Efficacy of acidified phytase supplemented cottonseed meal based diets on growth performance and proximate composition of Labeo rohita fingerlings

Abstract The growth of aquaculture sector is strongly dependent upon the continuous supply of inexpensive fish feed with balanced nutritional profile. However, fish meal (FM) is unable to satisfy this demand due to its scarce supply and high cost. In order to test the potential of cottonseed meal (CSM) as a fish meal replacer, a feeding trial of 12 weeks was conducted to check growth performance and proximate composition of Labeo rohita fingerlings. The protein ration of the test feed was satisfied by replacing FM with CSM at 0, 25, 50 and 75%. Sixteen test diets viz., TD1 (control), TD2, TD3, TD4, TD5, TD6, TD7, TD8, TD9, TD10, TD11, TD12, TD13, TD14, TD15 and TD16 were supplemented with citric acid (CA; 0 and 2.5%) and phytase (PHY; 0 and 750 FTU/kg) in a completely randomized design with 3×3 factorial arrangement. The highest weight gain (11.03g), weight gain% (249.21%), specific growth rate (1.39) and best feed conversion ratio (1.20) were recorded by fish fed with TD12. Furthermore, the same level increased the crude protein (59.26%) and fat (16.04%) being significantly different (p 0.05) than that of control. Conclusively, the addition of acidified phytase (CA; 2.5%, PHY; 750 FTU/kg) in TD12 (CSM=50%) led to the improved growth and proximate composition of L. rohita fingerlings.

Resumo O crescimento do setor de aquicultura é fortemente dependente do fornecimento contínuo de rações baratas para peixes com perfil nutricional equilibrado. Porém, a farinha de peixe (FM) não consegue atender a essa demanda devido à sua escassa oferta e alto custo. Com o objetivo de testar o potencial da farinha de semente de canola (MSC) como substituto da farinha de peixe, um ensaio alimentar de 12 semanas foi conduzido para verificar o desempenho de crescimento e a composição centesimal de alevinos de Labeo rohita. A ração de proteína da ração teste foi satisfeita substituindo FM por CSM em 0, 25, 50 e 75%. Dezesseis dietas de teste, viz., TD1 (controle), TD2, TD3, TD4, TD5, TD6, TD7, TD8, TD9, TD10, TD11, TD12, TD13, TD14, TD15 e TD16 foram suplementadas com ácido cítrico (CA; 0 e 2,5%) e fitase (PHY; 0 e 750 FTU / kg) em um delineamento inteiramente casualizado com arranjo fatorial 3 × 3. O maior ganho de peso (11,03g), % de ganho de peso (249,21%), taxa de crescimento específico (1,39) e melhor taxa de conversão alimentar (1,20) foram registrados por peixes alimentados com TD12. Além disso, o mesmo nível aumentou a proteína bruta (59,26%) e a gordura (16,04%), sendo significativamente diferente (p 0,05) do controle. Conclusivamente, a adição de fitase acidificada (CA; 2,5%, PHY; 750 FTU / kg) em TD12 (CSM = 50%) levou a um melhor crescimento e composição próxima de alevinos de L. rohita.

The effect of Cissus quadrangularis Salisb. extract on maturation of rat mesenchymal stem cells / O efeito do extrato de Cissus quadrangularis Salisb. na proliferação e diferenciação de células-tronco mesenquimais de rato

Degenerative diseases, such as osteoporosis, could be treated by stem cells. The aim of this study was to identify the gene expression of bone marrow mesenchymal stem cells (BM-MSC) derived from Sprague Dawley rats and to assess the effect of Cissus quadrangularis Salisb. extract on their maturation into bone cells. The BM-MSC were divided into three groups: (a) BM-MSCs + osteoblast cell growth basal medium as the positive control; (b) BM-MSCs + Dulbecco's modified eagle's medium (DMEM) + 0.3 mg/mL methanol extract of C. quadrangularis as methanol group; and (c) BM-MSC + DMEM + 0.3 mg/mL ethyl acetate extract of C. quadrangularis as ethyl acetate group. A relative quantification approach using was used to analyze the expression of the alp (alkaline phosphatase) gene, with the beta-actin gene was used to normalize the expression of the alp gene. The intra-assay variation was calculated to validate the RT-qPCR data. Our study found that the intra-assay variation value was acceptable, with most of the coefficients of variability (CV) value <5. Ethyl acetate solvent outperformed methanol solvent in extracting the active compound C. quadrangularis. In the ethyl acetate extract group, the expression of the alp gene increased three times compared to the positive control. In methanol extract group, the expression of alp gene was lower six times compared to positive control. This study suggests that C. quadrangularis extracts using ethyl acetate could induce the maturation of BM-MSCs. However, further studies are warrant to confirm this effect using different indicators.

Doenças degenerativas, como a osteoporose, podem ser tratadas por células-tronco. O objetivo deste estudo foi identificar a expressão gênica de células-tronco mesenquimais da medula óssea (BM-MSCs) derivadas de ratos Sprague Dawley, bem como sua proliferação e diferenciação em células ósseas afetadas pelo extrato de Cissus quadrangularis Salisb. As BM-MSCs foram divididas em três grupos: (1) BM-MSCs + meio basal de crescimento de osteoblastos como controle positivo; (2) BM-MSCs + meio de Eagle modificado por Dulbecco (DMEM) + extrato de metanol; e (3) BM-MSCs + DMEM + extrato de acetato de etila de C. quadrangularis. Uma abordagem de quantificação relativa foi usada para analisar a expressão do gene alp (fosfatase alcalina), com o gene da beta-actina sendo usado para normalizar a expressão do gene alp em cada tratamento. A variação intraensaio foi calculada para validar os dados de RT-qPCR. Os resultados mostraram que o valor da variação intraensaio é aceitável, com a maioria dos valores dos coeficientes de variabilidade (CV) < 5. Além disso, no tratamento T2, a expressão do gene alp aumentou três vezes em relação ao controle positivo. No tratamento T1, o gene alp apresentou nível de expressão seis vezes menor em comparação com o controle positivo. Além disso, o solvente acetato de etila superou o solvente metanol na extração do composto ativo C. quadrangularis Salisb. Este estudo demonstra que extratos de C. quadrangularis Salisb., particularmente com acetato de etila como solvente, podem induzir a proliferação e diferenciação de BM-MSCs. Com base na expressão do gene alp, C. quadrangularis tem o potencial de corrigir a perda óssea.

