Treatment of chronic immunomediated thrombocytopenia with mesenquimal stem cells in dog: case report / Tratamento de trombocitopenia imunomediada crônica com células-tronco mesenquimais em cão: relato de caso

Immunomediated thrombocytopenia is a systemic metabolic disorder in which the platelet count falls below reference values, as the patient's immune system destroys them. The main clinical signs in thrombocytopenia are petechial, hemorrhages, ecchymoses and suffusions. Hematomas can also occur in coagulation disorders. The diagnosis is based on clinical findings and hematological examinations. The treatment consists of the use of corticosteroids and immunosuppressants, delaying cell destruction, and may last for months, not always obtaining a cure for the disease. The present work reports the use of therapy with allogeneic mesenchymal stem cells, derived from the adipose tissue of dogs, for the treatment of chronic immunomediated thrombocytopenia, with an evolution of more than one year, in a Pinscher dog. The alternative treatment showed a good evolution, keeping platelets within the reference values during the treatment, giving the patient quality of life and removing the need for continuous medication for homeostasis after treatment.

A trombocitopenia imunomediada é uma desordem metabólica sistêmica, na qual a contagem plaquetária fica abaixo dos valores de referência, pois o sistema imunológico do paciente a destrói. O principal sinal clínico na trombocitopenia são hemorragias, petequiais, equimoses e sufusões. Hematomas podem ocorrer também em alterações da coagulação. O diagnóstico baseia-se nos achados clínicos e nos exames hematológicos. O tratamento consiste na utilização de corticosteroides e imunossupressores, o que retarda a destruição celular, mas pode se prolongar por meses, nem sempre obtendo cura da doença. O presente trabalho relata a utilização da terapia com células-tronco mesenquimais alogênicas, oriundas do tecido adiposo de cães, para tratamento de trombocitopenia imunomediada crônica, com evolução de mais de um ano, em um cão da raça Pinscher. O tratamento alternativo revelou boa evolução, pois manteve as plaquetas dentro dos valores de referência durante o tratamento, o que proporcionou qualidade de vida ao paciente e tornou desnecessárias medicações de uso contínuo para a homeostase após o tratamento.

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.

Chemical constituents and antifungal potential of Attalea geraensis Barb. Rodr. (Arecaceae) palm leaves, a species native to the Cerrado of Brazil / Constituintes químicos e potencial antifúngico de folhas da Attalea geraensis Barb. Rodr. (Arecaceae), uma espécie nativa do Cerrado do Brasil

Fungal diseases, especially those that affect the root systems of plants, caused by Rhizoctonia and Macrophomina are limiting factors for achieving high crop yields. Alternatives to controlling fungi with chemical products drive the search for new options for bioactive compounds from plants. Attalea geraensis, a palm tree from the Brazilian Cerrado, is rich in flavonoids with antifungal actions. The objective of this work is to identify the chemical classes present in the ethanolic extract of green leaves of A. geraensis and determine the antifungal potential of the extract against isolates of Macrophomina phaseolina (Tassi) Goid. and Rhizoctonia solani JG Kühn. Phytochemical prospection, flavonoid dereplication, and antifungal activity were carried out of the ethanolic extract of the green leaves of A. geraensis harvested in the Cerrado area of Brazil. Steroids, triterpenes, saponins, and anthraquinones are described here for the first time for the leaves of A. geraensis. The flavonoids quercetin, isorhamnetin, 3,7-dimethylquercetin, quercetin 3-galactoside, 5,7-dihydroxy-2-(4-hydroxy-3-methoxyphenyl)-3-{[3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}-4H-chromen-4-one, rhamnazin 3-galactoside, keioside, and rhamnazin 3-rutinoside were identified. Of these, only quercetin and isorhamnetin had already been identified in the leaves of A. geraensis. The results show a fungistatic potential for the species. The diversity of flavonoids present in the leaves of A. geraensis may be the result of a synergistic action between fungus and plant or there could be an antagonistic effect between flavonoids and the other identified chemical classes.

