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1.
Biol. Models Res. Technol ; 2(1): e00022022, 2022.
Artigo em Inglês | VETINDEX | ID: biblio-1402379

Resumo

Arboviruses are worldwide distributed arthropod-borne viruses representing a constant threat to public health. Among these arboviruses, the Chikungunya (CHIKV) and Zika (ZIKV) viruses have a high prevalence in Brazil being responsible for recent outbreaks resulting mainly in irreparable socioeconomic damages such as the high rate of cases of comorbidities and microcephaly in newborns, respectively. Therefore, it is necessary to understand the biology of these arboviruses and develop effective treatments against them; moreover, appropriate mice animal models are strongly encouraged. Here we reviewed the scientific literature aiming to improve the search for the best murine animal model, specific for the arboviruses, specifically, CHIKV and ZIKV. In this way, we performed a comparison between the various mice models currently available, among them: genetically modified immunosuppressed animals, as the A129 and AG129 which are knockout animals for the α/ß and α/ß/γ receptors, respectively, neonatal immunocompetent models C57BL/6 strains used between 6-8 days old for neuropathogenesis studies or 1 day old for vaccine safety studies and finally immunosuppressed induced by dexamethasone or interferon 1 blocker for pathogenesis studies. Mice models are the first option after in vitro analysis, as they are small animals, which facilitates handling and maintenance, in addition to being more inexpensive and abundantly available in different genetic strains, both wild and modified. If the results of this stage are promising, the studies move forward to the use of models with animals of greater complexity, such as rats, non-human primates and finally humans. For this review, we searched through articles in PubMed, Scopus and ScienceDirect databases using the criteria of date publications, titles, abstracts and complete manuscripts. The correct choice of these models during experimental planning is essential, since increases the confidence and the rational use of animals in experimentation in accordance to current bioethics guidelines.


Assuntos
Animais , Camundongos , Vírus Chikungunya , Modelos Animais , Zika virus , Animais de Laboratório , Camundongos
2.
Anim. Reprod. (Online) ; 17(3): [e20200069], 2020. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461517

Resumo

Reproductive efficiency is critically dependent on embryo survival, establishment of a successful pregnancy and placental development. Recent advances in gene editing technology have enabled investigators to use gene knockdown and knockout approaches to better understand the role of hormone signaling in placental function and fetal growth and development. In this review, an overview of ruminant placentation will be provided, including recent data highlighting the role of histone lysine demethylase 1A and androgen signaling in ruminant placenta and pregnancy. Studies in ruminant placenta establish a role for histone lysine demethylase 1A in controlling genetic networks necessary for important cellular events such as cell proliferation and angiogenesis, as well as androgen receptor signaling during early placentation.


Assuntos
Feminino , Animais , Histona Desmetilases , Placenta , Ruminantes/anatomia & histologia , Ruminantes/fisiologia , Ruminantes/genética , Androgênios
3.
Anim. Reprod. ; 17(3): [e20200069], 2020. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: vti-27859

Resumo

Reproductive efficiency is critically dependent on embryo survival, establishment of a successful pregnancy and placental development. Recent advances in gene editing technology have enabled investigators to use gene knockdown and knockout approaches to better understand the role of hormone signaling in placental function and fetal growth and development. In this review, an overview of ruminant placentation will be provided, including recent data highlighting the role of histone lysine demethylase 1A and androgen signaling in ruminant placenta and pregnancy. Studies in ruminant placenta establish a role for histone lysine demethylase 1A in controlling genetic networks necessary for important cellular events such as cell proliferation and angiogenesis, as well as androgen receptor signaling during early placentation.(AU)


Assuntos
Animais , Feminino , Ruminantes/anatomia & histologia , Ruminantes/genética , Ruminantes/fisiologia , Placenta , Histona Desmetilases , Androgênios
4.
Acta cir. bras. ; 34(10): e201901003, 2019. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: vti-24043

Resumo

Purpose:To evaluate that Connexin (Cx43) plays a role in lesions after hepatic ischemia/reperfusion (IR) injury.Methods:We use Cx43 deficient model (heterozygotes mice) and compared to a wild group. The groups underwent 1 hour ischemia and 24 hours reperfusion. The heterozygote genotype was confirmed by PCR. We analyzed the hepatic enzymes (AST, ALT, GGT) and histology.Results:The mice with Cx43 deficiency showed an ALT mean value of 4166 vs. 307 in the control group (p<0.001); AST mean value of 7231 vs. 471 in the control group (p<0.001); GGT mean value of 9.4 vs. 1.7 in the control group (p=0.001); histology showed necrosis and inflammation in the knockout group.Conclusions:This research demonstrated that the deficiency of Cx43 worses the prognosis for liver injury. The topic is a promising target for therapeutics advancements in liver diseases and procedures.(AU)


Assuntos
Animais , Camundongos , Conexina 43/análise , Junções Comunicantes , Isquemia , Reperfusão , Fígado/lesões , Camundongos Knockout , Comunicação Celular
5.
Acta cir. bras. ; 33(11): 991-999, Nov. 2018. ilus, graf
Artigo em Inglês | VETINDEX | ID: vti-18420