Efficacy of acidified phytase supplemented cottonseed meal based diets on growth performance and proximate composition of Labeo rohita fingerlings / Eficácia de dietas à base de algodão com fitase acidificada suplementada no desempenho do crescimento e na composição proximada de dedos de Labeo rohita

The growth of aquaculture sector is strongly dependent upon the continuous supply of inexpensive fish feed with balanced nutritional profile. However, fish meal (FM) is unable to satisfy this demand due to its scarce supply and high cost. In order to test the potential of cottonseed meal (CSM) as a fish meal replacer, a feeding trial of 12 weeks was conducted to check growth performance and proximate composition of Labeo rohita fingerlings. The protein ration of the test feed was satisfied by replacing FM with CSM at 0, 25, 50 and 75%. Sixteen test diets viz., TD1 (control), TD2, TD3, TD4, TD5, TD6, TD7, TD8, TD9, TD10, TD11, TD12, TD13, TD14, TD15 and TD16 were supplemented with citric acid (CA; 0 and 2.5%) and phytase (PHY; 0 and 750 FTU/kg) in a completely randomized design with 3×3 factorial arrangement. The highest weight gain (11.03g), weight gain% (249.21%), specific growth rate (1.39) and best feed conversion ratio (1.20) were recorded by fish fed with TD12. Furthermore, the same level increased the crude protein (59.26%) and fat (16.04%) being significantly different (p<0.05) than that of control. Conclusively, the addition of acidified phytase (CA; 2.5%, PHY; 750 FTU/kg) in TD12 (CSM=50%) led to the improved growth and proximate composition of L. rohita fingerlings.

O crescimento do setor de aquicultura é fortemente dependente do fornecimento contínuo de rações baratas para peixes com perfil nutricional equilibrado. Porém, a farinha de peixe (FM) não consegue atender a essa demanda devido à sua escassa oferta e alto custo. Com o objetivo de testar o potencial da farinha de semente de canola (MSC) como substituto da farinha de peixe, um ensaio alimentar de 12 semanas foi conduzido para verificar o desempenho de crescimento e a composição centesimal de alevinos de Labeo rohita. A ração de proteína da ração teste foi satisfeita substituindo FM por CSM em 0, 25, 50 e 75%. Dezesseis dietas de teste, viz., TD1 (controle), TD2, TD3, TD4, TD5, TD6, TD7, TD8, TD9, TD10, TD11, TD12, TD13, TD14, TD15 e TD16 foram suplementadas com ácido cítrico (CA; 0 e 2,5%) e fitase (PHY; 0 e 750 FTU / kg) em um delineamento inteiramente casualizado com arranjo fatorial 3 × 3. O maior ganho de peso (11,03g), % de ganho de peso (249,21%), taxa de crescimento específico (1,39) e melhor taxa de conversão alimentar (1,20) foram registrados por peixes alimentados com TD12. Além disso, o mesmo nível aumentou a proteína bruta (59,26%) e a gordura (16,04%), sendo significativamente diferente (p <0,05) do controle. Conclusivamente, a adição de fitase acidificada (CA; 2,5%, PHY; 750 FTU / kg) em TD12 (CSM = 50%) levou a um melhor crescimento e composição próxima de alevinos de L. rohita.

Efficacy of acidified phytase supplemented cottonseed meal based diets on growth performance and proximate composition of Labeo rohita fingerlings / Eficácia de dietas à base de algodão com fitase acidificada suplementada no desempenho do crescimento e na composição proximada de dedos de Labeo rohita

Abstract The growth of aquaculture sector is strongly dependent upon the continuous supply of inexpensive fish feed with balanced nutritional profile. However, fish meal (FM) is unable to satisfy this demand due to its scarce supply and high cost. In order to test the potential of cottonseed meal (CSM) as a fish meal replacer, a feeding trial of 12 weeks was conducted to check growth performance and proximate composition of Labeo rohita fingerlings. The protein ration of the test feed was satisfied by replacing FM with CSM at 0, 25, 50 and 75%. Sixteen test diets viz., TD1 (control), TD2, TD3, TD4, TD5, TD6, TD7, TD8, TD9, TD10, TD11, TD12, TD13, TD14, TD15 and TD16 were supplemented with citric acid (CA; 0 and 2.5%) and phytase (PHY; 0 and 750 FTU/kg) in a completely randomized design with 3×3 factorial arrangement. The highest weight gain (11.03g), weight gain% (249.21%), specific growth rate (1.39) and best feed conversion ratio (1.20) were recorded by fish fed with TD12. Furthermore, the same level increased the crude protein (59.26%) and fat (16.04%) being significantly different (p<0.05) than that of control. Conclusively, the addition of acidified phytase (CA; 2.5%, PHY; 750 FTU/kg) in TD12 (CSM=50%) led to the improved growth and proximate composition of L. rohita fingerlings.