Doenças fúngicas, especialmente as que afetam os sistemas radiculares das plantas, causadas por Rhizoctonia e Macrophomina, são fatores limitantes para obtenção de grande produtividade das culturas. Alternativas ao controle dos fungos com produtos químicos impulsionam a pesquisa de novas opções de compostos bioativos oriundos de plantas. A Attalea geraensis, uma palmeira do Cerrado brasileiro, é rica em flavonoides com ações antifúngicas. O objetivo do trabalho foi identificar as classes químicas presentes no extrato etanólico das folhas verdes de A. geraensis e determinar o potencial antifúngico do extrato frente a isolados de Macrophomina phaseolina (Tassi) Goid. e Rhizoctonia solani J.G. Kühn. Realizou-se a prospecção fitoquímica, desreplicação de flavonoides e atividade antifúngica a partir do extrato etanólico das folhas verdes da A. geraensis, colhida em área de Cerrado do Brasil. Os esteroides, triterpenos, saponinas e antraquinonas estão sendo descritos pela primeira vez para as folhas de A. geraensis. Foram identificados os flavonoides quercetina, isoramnetina, 3,7-dimetilquercetina, quercetina 3-galactosídeo, 5,7-dihidroxi-2-(4-hidroxi-3-metoxifenil)-3-{[3,4,5-trihidroxi-6-(hidroximetil)oxan-2-il]oxi}-4H-cromen-4-ona, ramnazina 3-galactosídeo, keiosídeo e ramnazina 3-rutinosídeo. Destes, somente a quercetina e isorhamnetin já haviam sido identificadas nas folhas da A. geraensis. Os resultados indicam potencial fungistático para a espécie. Infere-se que a diversidade de flavonoides presentes nas folhas de A. geraensis pode ser resultado da ação sinérgica entre fungo e planta ou que haja um efeito antagonista entre os flavonoides e as demais classes químicas identificadas.

Cell therapy with adipose tissue-derived human stem cells in the urinary bladder improves detrusor contractility and reduces voiding residue / A terapia celular com células-tronco humanas derivadas do tecido adiposo na bexiga urinária melhora a contratilidade do detrusor e reduz o resíduo miccional

Detrusor hypocontractility (DH) is a disease without a gold standard treatment in traditional medicine. Therefore, there is a need to develop innovative therapies. The present report presents the case of a patient with DH who was transplanted with 2 x 106 adipose tissue-derived mesenchymal stem cells twice and achieved significant improvements in their quality of life. The results showed that cell therapy reduced the voiding residue from 1,800 mL to 800 mL, the maximum cystometric capacity from 800 to 550 mL, and bladder compliance from 77 to 36.6 mL/cmH2O. Cell therapy also increased the maximum flow from 3 to 11 mL/s, the detrusor pressure from 08 to 35 cmH2O, the urine volume from 267 to 524 mL and the bladder contractility index (BCI) value from 23 to 90. The International Continence on Incontinence Questionnaire - Short Form score decreased from 17 to 8. Given the above, it is inferred that the transplantation of adipose tissue-derived mesenchymal stem cells is an innovative and efficient therapeutic strategy for DH treatment and improves the quality of life of patients affected by this disease.

A Hipocontratilidade Detrusora (HD) é uma doença sem um tratamento padrão-ouro na medicina tradicional. Logo, há a necessidade de desenvolvimento de terapias inovadoras. O presente relato apresenta um caso de paciente com HD transplantado duas vezes com 2 x 106 células-tronco mesenquimais derivadas do tecido adiposo que obteve melhoras significativas em sua qualidade de vida. Os resultados demonstraram que a terapia celular reduziu o resíduo miccional de 1.800mL para 800mL; a Capacidade Cistométrica Máxima de 800 para 550mL; a complacência de 77 para 36,6mL/cmH2O. A terapia celular também aumentou o fluxo máximo de 3 para 11mL/s; a pressão detrusora de 08 para 35cmH2O; o volume urinado de 267 para 524mL e o índice de contratilidade vesical (BCI) de 23 para 90. O score do International Continence on Incontinence Questionare - Short Form passou de 17 para 8. Diante do exposto, infere-se que o transplante de células-tronco mesenquimais derivadas do tecido adiposo é uma estratégia terapêutica inovadora e eficiente para o tratamento da HD e para melhoria da qualidade de vida de pacientes acometidos por essa doença.

Efficacy of Ficus carica leaf extract on morphological and molecular behavior of mice germ stem cells

Infertility is one of the most prevalent health disorders in reproductive-age males and females. Ficus carica (Fc), an herbal plant, has been used traditionally for the treatment of different diseases such as infertility especially in Iranian folk medicine. This study examined the effects of Fc leaf extract on the proliferation of mice spermatogonial stem cells (SSCs). Phenolic, flavonoid content, major polyphenolic compounds and antioxidant activity of the extract was evaluated respectively by Folin-Ciocateu, aluminum chloride, HPLC and the FRAP and DPPH methods. Testicular cells of neonate mice were extracted and their identity was confirmed using cytokeratin for Sertoli and Oct-4, CDHI and PLZF for SSCs. Effects of Fc (0.0875, 0.175, 0.35, 0.71 and 1.42 mg/ml) was evaluated at third, 7th, 9th and 14th days of culture by colony assay. The expression of the Mvh, GFRα1 and Oct-4 genes and the viability and proliferation of cultured cells was assessed at the end of the culture period. The extract has a rich phenolic and flavonoid content such as Rutin, Psoralen, Bergapten and Caffeoylmalic acid using HPLC analysis. It also had a potent reducing and radical scavenging activity. Morphology of colonies was similar in all groups. Higher viability, proliferation, colony number and diameter of SSCs was seen in the presence of Fc leaf extract in a dosedependent manner so that higher number and diameter of colonies were observed in two higher doses of 0.71 and 1.42 mg/ml, separately for each time point relative to other groups. The Mvh, Oct-4 and GFRα1 genes expression had no significant differences between groups. It seems that Fc leaf extract not only had no any cytotoxic effects on the viability and proliferation of SSCs but also support their stemness state. So, this culture system can be employed for enrichment of germ stem cells for use in clinical applications.(AU)