Resumo

Purpose:To determine whether the absence of transglutaminase 2 enzyme (TG2) in TG2 knockout mice (TG2-/-) protect them against early age-related functional and histological arterial changes.Methods:Pulse wave velocity (PWV) was measured using non-invasive Doppler and mean arterial pressure (MAP) was measured in awake mice using tail-cuff system. Thoracic aortas were excised for evaluation of endothelial dependent vasodilation (EDV) by wire myography, as well as histological analyses.Results:PWV and MAP were similar in TG2-/-mice to age-matched wild type (WT) control mice. Old WT mice exhibited a markedly attenuated EDV as compared to young WT animals. The TG2-/-young and old mice had enhanced EDV responses (p<0.01) as compared to WT mice. There was a significant increase in TG2 crosslinks by IHC in WT old group compared to Young, with no stain in the TG2-/-animals. Optical microscopy examination of Old WT mice aorta showed thinning and fragmentation of elastic laminae. Young WT mice, old and young TG2-/-mice presented regularly arranged and parallel elastic laminae of the tunica media.Conclusion:The genetic suppression of TG2 delays the age-induced endothelial dysfunction and histological modifications.(AU)


Assuntos
Animais , Camundongos , Transglutaminases/uso terapêutico , Camundongos Knockout , Artérias/patologia , Fatores Etários , Fatores Relaxantes Dependentes do Endotélio , Envelhecimento
6.
Anim. Reprod. (Online) ; 15(1): 45-50, Jan.-Mar. 2018. ilus, tab
Artigo em Inglês | VETINDEX | ID: biblio-1461336

Resumo

Bitches with uteri devoid of endometrial glands should be sterile, and consequently could contribute to the population control of dogs. Considering that an inadequate exposure of the female reproductive system to steroids can lead to the formation of the uterine gland knock-out (UGKO) phenotype in some species, the aim of this study was to evaluate the effect of serial applications of medroxyprogesterone acetate (MPA) from birth until the age of six months on the development of endometrial glands in bitches. For this purpose, 16 female mongrel dogs from different litters were distributed into either an MPA group (n = 8), animals treated with 10 mg kg sc (Promone-E®, Pfizer, Brasil) at 3-week intervals, from day one after birth until the age of six months, or a control group (n = 8), composed of animals that only received a 0.9% NaCl solution in place of MPA. At six months of age, ovariohysterectomy was performed and uterine horn samples were collected for histological and immunohistochemical examinations. The bitches from the MPA-treated group presented a 35% decrease in the number of endometrial glands, a larger diameter of the endometrial glands, a greater epithelial height, as well as a greater thickness of the uterine wall, endometrium, and myometrium. However, no significant differences were observed between the two groups in the expression of ER-α, ER-β, and PR on the surface epithelium and endometrial stroma. Therefore, the serial application of MPA from birth until the age of 6 months do not completely ablate the development of the endometrial glands in bitches, but impair it by 35%.


Assuntos
Animais , Recém-Nascido , Cães , Cães/embriologia , Cães/genética , Endométrio , Progesterona/administração & dosagem , Progesterona/análise
7.
Anim. Reprod. (Online) ; 15(supl. 1): 984-995, set. 2018. ilus
Artigo em Inglês | VETINDEX | ID: biblio-1461415

Resumo

The beginning of this century has witnessed great advances in the understanding of ovarian physiology and embryo development, in the improvement of assisted reproductive technologies (ARTs), and in the arrival of the revolutionary genome editing technology through zygote manipulation. Particularly in sheep and goats, the current knowledge on follicular dynamics enables the design of novel strategies for ovarian control, enhancing artificial insemination and embryo production programs applied to genetic improvement. In vitro embryo production (IVEP) has evolved due to a better understanding of the processes that occur during oocyte maturation, fertilization and early embryo development. Moreover, interesting advances have been achieved in embryo and oocyte cryopreservation, thereby reducing the gap between the bench and on-farm application of IVEP technology. Nevertheless, the major breakthrough of this century has been the arrival of the CRISPR/Cas system for genome editing. By joining diverse disciplines such as molecular biology, genetic engineering and reproductive technologies, CRISPR allows the generation of knock-out and knock-in animals in a novel way never achieved before. The innumerable applications of this disruptive biotechnology are challenging the imagination of those who intend to build the animals of the future.


Assuntos
Animais , Ovinos/embriologia , Transferência Embrionária , Transferência Embrionária/veterinária , Técnicas Reprodutivas , Criopreservação , Criopreservação/veterinária
8.
Anim. Reprod. ; 15(supl. 1): 984-995, set. 2018. ilus
Artigo em Inglês | VETINDEX | ID: vti-20558

Resumo

The beginning of this century has witnessed great advances in the understanding of ovarian physiology and embryo development, in the improvement of assisted reproductive technologies (ARTs), and in the arrival of the revolutionary genome editing technology through zygote manipulation. Particularly in sheep and goats, the current knowledge on follicular dynamics enables the design of novel strategies for ovarian control, enhancing artificial insemination and embryo production programs applied to genetic improvement. In vitro embryo production (IVEP) has evolved due to a better understanding of the processes that occur during oocyte maturation, fertilization and early embryo development. Moreover, interesting advances have been achieved in embryo and oocyte cryopreservation, thereby reducing the gap between the bench and on-farm application of IVEP technology. Nevertheless, the major breakthrough of this century has been the arrival of the CRISPR/Cas system for genome editing. By joining diverse disciplines such as molecular biology, genetic engineering and reproductive technologies, CRISPR allows the generation of knock-out and knock-in animals in a novel way never achieved before. The innumerable applications of this disruptive biotechnology are challenging the imagination of those who intend to build the animals of the future.(AU)


Assuntos
Animais , Ovinos/embriologia , Transferência Embrionária , Transferência Embrionária/veterinária , Técnicas Reprodutivas , Criopreservação , Criopreservação/veterinária
9.
Anim. Reprod. ; 15(1): 45-50, Jan.-Mar. 2018. ilus, tab
Artigo em Inglês | VETINDEX | ID: vti-16410