Resumo O crescimento do setor de aquicultura é fortemente dependente do fornecimento contínuo de rações baratas para peixes com perfil nutricional equilibrado. Porém, a farinha de peixe (FM) não consegue atender a essa demanda devido à sua escassa oferta e alto custo. Com o objetivo de testar o potencial da farinha de semente de canola (MSC) como substituto da farinha de peixe, um ensaio alimentar de 12 semanas foi conduzido para verificar o desempenho de crescimento e a composição centesimal de alevinos de Labeo rohita. A ração de proteína da ração teste foi satisfeita substituindo FM por CSM em 0, 25, 50 e 75%. Dezesseis dietas de teste, viz., TD1 (controle), TD2, TD3, TD4, TD5, TD6, TD7, TD8, TD9, TD10, TD11, TD12, TD13, TD14, TD15 e TD16 foram suplementadas com ácido cítrico (CA; 0 e 2,5%) e fitase (PHY; 0 e 750 FTU / kg) em um delineamento inteiramente casualizado com arranjo fatorial 3 × 3. O maior ganho de peso (11,03g), % de ganho de peso (249,21%), taxa de crescimento específico (1,39) e melhor taxa de conversão alimentar (1,20) foram registrados por peixes alimentados com TD12. Além disso, o mesmo nível aumentou a proteína bruta (59,26%) e a gordura (16,04%), sendo significativamente diferente (p <0,05) do controle. Conclusivamente, a adição de fitase acidificada (CA; 2,5%, PHY; 750 FTU / kg) em TD12 (CSM = 50%) levou a um melhor crescimento e composição próxima de alevinos de L. rohita.

Efficacy of acidified phytase supplemented cottonseed meal based diets on growth performance and proximate composition of Labeo rohita fingerlings / Eficácia de dietas à base de algodão com fitase acidificada suplementada no desempenho do crescimento e na composição proximada de dedos de Labeo rohita

The growth of aquaculture sector is strongly dependent upon the continuous supply of inexpensive fish feed with balanced nutritional profile. However, fish meal (FM) is unable to satisfy this demand due to its scarce supply and high cost. In order to test the potential of cottonseed meal (CSM) as a fish meal replacer, a feeding trial of 12 weeks was conducted to check growth performance and proximate composition of Labeo rohita fingerlings. The protein ration of the test feed was satisfied by replacing FM with CSM at 0, 25, 50 and 75%. Sixteen test diets viz., TD1 (control), TD2, TD3, TD4, TD5, TD6, TD7, TD8, TD9, TD10, TD11, TD12, TD13, TD14, TD15 and TD16 were supplemented with citric acid (CA; 0 and 2.5%) and phytase (PHY; 0 and 750 FTU/kg) in a completely randomized design with 3×3 factorial arrangement. The highest weight gain (11.03g), weight gain% (249.21%), specific growth rate (1.39) and best feed conversion ratio (1.20) were recorded by fish fed with TD12. Furthermore, the same level increased the crude protein (59.26%) and fat (16.04%) being significantly different (p<0.05) than that of control. Conclusively, the addition of acidified phytase (CA; 2.5%, PHY; 750 FTU/kg) in TD12 (CSM=50%) led to the improved growth and proximate composition of L. rohita fingerlings.(AU)

O crescimento do setor de aquicultura é fortemente dependente do fornecimento contínuo de rações baratas para peixes com perfil nutricional equilibrado. Porém, a farinha de peixe (FM) não consegue atender a essa demanda devido à sua escassa oferta e alto custo. Com o objetivo de testar o potencial da farinha de semente de canola (MSC) como substituto da farinha de peixe, um ensaio alimentar de 12 semanas foi conduzido para verificar o desempenho de crescimento e a composição centesimal de alevinos de Labeo rohita. A ração de proteína da ração teste foi satisfeita substituindo FM por CSM em 0, 25, 50 e 75%. Dezesseis dietas de teste, viz., TD1 (controle), TD2, TD3, TD4, TD5, TD6, TD7, TD8, TD9, TD10, TD11, TD12, TD13, TD14, TD15 e TD16 foram suplementadas com ácido cítrico (CA; 0 e 2,5%) e fitase (PHY; 0 e 750 FTU / kg) em um delineamento inteiramente casualizado com arranjo fatorial 3 × 3. O maior ganho de peso (11,03g), % de ganho de peso (249,21%), taxa de crescimento específico (1,39) e melhor taxa de conversão alimentar (1,20) foram registrados por peixes alimentados com TD12. Além disso, o mesmo nível aumentou a proteína bruta (59,26%) e a gordura (16,04%), sendo significativamente diferente (p <0,05) do controle. Conclusivamente, a adição de fitase acidificada (CA; 2,5%, PHY; 750 FTU / kg) em TD12 (CSM = 50%) levou a um melhor crescimento e composição próxima de alevinos de L. rohita.(AU)