Comparison of three different strategies to treat sciatic nerve regeneration: an experimental study

Purpose: To compare the effect of vein conduit filled with adipose tissue stem cells (ASC) on peripheral nerve injury regeneration. Methods: We analyzed 30 male Wistar rats surgically submitted to a 5-mm gap on the sciatic nerve. Then, the animals were divided into three groups: nerve autografting (AG, n=10), autogenous inverted glycerol-conserved vein (VG, n=10), and autogenous inverted glycerol-conserved vein + ASC (VASCG, n=10). The study endpoints were neuromotor functional analysis, gastrocnemius muscle weight, and sciatic nerve graft histomorphometry analysis. In the histologic analysis, we added a control group (naïve nerve). Results: Regarding functional analysis (Walking tract- score), the findings at week 3 showed a difference between the AG and the VG (-96.6 vs. -59.6, p=0.01, respectively) and between the VG and the inverted vein + VASCG (-59.9 vs. -88.92, p=0.02). At week 12, this study showed a difference between the AG and the VG (-64.8 vs. -47.3, p=0.004, respectively), and also a difference between the VG and the VASCG (-47.3 vs. -57.4, p=0.02, respectively). There was no difference in the histomorphometry analysis (nerve diameter, Schwann cells counting). The gastrocnemius muscles on the intervention side were more atrophic when compared to the gastrocnemius muscles on the control side. Conclusions: Our results suggested better functional recovery in the inverted vein group when compared to control group, and inverted vein + ASC group.

Interference of weeds in ruziziensis grass pastures

The aim of this study was to assess the effect of increasing periods of coexistence of weed plants with Urochloa ruziziensis on the canopy structure and productivity of a pasture already established with this forage species. The experiment was a randomized blocks design with four replications, and treatments consisted of seven increasing periods of coexistence of forage grass with weed plants: 0 (control), 15, 30, 45, 60, 75 and 90 days after regrowth (DAR). The main morpho-structural and productive characteristics of the forage plants were determined at the end of the experimental period (90 DAR). The ratio of the first green leaf height to the tiller height increases, while the leaf to stem ratio diminishes as the period of interaction with the infesting community increases. The number of green leaves per tiller and the tiller height diminishes as the period of coexistence with weed plants increases. The presence of weed plants interferes negatively with all parameters of the grass canopy structure and productivity of a grazing land already established with Urochloa ruziziensis, suggesting that measures of control of the infesting community should be adopted up to 17 days of regrowth of the forage plant.(AU)

Uso de células-tronco no tratamento de sequelas neurológicas causadas pela cinomose / Use of stem cells in the treatment of sequelates neurological caused by cinomosis

A cinomose canina é uma doença infectocontagiosa causada pelo vírus Paramyxoviridae, que acomete cães de todas as idades, entretanto principalmente cães filhotes não vacinados. Uma das sequelas da doença é neurológica que, por muitas vezes, é de difícil tratamento. A terapia com células-tronco é uma alternativa para atenuar os sinais e sequelas neurológicas, propiciando melhor qualidade de vida para o animal. As células-tronco são células com alta capacidade de auto renovação e diferenciação, e, portanto, uma alternativa de tratamento para as sequelas neurológicas derivada pela cinomose.

Canine distemper is an infectocontagious disease caused by Paramyxoviridae virus, which affects dogs of all ages, however mainly unvaccinated puppies. Neurological manifestation are common in canine distemper which is often difficult to treat. Stem cell therapy is an alternative to attenuate neurological signs and sequel, providing better quality of life for the animal. Stem cells have high capacity for self-renewal and differentiation, and an alternative treatment for neurological sequel derived by distemper.