Resumo

Bitches with uteri devoid of endometrial glands should be sterile, and consequently could contribute to the population control of dogs. Considering that an inadequate exposure of the female reproductive system to steroids can lead to the formation of the uterine gland knock-out (UGKO) phenotype in some species, the aim of this study was to evaluate the effect of serial applications of medroxyprogesterone acetate (MPA) from birth until the age of six months on the development of endometrial glands in bitches. For this purpose, 16 female mongrel dogs from different litters were distributed into either an MPA group (n = 8), animals treated with 10 mg kg sc (Promone-E®, Pfizer, Brasil) at 3-week intervals, from day one after birth until the age of six months, or a control group (n = 8), composed of animals that only received a 0.9% NaCl solution in place of MPA. At six months of age, ovariohysterectomy was performed and uterine horn samples were collected for histological and immunohistochemical examinations. The bitches from the MPA-treated group presented a 35% decrease in the number of endometrial glands, a larger diameter of the endometrial glands, a greater epithelial height, as well as a greater thickness of the uterine wall, endometrium, and myometrium. However, no significant differences were observed between the two groups in the expression of ER-α, ER-β, and PR on the surface epithelium and endometrial stroma. Therefore, the serial application of MPA from birth until the age of 6 months do not completely ablate the development of the endometrial glands in bitches, but impair it by 35%.(AU)


Assuntos
Animais , Recém-Nascido , Cães , Cães/embriologia , Cães/genética , Progesterona/administração & dosagem , Progesterona/análise , Endométrio
10.
Acta cir. bras. ; 33(6): 491-498, jun. 2018. graf
Artigo em Inglês | VETINDEX | ID: vti-734729

Resumo

Purpose: To investigate the possible role of IL-4 signaling pathway in vincristine-induced peripheral neuropathy. Methods: The mouse model of vincristine-induced peripheral neuropathy and interleukin (IL)-4 knockout mice were utilized to investigate the possible role of IL-4 signaling pathway in vincristine-induced peripheral neuropathy. Vincristine induced increased sensitivity to mechanical stimulation was measured by von Frey hair test 7 and 14 days after intraperitoneal administration of 0.1 mg/kg vincristine in mice. Relative expression levels of cytokines were detected by quantitative real-time PCR. STAT6 expression following vincristine treatment was assessed with western blotting. Results: We discovered that IL-4/STAT6 signaling was down-regulated in vincristine-treated mice. Deletion of IL-4 in mice increased the sensitivity to mechanical allodynia. IL-4 knockout mice also produced more pro-inflammatory cytokines, including IL-1β and TNF-α. Notably, co-administration of exogenous recombination IL-4 significantly prevented vincristine-induced mechanical allodynia. Conclusion: Anti-inflammatory cytokine IL-4 protects rodent model from vincristine-induced peripheral neuropathy via the stimulation of IL-4/STAT6 signaling and inhibition of the pro-inflammatory cytokines.(AU)


Assuntos
Animais , Masculino , Camundongos , Interleucina-4/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Doenças do Sistema Nervoso Periférico/tratamento farmacológico , Vincristina/efeitos adversos , Modelos Animais
11.
Anim. Reprod. ; 15(supl. 1): 660-679, set. 2018. ilus, graf
Artigo em Inglês | VETINDEX | ID: vti-19648

Resumo

Selection of a single dominant follicle from a cohort of growing follicles is a unique biological process, a key step in female reproductive function in monovular species, and lies at the core of reproductive technologies in cattle. Follicle growth and the number of follicles that ovulate are regulated by precise endocrine, paracrine, and autocrine mechanisms. Most of our current understanding about follicle selection focuses on the role of FSH, LH, and the IGF family in follicle growth and selection of the dominant follicle. However, more recently the role of members of the TGF-ß family has been highlighted, particularly in high fecundity genotypes in sheep. Intercellular signaling between the oocyte and granulosa cells (GC) regulates proliferation and differentiation due to actions of bone morphogenetic protein 15 (BMP15) and growth and differentiation factor 9 (GDF9) within the follicle. Mutations that either knockout or reduce the activity of BMP15 or GDF9 have been found to increase ovulation rate in heterozygotes and generally cause severe follicle abnormalities in homozygotes. A mutation in the intracellular kinase domain of the BMPR1B receptor (Booroola fecundity gene) increases ovulation rate in heterozygotes with further increases in ovulation in homozygotes. The physiological mechanisms linking these mutations to increased ovulation rates are still not well defined. A recently identified high fecundity bovine genotype, Trio, causes increased expression of SMAD6, an intracellular inhibitor of the BMP15/GDF9 signalling pathways. This bovine model has provided insights into the mechanisms associated with selection of multiple dominant follicles and multiple ovulations in carriers of fecundity alleles. The present review focuses on the mechanisms involved in follicle selection in ruminants with a special emphasis on the contribution made by multiple ovulation models in both cattle and sheep. The evaluation of multiple ovulation models in ruminants has allowed us to construct a new physiological model that relates changes in the BMP15/GDF9 signalling pathways to the physiological changes that result in selection of multiple dominant follicles. This model is characterized by acquisition of dominance at a smaller follicle size but at a similar time in the follicular wave with multiple follicles acquiring dominance in a hierarchal sequence, delaying FSH suppression and, thus allowing additional follicles to continue to grow and acquire dominance.(AU)


Assuntos
Animais , Bovinos , Ruminantes/fisiologia , Fase Folicular , Fertilidade , Hormônio Foliculoestimulante/análise , Bovinos/fisiologia
12.
Anim. Reprod. (Online) ; 15(supl. 1): 660-679, set. 2018. ilus, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461388