Secretion pattern of canine amniotic stem cells derived extracellular vesicles

Extracellular vesicles (EVs) derived from stem cells (SCs) have regenerative potential and the possibility of being used in treating chronic diseases. EVs present lower risk of tumorigenicity and easily to isolation and storage. Therefore, this research aims to compare the morphological characteristics of the EVs (up to 150nm) derived from stem cells obtained from canine amniotic membranes in different passages during the in vitro culture. For this, cells from the amniotic membranes were isolated, cultured, and characterized. In order to answer our aim, the number of cells was normalized at each passage to generate conditioned media for EVs separation. The cells were differentiated into adipogenic, chondrogenic, and osteogenic tissue, to characterize these cells as mesenchymal stem cells (MSC). Moreover, flow cytometry analysis was performed and showed that the MSC were positive for CD90, CD105 and negative for CD34, CD45, mesenchymal and hematopoietic markers, respectively. For EVs analysis, MSC in different passages (P0-P2) were culture until 80% of confluence, then the medium was replaced by EVs depleted medium. After 48h, culture medium was collected and centrifuged to separate EVs, followed by nanoparticle tracking analysis. The EVs were also characterized by western blot and transmission electron microscopy (TEM). EVs were positive for Alix and negative for Cytochrome C as well as presented the traditional cup-shape by transmission electronic microscopy. Our results demonstrated that the concentration in the different passages was increased in P0 compared to P1 and P2 (p<0.05). No differences were found in EVs size (P0=132nm, P1=130nm and P2=120nm). Together, these results demonstrate that P0 of MSC is enriched of EVs when compared to later passages, suggesting that this passage would be the best to be applied in pre-clinical tests. Despite that, more studies are necessary to identify the EVs content and how the cells will respond to treatment with them.(AU)

Evaluation of mesenchymal cells and dapsone for the treatment of dermonecrotic wounds caused by Loxosceles laeta venom in rabbits / Avaliação de células mesenquimais e dapsona para o tratamento de feridas dermonecróticas causadas pelo veneno de Loxosceles laeta em coelhos

This study aimed to evaluate the efficacy of mesenchymal stem cells (MSC), alone or associated with dapsone (DAP), in treating dermonecrotic wounds caused by Loxosceles laeta venom. Twenty-five male rabbits were distributed into five groups. Negative control received ultrapure water (C-), whilst all other groups were injected with 20 μg of L. laeta venom. After 4 hours, each group received one of the following treatments: PBS (C+), DAP, MSC, and DAP+MSC. Animals were evaluated daily and photographic records made for analysis of wound area. Twelve days after, animals were euthanized and skin samples removed for histological analysis. We observed that DAP showed the best percentage of wound contraction at day 3. In the treatments using MSCs, a negative value of wound contraction was observed for the isolated MSCs, as well as a lower contraction value for the association of the MSC + DAP when compared to PBS, probably, by the increase in initial infammation after the application of stem cells, due to the fact that MSCs secrete a broad spectrum of bioactive molecules such as cytokines and growth factors that favor regeneration. Histologically, it was observed that animals of C+ showed extensive areas of necrosis, ulcers, neutrophilic infiltrate, and mineralization. Collagen deposition showed increase in MSC+DAP treatment, however vascularization remained unchanged. This is the first report using MSC and MSC+DAP as a treatment for cutaneous loxoscelism and more studies are needed to determine its use as an alternative therapy for dermonecrotic lesions caused by Loxosceles spider.

Este estudo teve como objetivo avaliar a eficácia das células-tronco mesenquimais (CTMs), isoladas ou associadas à dapsona (DAP), no tratamento de feridas dermonecróticas causadas pelo veneno de Loxosceles laeta. Vinte e cinco coelhos machos foram distribuídos em cinco grupos. O controle negativo recebeu água ultrapura (C-), enquanto todos os outros grupos foram injetados com 20 μg de veneno de L. laeta. Após 4 horas, cada grupo recebeu um dos seguintes tratamentos: PBS (C+), DAP, CTMs e DAP + CTMs. Os animais foram avaliados diariamente durante 12 dias, e feitos registros fotográficos para análise da ferida e no 12º dia, foram eutanasiados e, retiradas amostras de pele para análise histológica. Observou-se que a DAP apresentou o melhor percentual de contração da ferida no terceiro dia. Nos tratamentos com CTMs, observou-se uma contração negativa da ferida tanto para as CTMs isoladas, bem como a associação CTMs + DAP em relação ao PBS, possivelmente, pelo aumento da infamação inicial após a aplicação de células-tronco. Isso é devido ao fato de que as CTMs secretam um amplo espectro de moléculas bioativas como citocinas e fatores de crescimento que favorecem a regeneração. Histologicamente, observou-se que os animais de C+ apresentaram extensas áreas de necrose, úlceras, infiltrado neutrofílico, além de mineralização. Houve aumento de deposição de colágeno no tratamento CTMs + DAP, no entanto, a vascularização permaneceu inalterada. Este é o primeiro relato usando CTMs e CTMs + DAP como tratamento para loxoscelismo cutâneo e mais estudos são necessários para determinar seu uso como terapia alternativa para lesões demonecróticas causadas pela aranha Loxosceles.

Células-tronco mesenquimais para tratamento de displasia coxofemoral em cão / Mesenchymal stem cells for treatment of coxofemoral dysplasia in a dog

Background: Coxofemoral dysplasia (CFD) is the abnormal development of the hip joint, mostly affecting large breeds, and is characterized by subluxation or complete luxation of the femoral head. Among the conservative therapeutic options, cell therapies with stem cells for CFD provides potential by the countless possibilities of therapeutic application, especially those related to the chronic and/or degenerative diseases treatment, which could be a key point for the bone and joint repair. The objective was to report a bilateral CFD case in a dog, treated with intra-articular application of mesenchymal stem cells (MSC), with 0, 30, 60 and 90 days, and further evaluations. Case: A 2-year-old male Akita dog was referred to the Veterinary Hospital Harmonia (HVH) located in the city of Recife, Pernambuco, presenting clinical signs of hip pain, lameness and pelvic limbs hopping. By orthopedic examination, pain was observed at the cranial and caudal extension of the pelvic limb, and at flexion, abduction and adduction of the limb, as well as moderate muscle atrophy and presence of joint crackling. By coxofemoral radiography performed in ventrodorsal projection, it were detected bilateral femoral head subluxation, thickening of the femoral neck and the presence of intraarticular osteophytes. These findings are compatible with the degeneration caused by the presence of CFD. Laboratory tests performed such as hemogram and biochemical had no changes. Thus, the patient had intra-articular application of allogeneic MSC, derived from adipose tissue, obtained by private company. For stem cell applications, arthrocentesis of the hip joints was performed one at a time, using a 5 mL syringe and 16G needle for aspiration of 2 mL synovial fluid, for discard and application of stem cells. After aspiration, the syringe containing the MSC were fitted to the 16G needle for the implant. After the...