Air layering in Caryocar brasiliense - effect of stem diameter / Alporquia em pequizeiro - efeito do diâmetro do caule

Seed propagation of pequizeiro (Caryocar brasiliense) has many disadvantages, including low germination rates and high genetic variability of the resulting plants. Therefore, vegetative propagation techniques, such as the air layering process, are potentially important for this species. This study evaluated the effects of stem diameter and parental plant on the air layering efficacy of pequizeiro. The air layering process was carried out in mid-December 2019, in the following stem diameters: less than 15 mm; 15-19 mm; 20-24 mm; 25-29 mm and above 29 mm. The treatments comprised four replicates of 10 air layers. After 3 months, the percentages of survival, callus formation and rooting were evaluated, in addition to the rooting vigor. The percentages of survival and callus formation observed in the air layering, regardless the parental plant, were higher than 85%, showing the high potential of this technique in the propagation of pequizeiro. The callus differentiation rate in roots varied between the parental plants. Considering the evaluated parameters, stem diameters ranging from 20 to 24 mm are the most suitable ones for the air layering process in pequizeiro. These results indicated the high potential of the air layering technique for the commercial propagation of pequizeiro.

A propagação seminal de pequizeiro acarreta grande variabilidade na taxa de germinação e no genótipo das mudas produzidas, mostrando a necessidade do desenvolvimento de técnicas para a propagação vegetativa dessa espécie, como a alporquia. Os objetivos deste estudo foram avaliar a eficácia da alporquia, bem como o efeito do diâmetro do caule e da planta-mãe na sobrevivência, no calejamento e no enraizamento de alporques de pequizeiro. O processo de alporquia foi realizado em meados de dezembro de 2019, nos seguintes diâmetros de caule: inferior a 15 mm; 15-19 mm; 20-24 mm; 25-29 mm e superior a 29 mm. Os tratamentos foram testados em quatro repetições de 10 alporques. Após três meses, avaliaram-se as porcentagens de sobrevivência, de calejamento e de enraizamento, além do vigor de enraizamento. As porcentagens de sobrevivência e de calejamento nos alporques, independentemente da planta matriz, foram superiores a 85%, mostrando o grande potencial dessa técnica na reprodução de pequizeiro. A taxa de diferenciação de calos em raízes variou entre as matrizes. Considerando os parâmetros avaliados, diâmetros de caule de 20 a 24 mm são os mais adequados para a alporquia em pequizeiro. Esses resultados indicam o potencial da técnica de alporquia para a produção comercial de mudas de pequizeiro.

Triiodothyronine does not influence in vitro chondrogenic differentiation of adipose tissue-derived stem cells from young female rat / Triiodotironina não influencia na diferenciação condrogênica de células tronco do tecido adipose de ratas jovens

The objective of this study was to investigate the in vitro action of triiodothyronine (T3) on the chondrogenic differentiation of adipose tissue-derived stem cells (ASCs) of female rats, with different time periods and doses. ASCs were extracted from female Wistar rats and were cultured in chondrogenic medium with and without the presence of T3. Five groups were established: 1) ASCs without T3; and 2,3,4,5) ASCs with 0.01, 1, 100 and 1,000 nM T3, respectively). After 7, 14 and 21 days, cell morphology, chondrogenic matrix formation, and expression of Sox9, aggrecan, collagen II, and collagen X were evaluated. The Student-Newman-Keuls test was used. ASCs showed CD54, CD73, and CD90 before chondrogenic differentiation. The hormone treatment did not alter chondrogenic matrix formation, Sox9 expression at 14 or 21 days, or expression of collagen II or collagen X at any time. However, the 0.01, 1, and 1000 nM T3 doses decreased Sox9 expression at 7 days. In conclusion, chondrogenic differentiation of ASCs of female rats is not influenced by T3.

O objetivo do presente trabalho foi verificar o efeito in vitro da triiodotironina (T3) na diferenciação condrogênica de células tronco mesenquimais do tecido adiposo (CTM-TA) de ratas, durante vários períodos e em várias doses. CTM-TA foram coletadas de ratas Wistar e cultivadas em meio condrogênico com ou sem a presença de T3. Constitui-se cinco grupos: 1) CTM-TA sem T3; e 2,3,4,5) CTM-TA com T3 (0,01; 1; 100 e 1000 nM, respectivamente). Após sete, 14 e 21 dias, foram avaliados morfologia celular, formação de matriz condrogênica e expressão de Sox9, agrecano, colágeno II e colágeno X. Para as análises foi utilizado o teste de Student Newman Keuls. CTM-TA expressaram CD54, CD73 e CD90 antes da diferenciação condrogênica. O tratamento hormonal não alterou a formação de matriz condrogênica e a expressão de Sox9 aos 14 e 21 dias e expressão dos colágenos II e X em nenhum dos períodos avaliados. No entanto, as doses de 0,01; 1 e 1000 nM T3 diminuíram a expressão de Sox9 aos 7 dias. Conclui-se que a diferenciação condrogênica de CTM-TA de ratas não é influenciada pela T3.