Resumo

Selection of a single dominant follicle from a cohort of growing follicles is a unique biological process, a key step in female reproductive function in monovular species, and lies at the core of reproductive technologies in cattle. Follicle growth and the number of follicles that ovulate are regulated by precise endocrine, paracrine, and autocrine mechanisms. Most of our current understanding about follicle selection focuses on the role of FSH, LH, and the IGF family in follicle growth and selection of the dominant follicle. However, more recently the role of members of the TGF-ß family has been highlighted, particularly in high fecundity genotypes in sheep. Intercellular signaling between the oocyte and granulosa cells (GC) regulates proliferation and differentiation due to actions of bone morphogenetic protein 15 (BMP15) and growth and differentiation factor 9 (GDF9) within the follicle. Mutations that either knockout or reduce the activity of BMP15 or GDF9 have been found to increase ovulation rate in heterozygotes and generally cause severe follicle abnormalities in homozygotes. A mutation in the intracellular kinase domain of the BMPR1B receptor (Booroola fecundity gene) increases ovulation rate in heterozygotes with further increases in ovulation in homozygotes. The physiological mechanisms linking these mutations to increased ovulation rates are still not well defined. A recently identified high fecundity bovine genotype, Trio, causes increased expression of SMAD6, an intracellular inhibitor of the BMP15/GDF9 signalling pathways. This bovine model has provided insights into the mechanisms associated with selection of multiple dominant follicles and multiple ovulations in carriers of fecundity alleles. The present review focuses on the mechanisms involved in follicle selection in ruminants with a special emphasis on the contribution made by multiple ovulation models in both cattle and sheep. The evaluation of multiple ovulation models in ruminants has allowed us to construct a new physiological model that relates changes in the BMP15/GDF9 signalling pathways to the physiological changes that result in selection of multiple dominant follicles. This model is characterized by acquisition of dominance at a smaller follicle size but at a similar time in the follicular wave with multiple follicles acquiring dominance in a hierarchal sequence, delaying FSH suppression and, thus allowing additional follicles to continue to grow and acquire dominance.


Assuntos
Animais , Bovinos , Bovinos/fisiologia , Fase Folicular , Fertilidade , Hormônio Foliculoestimulante/análise , Ruminantes/fisiologia
13.
Arq. bras. med. vet. zootec. (Online) ; 69(6): 1565-1572, nov.-dez. 2017. graf
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-910569

Resumo

This study aimed to evaluate aspects of host immune response using an experimental infection model of Corynebacterium pseudotuberculosis (CP) in C57/Black6 wild-type and knockout for nitric oxide (KO-NO) mice. 28 mice were evaluated: 4 wild-type controls; 10 wild-type infected with CP; 4 KO-NO controls; 10 KO-NO infected with CP. Infection procedures were carried out by intraperitoneal inoculation using 107. Infected C57/Black6 KO-NO mice began to die after the 5° day post-inoculation, up until the 14º day. Neutrophils were found in increased numbers in the infiltrate of KO-NO murine peritoneal cavities. Examination of splenic tissue revealed an accumulation of lymphocytes, predominantly CD8 T-cells, in experimental animal groups. KO-NO animals were found to have a predominance of granulomas 7 days post-inoculation, primarily in the lymph nodes. In addition, greater amounts of bacteria were recovered from the mesenteric lymph nodes of KO-NO mice. There was no statistically significant difference in the levels of total IgG and its subclasses 14 days post-inoculation between KO-NO and wild groups. The results suggest the importance of nitric oxide in the process of controlling CP infection, as KO-NO animals were observed to be markedly more affected by infection with this bacterium.(AU)


O objetivo deste estudo foi avaliar os aspectos da resposta imune do hospedeiro, mediante o uso de um modelo experimental de infecção de Corynebacterium pseudotuberculosis (CP) em camundongos C57/Black6 do tipo selvagem e em C57/Black6 knockout para o óxido nítrico (KO-NO). Foram avaliados 28 camundongos: quatro controles de tipo selvagem; 10 do tipo selvagem infectados com CP; quatro controles KO-NO; e 10 KO-NO infectados com CP. A infecção foi realizada via intraperitoneal, usando-se 107. Os animais C57/Black6 KO-NO infectados começaram a vir a óbito no quinto dia pós-inoculação, o que aconteceu até o 14º dia. Um número maior de neutrófilos foi encontrado na sua cavidade peritoneal. O exame do baço revelou um acúmulo de linfócitos, predominantemente células T CD8, nos grupos de animais experimentais. Nos animais KO-NO, foi observada a presença de granulomas, sete dias pós-inoculação, principalmente nos gânglios linfáticos. Além disso, uma maior quantidade de bactérias foi detectada dos linfonodos mesentéricos desses animais. Não houve diferença estatisticamente significante nos níveis séricos IgG total e em suas subclasses aos 14 dias pós-inoculação nos grupos KO-NO e selvagem. Os resultados obtidos sugerem a importância do óxido nítrico no processo de controle da infecção por CP.(AU)


Assuntos
Animais , Camundongos , Corynebacterium pseudotuberculosis/imunologia , Linfadenite/prevenção & controle , Camundongos Knockout/imunologia , Óxido Nítrico/uso terapêutico , Linfadenite/veterinária
14.
Arq. bras. med. vet. zootec. (Online) ; 69(6): 1565-1572, Nov.-Dez. 2017. graf
Artigo em Inglês | VETINDEX | ID: vti-735001

