pele após o procedimento, uma vez que existem relatos de que o produto é um potente estimulador da angiogênese. O grupo experimental foi composto por oito cadelas adultas, onde foram confeccionados dois flapes de avanço (de padrão subdérmico) na região abdominal ventral. Em um dos flapes, considerado controle (FC) não foi aplicado nenhum produto, enquanto que no flape contralateral, denominado tratado (FT), foi usado o gel de PRP. As áreas foram macroscopicamente avaliadas a cada dois dias até o 7º dia de pós-operatório em relação à coloração da pele e presença de área de necrose, e com 10 dias ambos os flapes foram coletados por biópsia e submetidos ao exame histológico e morfometria dos vasos sanguíneos. Os vasos contados em cada grupo foram estatisticamente analisados pelo teste de F ao nível de 1% de probabilidades. Os resultados demonstraram que não houve diferença significativa nas alterações macroscópicas das feridas e na morfometria vascular dos FC e FT, sugerindo dessa maneira que dentro das condições experimentais nas quais a pesquisa foi executada, que o gel de PRP não incrementou a angiogênese de flapes de avanço em cadelas.(AU)

Vaccarin hastens wound healing by promoting angiogenesis via activation of MAPK/ERK and PI3K/AKT signaling pathways in vivo

Purpose To explore the potential role and unclear molecular mechanisms of vaccarin in wound healing. Methods Rats skin excision model to study the effects of vaccarin on wound healing in vivo . Hematoxylin and eosin staining was performed to evaluate Histopathologic characteristics. Immunohistochemistry was employed to assess the effects of vaccarin in accelerating angiogenesis. Western blot was used to evaluate relative protein expressed levels. Results Vaccarin could significantly promote wound healing and endothelial cells and fibroblasts proliferation in the wound site. Immunohistochemistry and Western blot studies showed that the nodal proteins and receptor (bFGFR) related to angiogenesis signaling pathway were activated, and the microvascular density in the wound site was markedly higher than that in the control group. Conclusions The present study was the first to demonstrate that vaccarin is able to induce angiogenesis and accelerate wound healing in vivo by increasing expressions of p-Akt, p-Erk and p-bFGFR. This process is mediated by MAPK/ERK and PI3K/AKT signaling pathways.(AU)

Microvessel density assessment in canine mammary carcinomas as a predictive factor for metronomic chemotherapy / Mensuração da densidade microvascular em carcinomas mamários caninos como fator preditivo à quimioterapia metronômica

The canine mammary tumor is the neoplasia that most commonly affects intact female dogs (not spayed) in routine veterinary practice. Canine mammary tumor therapy is a challenge because only few effective treatments have been described for high-grade tumors in veterinary medicine. One such therapeutic option that slows down tumor growth is metronomic chemotherapy (MC), a therapeutic modality that acts by decreasing tumor angiogenesis. Quantification of intratumoral microvessel density (MVD) has been proposed as a means to evaluate the effectiveness of MC in human and canine tumors. In this study, MVD is proposed as a predictive factor for the effectiveness of MC in canine mammary tumors. Twenty female dogs with mammary carcinoma were equally distributed into a mastectomy group treated only with mastectomy, and a MC group treated with MC (cyclophosphamide and piroxicam) orally and daily for 28 days followed by mastectomy. Mammary tumors were classified and graded histologically. MVD was ascertained by CD31 immunostaining. The analysis showed statistically significant difference in MVD scores between groups, corroborating a quantitative reduction in tumor microvasculature in the group treated with MC. Our findings suggest that MVD may be an important predictive factor for the selection of female dogs with malignant mammary tumors that may benefit from MC.(AU)

O tumor mamário canino é a neoplasia que mais comumente acomete cadelas não castradas na rotina veterinária. A terapia para a neoplasia mamária canina é um desafio, com poucos tratamentos eficazes descritos em medicina veterinária, especialmente para as neoplasias de alto grau. Uma opção terapêutica que diminui o crescimento tumoral é a quimioterapia metronômica (QM). A QM é uma modalidade terapêutica que age pelo decréscimo da angiogênese tumoral. Para avaliação da eficácia da QM tem sido proposta a quantificação da densidade microvascular (DMV) em neoplasias caninas. Neste estudo foi proposta a mensuração da DMV como fator preditivo à utilização da QM em neoplasias mamárias caninas. Foram utilizadas 20 (vinte) cadelas com carcinomas mamários malignos sendo um grupo mastectomia, tratado somente por mastectomia, e um grupo QM, em que foi empregada a QM (ciclofosfamida e piroxicam) ambos oralmente e de uso diário por 28 dias seguida de mastectomia. As neoplasias mamárias foram histologicamente classificadas e graduadas. A DMV foi obtida por imunomarcação utilizando o anticorpo CD31. A análise estatística mostrou diferença na DMV entre os grupos, evidenciando redução quantitativa na microvasculatura tumoral no grupo tratado com quimioterapia metronômica. Com base neste resultado é possível afirmar que a DMV pode vir a ser um fator preditivo de importância para a seleção de cadelas portadoras de neoplasia mamária maligna que possam vir a se beneficiar da quimioterapia metronômica.(AU)