O extrato aquoso da casca de pequi Caryocar brasiliense melhora a retração de feridas / Aqueous extract of stem bark from Caryocar Brasiliense improves the wound retraction / El extracto acuoso de la cáscara de Caryocar brasiliense mejora la retracción de heridas

Este estudo foi conduzido para verificar o efeito do extrato aquoso e da pomada à base de casca de Caryocar brasiliense sobre a retração de feridas em coelhos. Foram utilizados 12 coelhos Nova Zelândia Branco divididos em dois grupos e quatro lesões foram produzidas na região dorsal de cada animal. As lesões à direita foram tratadas com extrato aquoso (grupo 1) ou com a pomada (grupo 2) e as lesões à esquerda foram tratadas com solução salina 0,9% (grupo controle). Os níveis de colágeno e fibroblastos foram menores (P<0,05) em lesões tratadas com extrato aquoso, comparado com o grupo controle. Aos 7 e 14 dias após o procedimento, a retração das lesões era maior (P<0,05) quando tratadas com extrato aquoso e nas lesões tratadas com a pomada, a melhora ocorreu apenas no 7º dia, comparado com o tratamento controle. Concluiu-se que o extrato aquoso de casca de Caryocar brasiliense melhorou a retração de feridas por um período de tempo maior do que a pomada.

This study was carried out to verify the effect of the aqueous extract and the ointment of the stem bark from Caryocar brasiliense on the wound healing in rabbits. Twelve New Zealand rabbits were divided in two groups and four lesions were produced on the back of each animal. The lesions on the right side were treated with aqueous extract (group 1) or with ointment (group 2), while the lesions on the left side were the control group (treated with 0.9% saline solution). Collagen and fibroblast levels were lower (P<0.05) in wounds treated with aqueous extract, compared to the control group. On days 7 and 14 after the procedures, the wound retraction was better (P<0.05) when treated with aqueous extract, while in wounds treated with the ointment, the improvement occurred only on the 7th day. The aqueous extract of stem bark from Caryocar brasiliense improved the wound retraction longer than the ointment.

Este estudio fue conducido para verificar el efecto del extracto acuoso y de la pomada basada en cáscara de Caryocar brasiliense sobre la retracción de heridas en conejos. Fueron utilizados 12 conejos Nueva Zelanda Blanco divididos en dos grupos y cuatro lesiones fueron producidas en la región dorsal de cada animal. Las lesiones a la derecha fueron tratadas com extracto acuoso (grupo 1) o con la pomada (grupo 2) y las lesiones a la izquierda fueron tratadas con solución salina 0,9% (grupo control). Los niveles de colágeno y fibroblastos fueron menores (P<0,05) en lesiones tratadas con extracto acuoso, comparado con el grupo control. A los 7 y 14 días después del procedimiento, la retracción de las lesiones era mayor (P<0,05) cuando tratadas con extracto acuoso y en las lesiones tratadas con la pomada, la mejora ocurrió solamente el 7° día, comparado con el grupo control. Se concluyó que el extracto acuoso de la cáscara de Caryocar brasiliense mejoro la retracción de heridas por un período de tiempo mayor do que la pomada.

Mesenchymal stem cells have ameliorative effect on the colitis model via Nrf2/HO-1 pathway

Purpose: To evaluate the ameliorative effect of mesenchymal stem cells (MSCs) on acetic acid colitis model via Nrf2/HO-1 pathway in rats. Methods: In this study, 30 rats were divided into three groups. Acute colitis was induced by rectal administration of 4% solution of acetic acid. MSCs were injected intraperitoneally in the treatment group. Results: Increased levels of tumor necrosis factor-α (TNF-α), pentraxin-3, and malondialdehyde (MDA) in colitis group were revealed biochemically. Increased level of TNF-α and decreased levels of Nrf2 and interleukin-10 (IL-10) were observed in rectum tissues. Increased fibrous tissue proliferation, vascularization and inflammatory cell infiltration were described in the colitis group. Significant improvement was observed in MSCs treated group histopathologically. Increased immunopositivity of TNF-α, vascular endothelial growth factor (VEGF) and CD68 markers was observed in the colitis group cells, and decreased level of this positivity was observed in MSCs treated group. Conclusions: Biochemical, histopathological and immunohistochemical results strongly support the ameliorative effect of MSCs against acetic induced colitis model via Nrf2/HO-1 pathway in rats.

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.