Resumo

This study aimed to evaluate aspects of host immune response using an experimental infection model of Corynebacterium pseudotuberculosis (CP) in C57/Black6 wild-type and knockout for nitric oxide (KO-NO) mice. 28 mice were evaluated: 4 wild-type controls; 10 wild-type infected with CP; 4 KO-NO controls; 10 KO-NO infected with CP. Infection procedures were carried out by intraperitoneal inoculation using 107. Infected C57/Black6 KO-NO mice began to die after the 5° day post-inoculation, up until the 14º day. Neutrophils were found in increased numbers in the infiltrate of KO-NO murine peritoneal cavities. Examination of splenic tissue revealed an accumulation of lymphocytes, predominantly CD8 T-cells, in experimental animal groups. KO-NO animals were found to have a predominance of granulomas 7 days post-inoculation, primarily in the lymph nodes. In addition, greater amounts of bacteria were recovered from the mesenteric lymph nodes of KO-NO mice. There was no statistically significant difference in the levels of total IgG and its subclasses 14 days post-inoculation between KO-NO and wild groups. The results suggest the importance of nitric oxide in the process of controlling CP infection, as KO-NO animals were observed to be markedly more affected by infection with this bacterium.(AU)


O objetivo deste estudo foi avaliar os aspectos da resposta imune do hospedeiro, mediante o uso de um modelo experimental de infecção de Corynebacterium pseudotuberculosis (CP) em camundongos C57/Black6 do tipo selvagem e em C57/Black6 knockout para o óxido nítrico (KO-NO). Foram avaliados 28 camundongos: quatro controles de tipo selvagem; 10 do tipo selvagem infectados com CP; quatro controles KO-NO; e 10 KO-NO infectados com CP. A infecção foi realizada via intraperitoneal, usando-se 107. Os animais C57/Black6 KO-NO infectados começaram a vir a óbito no quinto dia pós-inoculação, o que aconteceu até o 14º dia. Um número maior de neutrófilos foi encontrado na sua cavidade peritoneal. O exame do baço revelou um acúmulo de linfócitos, predominantemente células T CD8, nos grupos de animais experimentais. Nos animais KO-NO, foi observada a presença de granulomas, sete dias pós-inoculação, principalmente nos gânglios linfáticos. Além disso, uma maior quantidade de bactérias foi detectada dos linfonodos mesentéricos desses animais. Não houve diferença estatisticamente significante nos níveis séricos IgG total e em suas subclasses aos 14 dias pós-inoculação nos grupos KO-NO e selvagem. Os resultados obtidos sugerem a importância do óxido nítrico no processo de controle da infecção por CP.(AU)


Assuntos
Animais , Camundongos , Corynebacterium pseudotuberculosis/imunologia , Linfadenite/prevenção & controle , Camundongos Knockout/imunologia , Óxido Nítrico/uso terapêutico , Linfadenite/veterinária
15.
Artigo em Inglês | VETINDEX | ID: vti-20192

Resumo

ABSTRACT This study aimed to evaluate aspects of host immune response using an experimental infection model of Corynebacterium pseudotuberculosis (CP) in C57/Black6 wild-type and knockout for nitric oxide (KO-NO) mice. 28 mice were evaluated: 4 wild-type controls; 10 wild-type infected with CP; 4 KO-NO controls; 10 KO-NO infected with CP. Infection procedures were carried out by intraperitoneal inoculation using 107. Infected C57/Black6 KO-NO mice began to die after the 5° day post-inoculation, up until the 14º day. Neutrophils were found in increased numbers in the infiltrate of KO-NO murine peritoneal cavities. Examination of splenic tissue revealed an accumulation of lymphocytes, predominantly CD8 T-cells, in experimental animal groups. KO-NO animals were found to have a predominance of granulomas 7 days post-inoculation, primarily in the lymph nodes. In addition, greater amounts of bacteria were recovered from the mesenteric lymph nodes of KO-NO mice. There was no statistically significant difference in the levels of total IgG and its subclasses 14 days post-inoculation between KO-NO and wild groups. The results suggest the importance of nitric oxide in the process of controlling CP infection, as KO-NO animals were observed to be markedly more affected by infection with this bacterium.


RESUMO O objetivo deste estudo foi avaliar os aspectos da resposta imune do hospedeiro, mediante o uso de um modelo experimental de infecção de Corynebacterium pseudotuberculosis (CP) em camundongos C57/Black6 do tipo selvagem e em C57/Black6 knockout para o óxido nítrico (KO-NO). Foram avaliados 28 camundongos: quatro controles de tipo selvagem; 10 do tipo selvagem infectados com CP; quatro controles KO-NO; e 10 KO-NO infectados com CP. A infecção foi realizada via intraperitoneal, usando-se 107. Os animais C57/Black6 KO-NO infectados começaram a vir a óbito no quinto dia pós-inoculação, o que aconteceu até o 14º dia. Um número maior de neutrófilos foi encontrado na sua cavidade peritoneal. O exame do baço revelou um acúmulo de linfócitos, predominantemente células T CD8, nos grupos de animais experimentais. Nos animais KO-NO, foi observada a presença de granulomas, sete dias pós-inoculação, principalmente nos gânglios linfáticos. Além disso, uma maior quantidade de bactérias foi detectada dos linfonodos mesentéricos desses animais. Não houve diferença estatisticamente significante nos níveis séricos IgG total e em suas subclasses aos 14 dias pós-inoculação nos grupos KO-NO e selvagem. Os resultados obtidos sugerem a importância do óxido nítrico no processo de controle da infecção por CP.