Microvessel density assessment in canine mammary carcinomas as a predictive factor for metronomic chemotherapy / Mensuração da densidade microvascular em carcinomas mamários caninos como fator preditivo à quimioterapia metronômica

The canine mammary tumor is the neoplasia that most commonly affects intact female dogs (not spayed) in routine veterinary practice. Canine mammary tumor therapy is a challenge because only few effective treatments have been described for high-grade tumors in veterinary medicine. One such therapeutic option that slows down tumor growth is metronomic chemotherapy (MC), a therapeutic modality that acts by decreasing tumor angiogenesis. Quantification of intratumoral microvessel density (MVD) has been proposed as a means to evaluate the effectiveness of MC in human and canine tumors. In this study, MVD is proposed as a predictive factor for the effectiveness of MC in canine mammary tumors. Twenty female dogs with mammary carcinoma were equally distributed into a mastectomy group treated only with mastectomy, and a MC group treated with MC (cyclophosphamide and piroxicam) orally and daily for 28 days followed by mastectomy. Mammary tumors were classified and graded histologically. MVD was ascertained by CD31 immunostaining. The analysis showed statistically significant difference in MVD scores between groups, corroborating a quantitative reduction in tumor microvasculature in the group treated with MC. Our findings suggest that MVD may be an important predictive factor for the selection of female dogs with malignant mammary tumors that may benefit from MC.

O tumor mamário canino é a neoplasia que mais comumente acomete cadelas não castradas na rotina veterinária. A terapia para a neoplasia mamária canina é um desafio, com poucos tratamentos eficazes descritos em medicina veterinária, especialmente para as neoplasias de alto grau. Uma opção terapêutica que diminui o crescimento tumoral é a quimioterapia metronômica (QM). A QM é uma modalidade terapêutica que age pelo decréscimo da angiogênese tumoral. Para avaliação da eficácia da QM tem sido proposta a quantificação da densidade microvascular (DMV) em neoplasias caninas. Neste estudo foi proposta a mensuração da DMV como fator preditivo à utilização da QM em neoplasias mamárias caninas. Foram utilizadas 20 (vinte) cadelas com carcinomas mamários malignos sendo um grupo mastectomia, tratado somente por mastectomia, e um grupo QM, em que foi empregada a QM (ciclofosfamida e piroxicam) ambos oralmente e de uso diário por 28 dias seguida de mastectomia. As neoplasias mamárias foram histologicamente classificadas e graduadas. A DMV foi obtida por imunomarcação utilizando o anticorpo CD31. A análise estatística mostrou diferença na DMV entre os grupos, evidenciando redução quantitativa na microvasculatura tumoral no grupo tratado com quimioterapia metronômica. Com base neste resultado é possível afirmar que a DMV pode vir a ser um fator preditivo de importância para a seleção de cadelas portadoras de neoplasia mamária maligna que possam vir a se beneficiar da quimioterapia metronômica.

Effect of red propolis on hamster cheek pouch angiogenesis in a new sponge implant model

Purpose: To evaluate the effects of red propolis on cheek pouch angiogenesis in a hamster new model sponge implant. Methods: Forty eight animals divided into eight groups. (Groups I-IV), the animals were treated for 15 days before and 10 days after sponge implantation. (Groups V-VIII), the animals were treated for 10 days after sponge implantation (GI and GV: red propolis 100 mg/kg, GII and GVI: celecoxib 20 mg/kg, GIII and GVII: 1% gum arabic 5 mL/kg, GIV and GVIII: distilled water 5 mL/kg). On the 11th day of implantation, the animals were anesthetized for stereoscopic microscopic imaging and morphometric quantification of angiogenesis (SQAN), followed by histopathological evaluation (H&E). Results: In the SQAN analysis, no significant difference was found between the groups. However, on histology, propolis was found reduce the population of mastocytes in the qualitative analyses (p = 0,013) in the quantitative analyses to reduce the number of blood vessels (p = 0,007), and increase the macrophage count (p = 0,001). Conclusion: Red propolis inhibited inflammatory angiogenesis when administered before andcontinuously after sponge implant, and was shown to have immunomodulating effects on inflammatory cells (mastocytes and macrophages) in a new sponge implant hamster model.(AU)