Soybean development under different Aphelenchoides besseyi and Meloidogyne incognita populations / Desenvolvimento da soja sob diferentes populações de Aphelenchoides besseyi e Meloidogyne incognita

The green stem Aphelenchoides besseyi and gall Meloidogyne incognita phytonematodes cause soybean yield losses. Thus, the objective of this study was to evaluate the effects of different populations of A. besseyi and M. incognita on soybeans. The experiment had 10 treatment groups [T1: control, T2: A. besseyi (500), T3: M. incognita (2,000), T4: A. besseyi (250) + M. incognita (2,000), T5: A. besseyi (500) + M. incognita (2,000), T6: A. besseyi (250) + M. incognita (4000), T7: A. besseyi (500) + M. incognita (4,000) individuals, T8: A. besseyi (250) + M. incognita (8,000), T9: A. besseyi (500) + M. incognita (5,000), and T10: A. besseyi (1,000) + M. incognita (8,000) individuals] with six replicates. Inoculations were done on day 10 after soybean germination, and evaluations were conducted after 60 d. Plant height, shoot fresh weight, root fresh weight, number of pods (NP), pod fresh weight, number of grains (NG), grain yield (GY), amount of A. besseyi in the shoot, and M. incognita in the roots were analyzed. The 500 A. besseyi + 8,000 M. incognita treatment reduced all variables and led to considerable damage to soybean development. In addition, GY, NG, and NP were reduced in the different nematode population density groups, influencing soybean agronomic characteristics.(AU)

Entre as espécies de fitonematoides que provocam perdas de produção à cultura da soja estão os nematoides da haste verde (Aphelenchoides besseyi) e o das galhas (Meloidogyne incognita). Diante disso, o objetivo deste trabalho foi estudar os efeitos de diferentes níveis populacionais de A. besseyi e M. incognita na soja. O experimento foi realizado com nove combinações de diferentes níveis populacionais dos nematoides (T1 ­ Controle não inoculado, T2 - A. besseyi 500 indivíduos, T3 - M. incognita 2000 indivíduos, T4 - A. besseyi + M. incognita 250 + 2000 indivíduos, T5 - A. besseyi + M. incognita 500 + 2000 indivíduos, T6 - A. besseyi + M. incognita 250 + 4000 indivíduos, T7 - A. besseyi + M. incognita 500 + 4000 indivíduos, T8 - A. besseyi + M. incognita 250 + 8000 indivíduos, T9 - A. besseyi + M. incognita 500 + 8000 indivíduos e T10 - A. besseyi + M. incognita 1000 + 8000 indivíduos) e seis repetições. As variáveis analisadas foram: altura de planta, massa fresca da parte aérea (MPA), massa fresca de raízes (MR), número de vagens (V), massa fresca de vagens (MV), número de grãos (G), massa fresca de grãos (MG), quantidade de A. besseyi na parte aérea e de M. incognita nas raízes. O mapa de calor indicou que populações iniciais inoculadas de 500 A. besseyi + 8000 M. incognita apresentou maiores danos ao desenvolvimento da soja, reduzindo todas as variáveis analisadas. As variáveis MG, G e V reduziram na presença de diferentes densidades populacionais das espécies de fitonematoides nematoides, influenciando negativamente o desenvolvimento da soja.(AU)

Height management strategies of sudangrass 'BRS Estribo' / Estratégias de manejo de altura para capim-sudão cultivar BRS

The productive, morphological and chemical composition of Sudangrass 'BRS Estribo' submitted to different heights of management was evaluated. The design was in randomized blocks with four replicates in a 3x2 factorial scheme, with three pre-defoliation heights (40, 55 and 70 cm) and two post-defoliation heights (20 and 10 cm). There was no significance between post-defoliation heights for accumulated forage production (AFP), percentages of leaf blades (%F) and of stem and sheath (%S). The AFP at 70 cm pre-defoliation height (10,071.1 kg ha-1) was higher than at 40 cm height (7,471.3 kg ha-1) and did not differ from defoliation at 55 cm (8,562.3 kg ha-1). Pre-defoliation at 40 cm showed a higher %F (67.67%) than at the heights of 55 cm (61.74%) and 70 cm (55.15%), which differed from each other. The %S was higher for pre-defoliation at 70 cm (35.57%) than at the heights of 55 (32.08%) and 40 cm (27.75%), which also differed from each other. The different handling heights did not change the contents of dry matter, insoluble fiber in neutral detergent, insoluble fiber in acid detergent and ash, which presented averages of 15.87; 60.34; 28.53; and 10.48%, respectively. However, there was a significant interaction between pre-and post-defoliation heights for crude protein (CP) contents, which the management heights at 40 x 20 cm and 70 x 10 cm presented, respectively, the highest (18.78%) and lowest (12.22%) CP contents of the study. Pre-defoliation heights of 55 cm with 10 or 20 cm of residual height should be used to improve the botanical composition and crude protein content of Sudangrass BRS Estribo forage.