16.
Tese em Português | VETTESES | ID: vtt-220167

Resumo

O gênero Sarcocystis inclui aproximadamente 200 espécies, e destas, apenas 17 são conhecidas por parasitar serpentes. A descoberta de Sarcocystis nesbitti, que possui importância para saúde pública, impulsionou o desenvolvimento de novas pesquisas em serpentes. No Brasil, as pesquisas sobre Sarcocystis spp. em serpentes são escassas. Assim, objetivou-se com o corrente trabalho verificar a infecção por Sarcocystis spp. em Boa constrictor no Município de Salvador, e consequentemente, caracterizar as formas parasitárias por métodos morfológicos e moleculares. Foram avaliadas 65 amostras de fezes de jiboias obtidas no município de Salvador, empregando-se a técnica de centrifugo-flutuação em solução de sacarose. A frequência observada para a infecção foi de 1,53% (1/65). A partir do único animal infectado, foi possível analisar seis amostras fecais, observando-se a oscilação na quantidade de esporocistos eliminados, bem como uma frequência intermitente de eliminação. A avaliação morfométrica revelou a presença de esporocistos com dimensões significativamente diferentes (p < 0,05). A experimentação in vivo foi desenvolvida com duas espécies de roedores Mus musculus (camundongos Balb/c, Balb/c Nude e nocauteado INF- -KO) e Rattus norvegicus (Rato Wistar), não ocorrendo a infecção nos exemplares de camundongos. Como os tecidos dos ratos ainda estão sob processamento histológico, não foi possível gerar dados até o presente momento. A análise molecular de esporocistos da jiboia utilizando os marcadores 18S, 28S, e ITS-1 demonstraram alterações no cromatograma compatíveis com heterozigose. Ainda assim, após a edição das sequências, e associados ao marcador mitocrondial COX-1, as sequências revelaram alta identidade com Sarcocystis spp. isoladas de esporocistos de serpentes e sarcocistos de roedores. A análise filogenética da amostra da jiboia não demonstrou correspondência com outros organismos conhecidos, sendo designada como inédita. Com o intuito de obter melhores resultados para os loci ribossômicos, a clonagem do material está sendo desenvolvida e será acrescentada ao artigo para publicação.


The genus Sarcocystis has approximately 200 species and only 17 of them are known to infect snakes. The discovery of Sarcocystis nesbitti, which has public health relevance, boosted the development of new investigations in snakes. In Brazil, studies on Sarcocystis spp. infection in snakes are scarce. Then, the current work aimed to test the role of common boa (Boa sp.) from Salvador city as Sarcocystis spp. hosts, and consequently, to characterize the stages of the parasite by morphological and molecular methods. Fecal samples obtained from 65 boa from Salvador city were examined using a centrifugal-flotation method. The frequency of infection in the tested snakes was 1.53% (1/65). From the single positive animal, six fecal samples were available for analysis. Intermittent shedding of sporocysts and variation in the number of excreted sporocysts occurred in this snake. Morphomethric evaluation of the parasites revealed significant differences in sporocysts sizes (p < 0.05). In vivo experimentation was carried out with two species of rodents Mus musculus (Balb/c, Balb /c Nude and knockout INF- -KO mice) and Rattus norvegicus (Wistar rat), resulting in no infection in the mice. As the tissues of the rats are still under histological processing, it has not been possible to generate data so far. Molecular analysis of sporocysts derived from the common boa using 18S, 28S, and ITS-1 markers demonstrated changes in the chromatogram compatible with heterozygosis. Still, after editing these sequences, and associating them with the mitochondrial COX-1 marker, the sequences revealed a high identity with Sarcocystis spp. isolated from snake sporocysts and rodent sarcocysts. Phylogenetic analysis revealed that the common boa sample does not possess correspondence with any other known organism, representing a novel sequence. In order to obtain better results for ribosomal loci, the sample is being cloned and the results will be added to the article for publication.

17.
Tese em Português | VETTESES | ID: vtt-222396

Resumo

O presente trabalho compreende o Estudo de diferentes cepas de Neospora caninum do Brasil e da Escócia quanto às suas características genéticas, avaliação de patogenicidade, virulência e possibilidade de isolamento. O primeiro capítulo, consiste de um artigo publicado no periódico internacional Veterinary Parasitology e descreve uma tentativa de isolamento de cepas do parasito utilizando tecido placentário de cabras naturalmente infectadas, em um bioensaio utilizando gerbis. Apesar dos gerbis serem descritos na literatura como sendo altamente susceptíveis à infecção por N. caninum, somente foi obtido sucesso na infecção de animais após imunossupressão com altas doses de metilpredinisolona. Animais imunossuprimidos apresentaram sinais clínicos de emagrecimento, pelos arrepiados e alterações de propriocepção como Head tilt. Análises histopatológicas revelaram reações inflamatórias no encéfalo, consistindo de infiltrados inflamatórios mononucleares multifocais associados à manguitos perivasculares e estruturas parasitárias semelhantes a cistos e taquizoítos, juntamente com áreas de necrose associadas à infiltrado inflamatório de macrófagos e reação de cicatrização em musculatura esquelética. Apesar do sucesso na infecção dos gerbis, não foi possível obter o isolamento do parasito em cultivo celular em uma primeira tentativa, o que levou à segunda parte do trabalho. O capítulo dois consiste de um artigo publicado no periódico intenracional Experimental Parasitology. Foram utilizados tecidos dos gerbis infectados descritos anteriormente e novas amostras de sistema nervoso central de caprinos naturalmente infectados em um bioensaio em camundongos C57BL6 Knockout para o gene Interferon gamma (camundongos knockout- KO). Os camundongos Knockout por serem incapazes de desenvolver uma resposta imune adequada ao N. caninum, permitiram a rápida replicação de taquizoítos principalmente em macrófagos peritoneais, os camundongos apresentaram acentuada perde de peso, letargia e ascite 15 a 30 dias após inoculação, quando foram eutanasiados e o líquido peritoneal utilizado para infecção de monocamadas de células VERO. A grande quantidade de taquizoítos no líquido peritoneal de camundongos KO permitiu uma rápida adaptação e crescimento dos mesmos em cultura de células, dando origem a dois isolados de cabras nomeados NC-Goat1 e NC-Goat2. Os dois isolados foram testados quanto à sua Patogenicidade em um modelo de infecção experimental em camundongos BALB/c, amplamente utilizado na literatura. As cepas não apresentaram alta patogenicidade e virulência no modelo de infecção de camundongos, apesar das duas cepas terem sido associadas a doença clínica em caprinos, com abortos, natimortos e encefalite em caprinos jovens e adultos. O terceiro capítulo apresenta resultados de um estudo genético de genotipagem dos isolados NC-Goat1 e NC-Goat2, amostras clínicas de caprinos e bovinos do sul de Minas Gerais e amostras de bovinos da região de Dumfries e Galloway na Escócia. Utilizando a técnica de genotipagem por microssatélites, análises filogenéticas evidenciaram que as cepas de N. caninum presentes nos isolados e nas amostras clínicas de caprinos tinha relação próxima com os isolados brasileiros NC-SP1, NC-Bahia e NC-Goiás, enquanto que as amostras de bovinos de minas gerais não se correlacionaram proximamente às outras. Analisando as amostras provenientes de rebanhos escoceses, todos tenderam a se relacionar proximamente entre elas e com outras amostras escocesas descritas anteriormente na literatura. Análises adicionais utilizando marcadores microssatélites descritos pelo grupo de pesquisa do Instituto Moredun, evidenciaram genótipos mistos associados ao isolado padrão de N. caninum (NC-1), sugerindo que o isolado poderia ter sido composto por múltiplos genótipos ao isolamento e à medida que foi replicado em cultura celular convergiu em uma população mais homogênea de parasitos.