Efficacy of antimicrobial therapy in association with vaccination on the bacteriological cure of subclinical mastitis caused by Staphylococcus aureus in lactating cows / Eficácia da terapia antimicrobiana associada à vacinação na cura bacteriológica de mastite subclínica causada por Staphylococcus aureus em vacas em lactação

The objective of this study was to evaluate the efficacy of the antibiotic therapy associated with vaccination on the microbiological cure rate of subclinical mastitis caused by Staphylococcus aureus in lactating dairy cows. A total of five herds, from which 72 cows (120 mammary quarters - MQ) were diagnosed with S. aureus subclinical mastitis, were included in this study. Cows were randomly allocated to one of three treatment groups: a) Control (no treatment); b) ATB (antibiotic therapy); and c) ATB+VAC (antibiotic therapy plus vaccination against S. aureus). Intramammary treatment consisted of twice-daily infusion of ampicillin 75mg + cloxacillin 200mg, for 5 days. Parenteral treatment was done by injection of a single dose (7.5mg/kg) of enrofloxacin, on the first day of the treatment protocol. Vaccinated cows received three doses of a commercial vaccine 14 days before treatment (d-14), on the first day of treatment protocol (d1), and 14 days after the treatment protocol (d+14). Non-treated cows had a lower cure rate (0.06) than cows treated with ATB (0.84) and ATB+VAC (0.85). No difference in cure rate was observed between cows treated with ATB and ATB+VAC. On the other hand, vaccinated cows had lower somatic cell count (SCC) after 28 days of the treatment protocols (4.76 log10) than non-treated cows (5.37 log10). In conclusion, treatment with intramammary ampicillin and cloxacillin, associated with intramuscular enrofloxacin presented a high cure rate for SCM caused by S. aureus during lactation. The use of vaccination against S. aureus in association with antibiotic therapy did not increase the cure rate of MQ during lactation, but it was effective in reducing the SCC when compared to non-treated MQ. Although to ensure that the decrease of the SCC in ATB+VAC group was associated with the vaccination, the study should have included an additional group of only vaccinated cows, without antimicrobial therapy, with was not done in the present study, and therefore is one of the limitations of the experimental protocol used.

O objetivo deste estudo foi avaliar a eficácia da antibioticoterapia associada à vacinação sobre a taxa de cura microbiológica de mastite subclínica causada por Staphylococcus aureus em vacas leiteiras em lactação. Foram selecionados 5 rebanhos, dos quais 72 vacas (120 quartos mamários, QM) foram diagnosticadas com mastite subclínica por S. aureus e foram alocadas aleatoriamente em um de três grupos de tratamento: a) Controle (sem tratamento); b) ATB (antibioticoterapia); e c) ATB+VAC (antibioticoterapia mais vacinação contra S. aureus). O tratamento intramamário consistiu em infusão de ampicilina 75 mg + cloxacilina 200 mg duas vezes ao dia, durante 5 dias. O tratamento parenteral foi feito por injeção de uma dose única (7,5 mg/kg) de enrofloxacina, no primeiro dia do protocolo de tratamento. As vacas vacinadas receberam três doses de uma vacina comercial, 14 dias antes do tratamento (d-14), no primeiro dia do protocolo de tratamento (d1) e 14 dias após o protocolo de tratamento (d+14). A taxa de cura das vacas não tratadas foi menor (0,06) do que das vacas tratadas com ATB (0,84) e ATB+VAC (0,85). Não foi observada diferença de taxa de cura entre vacas tratadas com ATB e ATB+VAC. Por outro lado, as vacas vacinadas apresentaram menor a contagem de células somáticas (CCS) após 28 dias de tratamento (4,76 log10) do que em vacas não tratadas (5,37 log10). Em conclusão, o tratamento com ampicilina e cloxacilina intramamária, associados à enrofloxacina intramuscular, apresentou alta taxa de cura para MSC causada por S. aureus durante a lactação. A utilização da vacinação contra S. aureus associada à antibioticoterapia não aumentou a taxa de cura dos QM durante a lactação, mas foi eficaz na redução do CCS quando comparada à QM não tratados. Entretanto, para ter certeza que a diminuição da CCS no grupo ATB+VAC estivesse associada à vacinação, o estudo deveria ter incluído um grupo adicional de apenas vacas vacinadas, sem terapia antimicrobiana, o que não foi feito no presente estudo e, portanto, é um das limitações do protocolo experimental utilizado.

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.

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)

Mesenchymal Stem cells in the context of canine atopic dermatitis: A Review

ABSTRACT Canine atopic dermatitis (CAD) is a chronic inflammatory skin disease and has a high frequency among dermatological diseases. The interaction of genetic factors, skin and environmental conditions affect the expression of the disease, developing a complex pathology. Current multimodal treatment has numerous adverse effects and variations in its efficacy and safety, demonstrating the need to develop safe and effective therapeutic resources for patients with CAD. Mesenchymal stem cells (MSCs) are multipotent cells, with special characteristics, such as self-renewal, immunomodulatory properties, and de-differentiation, making them useful for several clinical problems. The discovery of the immunosuppressive effect of MSCs on T cells has opened the potential for new perspectives with its use as a therapeutic agent for immune diseases, such as CAD. The scarce number of research using the MSC as a treatment for CAD result in the lack of knowledge about the benefits and possible protocols to be followed for the use of this cell therapy. In this review, we highlighted the clinical studies and potential biological mechanisms of MSC-based cell therapy effects attenuating canine atopic dermatitis compared to conventional treatment, which might lead to a safe improvement of the animals clinical condition in a short period without causing adverse effects.