Avaliaram-se as características produtivas, morfológicas e a composição bromatológica do capim-sudão BRS Estribo submetido a diferentes alturas de manejo. O delineamento em blocos casualizados com quatro repetições em esquema fatorial 3x2, sendo três alturas de pré-desfolhação (40, 55 e 70 cm) e duas alturas de pós-desfolhação (20 e 10 cm). Não houve efeito de alturas de resíduo pós-desfolhação para a produção total de forragem acumulada (PTA), porcentagens de lâminas foliares (%F) e de colmo e bainha (%C). A PTA das plantas submetidas a pré-desfolhação de 70 cm (10.071,1 kg ha-1) foi maior que aquelas desfolhadas com 40 cm (7.471,3 kg ha-1) e não diferiu estatisticamente das desfolhadas com 55 cm (8.562,3 kg ha-1). A pré-desfolhação à 40 cm apresentou a maior %F (67,67%) que as alturas de 55 (61,74%) e 70cm (55,15%). Já a %C foi maior para altura de pré-desfolhação à 70 cm (35,57%) em relação às alturas de 55 (32,08%) e 40cm (27,75%). As diferentes alturas de manejo não alteraram os teores de matéria seca, fibra insolúvel em detergente neutro, fibra insolúvel em detergente ácido e cinzas da forragem, que apresentou médias de 15,87; 60,34;28,53; e 10,48%, respectivamente. Houve interação significativa entre as alturas de pré e pós-desfolhação para os teores de proteína bruta (PB), onde o manejo à 40 x 20 cm e 70 x 10 cm, apresentaram, respectivamente, o maior (18.78%) e menor (12,22%) teor de PB do estudo. Alturas de pré-desfolhação de 55 cm de altura deixando-se 10 ou 20 cm de resíduo devem ser preconizadas como forma de melhorar a composição morfológica e o teor de proteína da forragem.

Characterization of hematopoietic stem cells from the canine yolk sac

Abstract The characterization of hematopoietic stem cells (HSC) from the canine yolk sac (cYS) can contribute to future gene therapies because it is possible to obtain information about the beginning of the development of the circulatory system through the characterization. The cYS is a likely source of HSC, which is a source of blood cell development in mammals. Studies in this field have been conducted for decades; however, interest in cellular therapy is currently at its peak with greater visibility, and these cells are a promising therapeutic tool for the treatment of diseases related to animals and humans. The aim of this study was to isolate and characterize HSC from the cYS embryos at 30 to 45 days of gestational age. Our results showed that the cYS was macroscopically located in the ventral region with a central portion and extremities. The cells in culture presented a circular morphology and cell clusters. The average cell viability was 22.55% dead cells out of 6.5 × 104 total cells. The cells were also able to form colonies on methylcellulose. Flow cytometry analysis revealed the expression of CD34, CD117, and CD45. Our results suggest that the cYS can be used as a source of hematopoietic cells, and this study is very important to understand the mechanism and development of the hematopoietic system in dogs.

Quantification of adhesion of mesenchymal stem cells spread on decellularized vein scaffold

ABSTRACT Purpose: To evaluate methods that improve adipose-derived stem cells (ASCs) population in decellularized biological venous scaffold for tissue engineering in blood vessels, a model in rabbits. Methods: The ASC was expanded until the third passage. Inferior vena cava (IVC) was submitted to the decellularization process using 1% sodium dodecyl sulfate (SDS) or 2% sodium deoxycholate (SD) to compose 12 study groups (G): pure SD or SDS, exposed or not to 1% TritonX-100 (TX-100) and exposed or not to poly-l'lysine and laminin (PL). Scaffolds were covered with 1 × 105 or 1 × 106 ASCs diluted in 10 μL Puramatrix™. The histological analysis was done by cell counting in hematoxylin and eosin (HE) and nuclei count in immunofluorescence (IF) with 4',6-Diamidine-2'-phenylindole dihydrochloride (DAPI). Results: The study of groups in HE and IF showed similar results. For both analyses,IVC-SD-1 × 106 ASC and IVC-SD-PL-1 × 106 ASC provided the best results. The IF technique showed better sensitivity than HE, with a weak agreement between them. Conclusions: Decellularizing agent and the number of ASC influence scaffolds cellularization response and the best protocols as those ones using SD with or without the addition of PL.