The present work comprises the study of different strains of Neospora caninum, from Brazil and Scotland, regarding their genetic characteristics, pathogenicity, virulence and attempts at isolation. The first chapter consists of an article published in the journal Veterinary Parasitology and describes an attempt to isolate parasite strains using placental tissue from naturally infected goats in a bioassay using gerbils. Although gerbils are described in the literature as highly susceptible to N. caninum infection, successful infection of animals has only been achieved after immunosuppression with high doses of methylpredinisolone. Immunosuppressed animals showed clinical signs of weight loss, fur and proprioception changes such as Head tilt. Histopathological analysis revealed inflammatory reactions in the brain consisting of multifocal mononuclear inflammatory infiltrates associated with perivascular cuffs, cyst-like and tachyzoite-like parasitic structures, along with areas of necrosis associated with macrophage inflammatory infiltrate and skeletal muscle scar formation. Despite the success in gerbil infection, it was not possible to obtain isolation of the parasite in cell culture in a first attempt, which led to the second part of the work. The chapter two consists of an article published in the journal Experimental Parasitology. Tissues from the previously described infected gerbils and new samples from the central nervous system of naturally infected goats were used in a bioassay using C57BL6 interferon gamma Knockout mouse (KO mice). Knockout mice, being unable to develop an adequate immune response to N. caninum, allowed the rapid replication of tachyzoites mainly in peritoneal macrophages, showing marked weight loss, lethargy and ascites 15 to 30 days after inoculation, when they were euthanized, and the peritoneal fluid used for infection of VERO cell monolayers. The large amount of tachyzoites in the peritoneal fluid of KO mice allowed their rapid adaptation and growth in cell culture, resulting in two goat isolates named NCGoat1 and NC-Goat2. Both isolates were tested for pathogenicity in a widely used experimental BALB/c mouse infection model. The strains did not show high pathogenicity and virulence in the mouse infection model, although both strains were associated with clinical disease in goats, with abortions, stillbirths and encephalitis in young and adult goats. The third chapter presents the results of a genetic genotyping study of the isolates NCGoat1 and NC-Goat2, clinical samples from goats and cattle from southern Minas Gerais and samples from cattle from the Dumfries and Galloway region of Scotland. Using the microsatellite genotyping technique, phylogenetic analyzes showed that N. caninum strains present in the isolates and clinical samples of goats had a close relationship with the Brazilian isolates NC-SP1, NC-Bahia and NC-Goiás, while the samples from Minas Gerais cattle did not correlate closely to the others. Analysing samples from Scottish herds, all tended to relate closely to each other and to other Scottish samples previously described in the literature. Additional analyses using microsatellite markers described by the Moredun Institute research group showed mixed genotypes associated with the standard N. caninum isolate (NC-1), suggesting that the isolate could have been composed of multiple genotypes on isolation and as it was replicated in cell culture they converged to a more homogeneous population of parasites.

18.
Tese em Português | VETTESES | ID: vtt-220541

Resumo

Aedes aegypti é um mosquito amplamente distribuído no mundo que possui grande importância médica, apresentando competência vetorial para diversos vírus como da Febre Amarela (YFV), Dengue (DENV), Chikungunya (CHIKV) e Zika (ZIKV). Diversos métodos de controle foram desenvolvidos na tentativa de combater esses parasitos, no entanto, a utilização rotineira de inseticidas gerou a resistência múltipla à esses produtos. O presente estudo teve como objetivo verificar a susceptibilidade de Ae. aegypti provenientes de seis bairros do município de Castanhal-PA, a permetrina. Para isso, foram coletados exemplares de campo por meio de busca ativa por criadouros. As larvas e pupas foram transferidas para recipientes com água destilada e mantidas em laboratório com temperatura de 26 28ºC, e 12 horas de luminosidade para obtenção de F1. O bioensaio inseticida foi realizado com a geração F1 de Ae. aegypti, utilizando-se permetrina na concentração de 15µg/mL. Os mosquitos testados foram sensíveis ao efeito nocaute da permetrina. A mortalidade das populações de campo no tempo diagnóstico variou de 98,3% a 100%, tendo-se observado em apenas dois bairros a presença de mosquitos que morreram após o tempo diagnóstico. No entanto, a taxa de sobrevivência foi zero após 120 minutos de observação. Dessa forma, concluímos que todas as populações de Ae. aegypti testadas são sensíveis à permetrina.