Simazine, a triazine herbicide, disrupts swine granulosa cell functions

The triazine herbicide simazine is a pesticide commonly detected in surface and ground waters, although banned in most European countries since 2004. Concerns for humans and animal health result from its potential endocrine disrupting action, that can lead to reproductive disorders. The present in vitro study was undertaken to study simazine effects on swine granulosa cell function, namely cell viability, proliferation, steroidogenesis and NO production. Moreover, the ability of this substance to interfere with the angiogenetic process, a crucial event in reproductive function, was taken into account. Our data document that simazine treatment, at 0.1 or 10 μM concentration levels, stimulates granulosa cell proliferation and viability and impairs steroidogenesis, increasing in particular progesterone production. In addition, the in vitro angiogenesis bioassay revealed a significant simazine stimulatory effect on immortalized porcine Aortic Endothelial Cell proliferation. Collectively, these results show that simazine can display disruptive effects on ovarian cell functional parameters, possibly resulting in reproductive dysfunction. This hypothesis is also supported by the observed pro-angiogenetic properties of this herbicide, as already suggested for different endocrine disruptors.

Membrana de látex natural de Hevea brasiliensis auxilia no processo de reparação tecidual em bovinos / Natural latex rubber from Hevea brasiliensis aids tissue repair of bovine

Feridas cutâneas em bovinos são um constante desafio clínico cirúrgico por desencadearem perdas econômicas bastante significativas. O látex proveniente da seiva da seringueira (Hevea brasiliensis) apresenta potencial terapêutico para incrementar o processo de reparação tecidual. Portanto, pretendeu-se com esse estudo avaliar o tipo de reação tecidual e os possíveis mecanismos de angiogênese desencadeados pelo implante de uma membrana de látex natural em bovinos. Para tal, foram utilizados seis bovinos da raça Nelore, submetidos ao implante subcutâneo experimental de três fragmentos de membranas de látex natural. Foram coletadas amostras de tecido e da membrana aos 15, 30 e 45 dias após a implantação, para avaliações histológicas, ultraestruturais por microscopia eletrônica de varredura e imunoistoquímicas com anticorpos antimarcador de macrófagos (MAC), CYR 61 e VEGF. O implante de látex proporcionou aumento da angiogênese e reparação tecidual em bovinos, não mediada pela expressão do VEGF e CYR 61.(AU)

Cattle wounds are a constant surgical and clinical challenge, leading to important economical losses. The latex from the sap of the rubber tree (Hevea brasiliensis) has therapeutic potential to enhance tissue repair process. Therefore, we evaluated the type of tissue reaction and possible mechanisms of angiogenesis triggered by implanting natural latex rubber in bovine species. Six Nelore bovines were subjected to subcutaneous experimental implant of three fragments of natural rubber latex membranes. Tissue and rubber membrane samples were harvested at 15, 30 and 45 days implantation for histology, scanning electron microscopy and immunohistochemical evaluation with anti macrophage marker (MAC), anti CYR 61, anti VEGF antibodies. The latex membrane estimulates tissue reaction and repair and significant angiogenesis stimuli without activating CYR 61 and VEGF pathways.(AU)

Simazine, a triazine herbicide, disrupts swine granulosa cell functions

The triazine herbicide simazine is a pesticide commonly detected in surface and ground waters, although banned in most European countries since 2004. Concerns for humans and animal health result from its potential endocrine disrupting action, that can lead to reproductive disorders. The present in vitro study was undertaken to study simazine effects on swine granulosa cell function, namely cell viability, proliferation, steroidogenesis and NO production. Moreover, the ability of this substance to interfere with the angiogenetic process, a crucial event in reproductive function, was taken into account. Our data document that simazine treatment, at 0.1 or 10 μM concentration levels, stimulates granulosa cell proliferation and viability and impairs steroidogenesis, increasing in particular progesterone production. In addition, the in vitro angiogenesis bioassay revealed a significant simazine stimulatory effect on immortalized porcine Aortic Endothelial Cell proliferation. Collectively, these results show that simazine can display disruptive effects on ovarian cell functional parameters, possibly resulting in reproductive dysfunction. This hypothesis is also supported by the observed pro-angiogenetic properties of this herbicide, as already suggested for different endocrine disruptors.(AU)