The influence of adipocyte-derived stem cells (ASCs) on the ischemic epigastric flap survival in diabetic rats

ABSTRACT Purpose: To assess the effects of adipocyte-derived stem cell (ASC)-injection on the survival of surgical flaps under ischemia in diabetic rats. Methods: Diabetes was induced in 30 male Wistar rats using streptozotocin (55 mg/kg). After eight weeks, epigastric flap (EF) surgery was performed. The animals were divided into control (CG), medium-solution (MG), and ASC groups. The outcomes were: the survival area (SA), the survival/total area rate (S/TR), and expression levels (EL) of genes: C5ar1, Icam1, Nos2, Vegf-a. Results: In the ASC group, compared to CG, we observed improved flap SA (CG-420 mm2 vs. ASC-720 mm2; p=0.003) was observed. The S/TR analysis was larger in the ASC group (78%) than the CG (45%). This study showed an increase in the Vegf-a EL in the ASC group (2.3) vs. CG (0.93, p=0.0008). The Nos2 EL increased four-fold in the ASC group compared to CG, and C5ar1 EL decreased almost two-fold in the ASC group vs. the CG (p=0.02). There was no difference among the groups regarding Icam1 EL. Compared to the MG, the ASC group had a bigger flap SA (720 mm2 vs. 301 mm2, respectively), a bigger S/TR (78% vs. 32%, p=0.06, respectively) and increased EL of Vegf-a (2.3 vs. 1.3, respectively). No difference between ASC-group and MG was seen regarding Nos2 (p=0.08) and C5ar1 (p=0.05). Conclusions: This study suggests that ASCs increase the survival of EF under IR in diabetic rats.

Pigeon as a model to study peripheral projections from the horizontal semicircular canal vestibular apparatus to a brainstem target immunoreactive for AMPA

ABSTRACT Purpose: To evaluate whether the pigeon (Columba livia) is a good model for evaluating the vestibular system involved with postural maintenance during movement. Methods: This study maps the brainstem targets of the horizontal ampullary inputs from the vestibular periphery of the pigeon. We used biotin dextran amine (BDA) injection in horizontal semicircular canal (HSCC), immunohistochemistry for GluR2/3 and GluR4 AMPA and computerized histomorphology reconstruction. Results: Our results show the same distribution pattern with ipsilateral projections to vestibular nuclear complex (VNC) from the HSCC, with the majority of labeled fibers being, long, thin, with few varicosities and many ramifications. Horizontal semicircular canal projections achieve neurons belonging to all nuclei of the VNC with exception of dorsal portion of lateral vestibular nucleus and this area express GluR2/3 and GluR4 AMPA receptors reinforcing the idea of glutamate participation in these connections. Conclusions: Pigeon is an appropriated experimental model to study of projections of HSCC and reinforcing the information that the vestibular system has strong relation with the fast responses necessary for postural control. Moreover, its phylogenetic organization apparently conservation, also seems to be a fundamental characteristic for vertebrates.

The role of Rho-associated kinase inhibitor, Y-27632 on primary culture of ovine spermatogonial stem cells

Abstract The access to sufficient numbers of spermatogonial stem cells (SSCs) is a prerequisite for the study of their regulation and further biomanipulation. Rho kinase (ROCK) belongs to a family of serine/threonine kinases and involves in a wide range of fundamental cellular functions. The aim of the present study was to study the effect of ROCK inhibitor, Y-27632 (0.1-40 µM), during the primary culture of ovine SSCs. SSCs were collected from 3-5-month-old's lamb testes. The viability of SSCs, the apoptosis assay of SSCs, the intracellular reactive oxygen species (ROS) analysis, and the SSCs markers and apoptosis-related gene expressions were detected by MTT reduction assay, Annexin V-FITC/ Propidium Iodide (PI) dual staining, flow cytometry and real-time-PCR studies, respectively. Morphological analyses indicated that the 5-10 µM Y-27632 had an optimal effect on the number of presumptive SSCs colonies and the area covered by them after a 10 days culture. The cell viability, apoptosis and necrosis of SSCs after 10 days' culture were not affected in comparison with the control group, and the 20 µM of Y-27632 resulted in significantly decreased cell viability (P<0.05) and an increased necrosis of cells. On day 10 after culture, the expression of P53 was decreased with an increase from 0 to 10 µM in the Y-27632 dose. In the 20 µM Y-27632 group, the expressions of P53 and Bax were higher and the Bcl-2 was lower than other groups and these values were significantly different from 5 and 10 µM Y-27632 groups (P<0.05). The level of intracellular ROS was decreased with an increase in the Y-27632 dose from 5 to 20 µM in comparison with the control group. In conclusion, the present study demonstrated that Y-27632 at a concentration of 5-10 µM provided optimal culture conditions for the primary culture of ovine SSCs.