RESUMO A dermatite atópica canina (DAC) é uma doença inflamatória crônica da pele e tem alta frequência entre as doenças dermatológicas. A interação de fatores genéticos, pele e condições ambientais afetam a expressão da doença, desenvolvendo uma patologia complexa. O tratamento multimodal atual apresenta inúmeros efeitos adversos e variações em sua eficácia e segurança, demonstrando a necessidade de desenvolver recursos terapêuticos seguros e eficazes para pacientes com DAC. As células-tronco mesenquimais (CTM) são células multipotentes, com características especiais, como auto renovação, propriedades imunomoduladoras e desdiferenciação, tornando-se úteis para diversos problemas clínicos. A descoberta do efeito imunossupressor das CTMs sobre as células T abriu o potencial para novas perspectivas com sua utilização como agente terapêutico para doenças imunológicas, como a DAC. O escasso número de pesquisas utilizando o MSC como tratamento para DAC resulta no desconhecimento dos benefícios e dos possíveis protocolos a serem seguidos para a utilização dessa terapia celular. Nesta revisão, destacamos os estudos clínicos e os potenciais mecanismos biológicos dos efeitos da terapia celular baseada em MSC que atenuam a dermatite atópica canina em comparação com o tratamento convencional, podendo levar a uma melhora segura da condição clínica do animal em um curto período, sem causar efeitos adversos.

Percutaneous application of allogeneic adipose-derived mesenchymal stem cell in dogs submitted to minimally invasive plate osteosynthesis of the tibia

Purpose To evaluate clinical outcome following minimally invasive plate osteosynthesis (MIPO) associated with percutaneous transplantation of allogeneic adipose-derived mesenchymal stem cells (AD-MSC) at the tibial fracture site in dogs. Methods Thirty-six dogs presenting with nonarticular complete tibial fracture were included in this study. All fractures were treated by the same MIPO technique. The animals were divided in group 1 (n = 20) received a percutaneous application of 3 × 106 AD-MSC at the fracture site and group 2 (n = 16) did not receive any adjuvant treatment. Postoperative radiographic examinations were made at 15, 30, 60, 90 and 120 days. Results Fifty-eight percent of the patients were classified as skeletally immature. The median weight of the animals was 18.8 kg. The mean radiographic union time differed statistically between the AD-MSC group (28.5 days) and the control group (70.3 days). Sixty percent of dogs in group 1 and 56.25% of the group 2 were considered immature. Conclusions The use of allogeneic AD-MSC cell therapy and MIPO is a safe, viable and effective technique for promoting bone healing in nonarticular tibial fractures in dogs.(AU)

Mesenchymal Stem cells in the context of canine atopic dermatitis: A Review / Células-tronco mesenquimais no contexto da dermatite atópica canina: uma revisão

Canine atopic dermatitis (CAD) is a chronic inflammatory skin disease and has a high frequency among dermatological diseases. The interaction of genetic factors, skin and environmental conditions affect the expression of the disease, developing a complex pathology. Current multimodal treatment has numerous adverse effects and variations in its efficacy and safety, demonstrating the need to develop safe and effective therapeutic resources for patients with CAD. Mesenchymal stem cells (MSCs) are multipotent cells, with special characteristics, such as self-renewal, immunomodulatory properties, and de-differentiation, making them useful for several clinical problems. The discovery of the immunosuppressive effect of MSCs on T cells has opened the potential for new perspectives with its use as a therapeutic agent for immune diseases, such as CAD. The scarce number of research using the MSC as a treatment for CAD result in the lack of knowledge about the benefits and possible protocols to be followed for the use of this cell therapy. In this review, we highlighted the clinical studies and potential biological mechanisms of MSC-based cell therapy effects attenuating canine atopic dermatitis compared to conventional treatment, which might lead to a safe improvement of the animals clinical condition in a short period without causing adverse effects.(AU)

A dermatite atópica canina (DAC) é uma doença inflamatória crônica da pele e tem alta frequência entre as doenças dermatológicas. A interação de fatores genéticos, pele e condições ambientais afetam a expressão da doença, desenvolvendo uma patologia complexa. O tratamento multimodal atual apresenta inúmeros efeitos adversos e variações em sua eficácia e segurança, demonstrando a necessidade de desenvolver recursos terapêuticos seguros e eficazes para pacientes com DAC. As células-tronco mesenquimais (CTM) são células multipotentes, com características especiais, como auto renovação, propriedades imunomoduladoras e desdiferenciação, tornando-se úteis para diversos problemas clínicos. A descoberta do efeito imunossupressor das CTMs sobre as células T abriu o potencial para novas perspectivas com sua utilização como agente terapêutico para doenças imunológicas, como a DAC. O escasso número de pesquisas utilizando o MSC como tratamento para DAC resulta no desconhecimento dos benefícios e dos possíveis protocolos a serem seguidos para a utilização dessa terapia celular. Nesta revisão, destacamos os estudos clínicos e os potenciais mecanismos biológicos dos efeitos da terapia celular baseada em MSC que atenuam a dermatite atópica canina em comparação com o tratamento convencional, podendo levar a uma melhora segura da condição clínica do animal em um curto período, sem causar efeitos adversos.(AU)