Aedes aegypti is a mosquito widely distributed in the world that has great medical importance, presenting vectorial competence for various viruses such as Yellow Fever (YFV), Dengue (DENV), Chikungunya (CHIKV) and Zika (ZIKV). Several control methods have been developed in an attempt to combat these parasites, however, the routine use of insecticides has generated multiple insecticide resistance to these products. The present study aimed to verify the susceptibility of Ae. aegypti from six neighborhoods in the municipality of Castanhal-PA, permethrin. For this purpose, field specimens were collected through an active search for breeding sites. The larvae and pupae were transferred to containers with distilled water and kept in the laboratory with a temperature of 26 - 28ºC, and 12 hours of light to obtain F1. The insecticidal bioassay was carried out with the F1 generation of Ae. aegypti, using permethrin at a concentration of 15 µg / mL. The tested mosquitoes were sensitive to the knockout effect of permethrin. The mortality of field populations at the time of diagnosis varied from 98.3% to 100%, having been observed in only two neighborhoods the presence of mosquitoes that died after the time of diagnosis. However, the survival rate was zero after 120 minutes of observation. Thus, we conclude that all populations of Ae. aegypti tested, from the municipality of Castanhal, Pará, are sensitive to permethrin.

19.
Anim. Reprod. (Online) ; 12(1): 28-34, Jan.-Mar.2015. ilus, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461141

Resumo

The major endocrine regulators of the female reproductive tract are 17β-estradiol (E2) and progesterone (P4). This review discusses our recent work related to the roles of E2 and P4 and their receptors, estrogen receptor 1 (ESR1) and progesterone receptor (PR), respectively, in the neonatal uterus. Neonatal uterine cells in mice are mitogenically responsive to estrogens, but neonatal ovariectomy does not inhibit pre-weaning uterine cell proliferation, indicating that this process does not require endogenous estrogens. Neonatal terine cell proliferation could result from ligand-independent growth factor activation of ESR1, or be independent of ESR1 neonatally despite its obligatory role in adult uterine epithelial proliferation. To determine the role of ESR1 in uterine development, we analyzed cell proliferation and uterine gland development (adenogenesis) in wild-type (WT) and Esr1 knockout (Esr1KO) mice postnatally. Our results indicate that pre-weaning uterine cell proliferation and adenogenesis are independent of ESR1, but these processes become dependent on E2/ESR1 signaling for maintenance and further proliferation and uterine growth during puberty. How pre-weaning uterine cell proliferation and adenogenesis occur independently of E2/ESR1 signaling remains unknown, but ligand-independent activation of ESR1 is not involved in this process. The synthetic glucocorticoid dexamethasone (Dex) inhibits luminal epithelial (LE) proliferation in neonatal mouse uteri, but it has been unclear whether Dex effects were mediated by glucocorticoid receptor (GR) and/or PR. We have used PR knockout (PRKO) mice to test whether PR is required for Dex inhibition of LE proliferation. Our results indicate that maximal inhibitory Dex effects on uterine LE proliferation require PR, possibly reflecting Dex crosstalk with PR.


Assuntos
Feminino , Animais , Camundongos , Camundongos , Estrogênios/análise , Progesterona/análise , Proliferação de Células
20.
Anim. Reprod. ; 12(1): 28-34, Jan.-Mar.2015. ilus, graf
Artigo em Inglês | VETINDEX | ID: vti-745423

Resumo

The major endocrine regulators of the female reproductive tract are 17β-estradiol (E2) and progesterone (P4). This review discusses our recent work related to the roles of E2 and P4 and their receptors, estrogen receptor 1 (ESR1) and progesterone receptor (PR), respectively, in the neonatal uterus. Neonatal uterine cells in mice are mitogenically responsive to estrogens, but neonatal ovariectomy does not inhibit pre-weaning uterine cell proliferation, indicating that this process does not require endogenous estrogens. Neonatal terine cell proliferation could result from ligand-independent growth factor activation of ESR1, or be independent of ESR1 neonatally despite its obligatory role in adult uterine epithelial proliferation. To determine the role of ESR1 in uterine development, we analyzed cell proliferation and uterine gland development (adenogenesis) in wild-type (WT) and Esr1 knockout (Esr1KO) mice postnatally. Our results indicate that pre-weaning uterine cell proliferation and adenogenesis are independent of ESR1, but these processes become dependent on E2/ESR1 signaling for maintenance and further proliferation and uterine growth during puberty. How pre-weaning uterine cell proliferation and adenogenesis occur independently of E2/ESR1 signaling remains unknown, but ligand-independent activation of ESR1 is not involved in this process. The synthetic glucocorticoid dexamethasone (Dex) inhibits luminal epithelial (LE) proliferation in neonatal mouse uteri, but it has been unclear whether Dex effects were mediated by glucocorticoid receptor (GR) and/or PR. We have used PR knockout (PRKO) mice to test whether PR is required for Dex inhibition of LE proliferation. Our results indicate that maximal inhibitory Dex effects on uterine LE proliferation require PR, possibly reflecting Dex crosstalk with PR. (AU)


Assuntos
Animais , Feminino , Camundongos , Estrogênios/análise , Progesterona/análise , Proliferação de Células , Camundongos
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