Retorno de cavalos da raça Puro-Sangue Inglês às corridas após tratamento de fraturas slab do osso terceiro carpiano com PRP e células-tronco / Return of Thoroughbred racing horses to the races after treating slab fractures of the third carpal bone with PRP and stem cells

Background: Despite the fact that slab fracture of the third carpal bone is an event of great worldwide relevance in racehorses, the third carpal fracture doesn’t have data on treatments and return to racing in Brazil. The search for efficient treatments and which provide recovery providing horses return to racing is an objective of sports equine medicine. Regenerative therapies like Platelet Rich Plasma (PRP) and Stem Cells (MSc) have demonstrated a great potential in the treatment of several injuries. For the treatment of three Thoroughbred racing horses at Brazilian Jockey Club, with sagittal plane slab fractures of the third carpal bone, we used the association of PRP and Stem Cells for reduction of the rest and good return to conditions athletics of these horses. Cases: Three Thoroughbred racing horses, males, 4 and 5 years old, showed slab fracture third carpal bone in different moments of their activity at Brazilian Jockey Club. Usually, the treatment for this type of fracture is the arthroscopic repair and the conservative management. We performed after initial radiographic evaluation dorsoproximal-dorsodistal oblique (DPr-DDiO) “skyline”, intra-articular applications of PRP and MSCs. The treatments were performed according to the radiographic follow-up of the lesion evolution. No other treatment was performed concomitantly with cell therapy in the 3 treated patients. Three treatments were performed in the first patient and 2 in the second and third patients. Before each treatment for all patients, we performed radiography dorsoproximal-dorsodistal oblique (DPr-DDiO) ‘skyline’ of the carpus. After this step, the animal was sedated with 10% xylazine hydrochloride at a dose of 0.5 mg/kg intravenously. Antisepsis of the carpus was performed and applied with radiographic monitoring of the needle positioning, first the PRP in the volume of 2 mL and in sequence, 20 million cells MSCs suspended in autologous...

Reconstruction of parietal bone defects with adiposederived mesenchymal stem cells: experimental study

Purpose: This study assessed the regeneration potential of mesenchymal stem cells (MSC) from adipose tissue associated with platelet-rich plasma (PRP) in bone regeneration. Methods: Thirty Wistar rats (Rattus norvegicus albinos) were divided into five groups (according to the grafting material and time to euthanasia): (1) autograft - 14 days (control), (2) autograft - 28 days (control), (3) MSC + PRP - 14 days, (4) MSC + PRP + papaverine - 14 days and (5) MSC + PRP + papaverine - 28 days. After euthanasia, the graft was removed and histological slides were prepared. They were assessed by a blinded pathologist using a previously published histological scale as parameter. Results: There was some degree of neoformed bone trabeculae (NBT) in 93.3% of the samples, as well as osteoblastic activity (OA). The autograft groups (14 and 28 days) had higher levels in the formation of bone trabeculae. Nonparametric data were analyzed using the Wilcoxon-Mann-Whitney test and proved not to be statistically significant at p 0.05. Conclusions: Experimental parietal bone reconstruction, combining MSC, PRP and papaverine presented regeneration in all groups with no significant difference among them.(AU)

Chlorophyll content for millet leaf using hyperspectral imaging and an attention-convolutional neural network / Estimativa do teor de clorofila para folhas de milheto usando imagens hiperespectrais e uma rede neural atenção-convolucional

Chlorophyll is a major factor affecting photosynthesis; and consequently, crop growth and yield. In this study, we devised a chlorophyll-content detection model for millet leaves in different stages of growth based on hyperspectral data. The hyperspectral images of millet leaves were obtained under a wavelength range of 380-1000 nm using a hyperspectral imager. Threshold segmentation was performed with near-infrared (NIR) reflectance and normalized difference vegetation index (NDVI) to intelligently acquire the regions of interest (ROI). Furthermore, raw spectral data were preprocessed using multivariate scatter correction (MSC). A correlation coefficient-successive projections algorithm (CC-SPA) was used to extract the characteristic wavelengths, and the characteristic parameters were extracted based on the spectral and image information. A partial least squares regression (PLSR) prediction model was established based on the single characteristic parameter and multi-characteristic parameter fusion. The determination coefficient (Rv 2) and the root-mean-square error (RMSEv) of the validation set for the multi-characteristic parameter fusion model were reported to be 0.813 and 1.766, respectively, which are higher than those obtained by the single characteristic parameter model. Based on the multi-characteristic parameter fusion, an attention-convolutional neural network (attention-CNN) (Rv 2 = 0.839, RMSEv = 1.451, RPD = 2.355) was established, which is more effective than the PLSR (Rv 2 = 0.813, RMSEv = 1.766, RPD = 2.167) and least squares support vector machine (LS-SVM) models (Rv 2 = 0.806, RMSEv = 1.576, RPD = 2.061). These results indicated that the combination of hyperspectral imaging and attention-CNN is beneficial to the application of nutrient element monitoring of crops.(AU)

A clorofila é um fator importante que afeta a fotossíntese e, consequentemente, o crescimento e o rendimento das culturas. Neste estudo, um modelo de detecção de conteúdo de clorofila é construído para folhas de milheto em diferentes estágios de crescimento, com base em dados hiperespectrais. As imagens hiperespectrais dos diferentes estágios de crescimento das folhas de milheto foram obtidas para 380-1000 nm, utilizando um gerador de imagens hiperespectrais. Uma segmentação de limiar foi realizada com refletância no infravermelho próximo (NIR) e índice de vegetação com diferença normalizada (NDVI) para adquirir de forma inteligente as regiões de interesse (ROI). Além disso, os dados espectrais brutos foram pré-processados usando o método de correção de dispersão multivariada (MSC). Um algoritmo de projeção de coeficiente de correlação sucessivo (CC-SPA) foi utilizado para extrair os comprimentos de onda característicos, e os parâmetros característicos foram extraídos com base nas informações espectrais e de imagem. O modelo de previsão de regressão parcial dos mínimos quadrados (PLSR) foi estabelecido com base nos parâmetros de característica única e na fusão de parâmetros de característica múltipla. O coeficiente de determinação (Rv2) e o erro quadrático médio da raiz (RMSEv) do conjunto de validação para o modelo de fusão de parâmetros com várias características foram obtidos como 0,813 e 1,766, sendo melhores do que os do modelo de parâmetro de característica única. Com base na fusão de parâmetros com várias características, foi estabelecida uma rede neural atenção-convolucional (atenção-CNN) (Rv2 = 0,839, RMSEv = 1,451, RPD = 2,355) mais eficaz que o PLSR (Rv2 = 0,813, RMSEv = 1,766, RPD = 2,167) e mínimos quadrados que suportam modelos de máquina de vetores (LS-SVM) (Rv2 = 0,806, RMSEv = 1,576, RPD = 2,061)....(AU)

Study of comparative proteome between normal and inverted karyotypes of human mesenchymal stem cells

Multipotent mesenchymal stem cells have been expanded in vitro for cellular therapy in numerous clinical settings without standardized culture conditions or quality-control schemes. The in vitro expansion is necessary to obtain sufficient cells for clinical applications. However, the expansion may induce genetic and functional abnormalities which may affect the safety and functionality of MSC, especially the chromosomal stability. This study aimed to investigate the protein profile of umbilical cord-derived MSC with normal and inverted karyotypes after expansion in the laboratory. Mass spectrometry analysis was performed and the Bradford method, Scaffold software, String and Cytoscape databases were employed to measure and characterize the protein content of umbilical cord-derived MSC. Networks of protein interactions, hub and bottleneck proteins were identified by proteomics and systems biology approaches. We found that proteins related to cellular stress were super expressed in inverted karyotype cells. Moreover, a high expression of Serpine 1, RHOA, and CTSB was found in these cells, which are proteins related to cancer. The albumin and ubiquitin proteins have been associated with a positive prognosis in cancer and cellular stress, and were up- and down-regulated in normal karyotype cells, respectively. The results suggests that the paracentric inversion inv(3)(p25p13) induced some type of cellular stress and genetic instability in human mesenchymal stem cells. These analyses showed the importance of carrying out studies related to the genetic instability of human mesenchymal stem cells using the protein expression profile as a parameter.

Study of comparative proteome between normal and inverted karyotypes of human mesenchymal stem cells

Multipotent mesenchymal stem cells have been expanded in vitro for cellular therapy in numerous clinical settings without standardized culture conditions or quality-control schemes. The in vitro expansion is necessary to obtain sufficient cells for clinical applications. However, the expansion may induce genetic and functional abnormalities which may affect the safety and functionality of MSC, especially the chromosomal stability. This study aimed to investigate the protein profile of umbilical cord-derived MSC with normal and inverted karyotypes after expansion in the laboratory. Mass spectrometry analysis was performed and the Bradford method, Scaffold software, String and Cytoscape databases were employed to measure and characterize the protein content of umbilical cord-derived MSC. Networks of protein interactions, hub and bottleneck proteins were identified by proteomics and systems biology approaches. We found that proteins related to cellular stress were super expressed in inverted karyotype cells. Moreover, a high expression of Serpine 1, RHOA, and CTSB was found in these cells, which are proteins related to cancer. The albumin and ubiquitin proteins have been associated with a positive prognosis in cancer and cellular stress, and were up- and down-regulated in normal karyotype cells, respectively. The results suggests that the paracentric inversion inv(3)(p25p13) induced some type of cellular stress and genetic instability in human mesenchymal stem cells. These analyses showed the importance of carrying out studies related to the genetic instability of human mesenchymal stem cells using the protein expression profile as a parameter.(AU)

Characterization of mesenchymal stem cells derived from adipose tissue of a cougar (Puma concolor)

Adipose derived mesenchymal stem cells (AMSCs) have been isolated from domestic and wild cats. For wild cats, the isolation of AMSCs has been reported in the black-footed cats (Felis nigripes) and guigna (Leopardus guigna). Stromal vascular fraction (SVF) isolated from cougar adipose tissue have been used to restore elbow functionality in the cougar (Puma concolor) but multipotent characteristics of these cells have not been described. The present study describes for the first time the isolation and characterization of mesenchymal stem cells derived from adipose tissue of cougar. AMSCs and fibroblasts from six months female cougar were isolated and cultured in DMEM/F12, supplemented with FBS 10% + 1% Antibiotic/Antifungal + 2.4 mM L-Glutamine + 2.4 mM pyruvate up to passage 5. Expression of pluripotent and surface marker genes was evaluated at mRNA level. Mesodermal differentiation (adipogenic, osteogenic and chondrogenic) was described. AMSCs expressed mRNA of pluripotent genes Oct4, Nanog, Sox2 and Klf4 and surface markers Cd44, Cd90, Cd105 and MHCII. Fibroblasts showed similar mRNA expression with the exception of Sox2. AMSCs obtained from cougar exhibit multipotency features similar to domestic cats MSC, nevertheless, other analyses are required. AMSCs from cougar could be a source of interest for treatment of individuals that remain in captivity or arrive to wildlife rehabilitation centers.