RESUMO
At present, the zebrafish embryo is increasingly used as an alternative animal model to screen for developmental toxicity after exposure to xenobiotics. Since zebrafish embryos depend on their own drug-metabolizing capacity, knowledge of their intrinsic biotransformation is pivotal in order to correctly interpret the outcome of teratogenicity assays. Therefore, the aim of this in vitro study was to assess the activity of cytochrome P450 (CYP)-a group of drug-metabolizing enzymes-in microsomes from whole zebrafish embryos (ZEM) of 5, 24, 48, 72, 96 and 120 h post-fertilization (hpf) by means of a mammalian CYP substrate, i.e., benzyloxy-methyl-resorufin (BOMR). The same CYP activity assays were performed in adult zebrafish liver microsomes (ZLM) to serve as a reference for the embryos. In addition, activity assays with the human CYP3A4-specific Luciferin isopropyl acetal (Luciferin-IPA) as well as inhibition studies with ketoconazole and CYP3cide were carried out to identify CYP activity in ZLM. In the present study, biotransformation of BOMR was detected at 72 and 96 hpf; however, metabolite formation was low compared with ZLM. Furthermore, Luciferin-IPA was not metabolized by the zebrafish. In conclusion, the capacity of intrinsic biotransformation in zebrafish embryos appears to be lacking during a major part of organogenesis.
Assuntos
Citocromo P-450 CYP3A/metabolismo , Sondas Moleculares/metabolismo , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , Animais , Biotransformação/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Feminino , Luciferina de Vaga-Lumes/metabolismo , Humanos , Cetoconazol/farmacologia , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Oxazinas/metabolismo , Recombinação Genética/genética , Especificidade por Substrato/efeitos dos fármacosRESUMO
Zebrafish embryos are increasingly used for developmental toxicity screening of candidate drugs and are occasionally co-incubated with a metabolic activation system at 32°C for 1, 2 or 4h, depending on their developmental stage. As this temperature is higher than the optimal temperature for zebrafish embryonic development (26-28.5°C), we investigated whether continuous incubation of zebrafish embryos from 2.5 until 96h post fertilization (hpf) at high temperatures (30.5-36.5°C) causes malformations. At 32.5°C tail malformations were observed as early as 24hpf, and these became even more prominent at 34.5 and 36.5°C. Cardiovascular and head malformations, edema and blood accumulations throughout the body were present at 36.5°C. Finally, temperatures higher than 28.5°C accelerated embryonic development except for 36.5°C, at which a lower hatching rate and hatching enzyme activity were observed. In conclusion, incubation of zebrafish embryos at 32.5°C and above from 2.5 until 96hpf causes malformations as early as 24hpf.
Assuntos
Bioensaio , Embrião não Mamífero/anormalidades , Temperatura , Testes de Toxicidade/métodos , Peixe-Zebra/anormalidades , Animais , Catepsina L/metabolismo , Embrião não Mamífero/enzimologia , Fatores de Tempo , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/metabolismoRESUMO
Anatomical dissection of the temporal regions of 62 cattle demonstrated that lymph nodes and hemal nodes are present in 89% of the animals (bilaterally in 65% and unilaterally in 24% of the cases). Lymph nodes accounted for 60% and hemal nodes for 40% of all examined nodes. They are nearly always round with diameters ranging from 1 to 9mm. Injections of India ink showed that their drainage area consists of the forehead, the upper eyelid, the base of the horn and the temporal muscle. Immunohistochemistry and flow cytometry revealed that the distribution and percentages of the different cell populations in the lymph nodes and hemal nodes are similar to other cranial lymph nodes. Based on its anatomical location the name temporal lymph node (lymphonodus temporalis) is proposed.
Assuntos
Bovinos/anatomia & histologia , Tecido Linfoide/anatomia & histologia , Animais , Dissecação/veterinária , Citometria de Fluxo/veterinária , Imuno-Histoquímica/veterináriaRESUMO
Three dogs were presented for clinical examination with bilateral buccal nodules which were identified as enlarged buccal lymph nodes. As little is known about this pathology, 150 dogs were examined by anatomical dissection for the presence of buccal lymph nodes. They were found in 13 dogs, occurring bilaterally in six dogs and unilaterally in seven dogs. Two buccal lymph nodes were bilobulated and one was double. The lymph nodes were always located dorsal to the zygomatic muscle and rostral to the masseter muscle in the region where the superior labial vein drains into the facial vein. Histology demonstrated a large amount of intranodal adipose tissue scattered throughout the lymphoid tissue. The canine buccal lymph node should not be confused with the accessory parotid or ventral buccal salivary gland and is clinically important as it can enlarge due to tumour metastasis or inflammation of the buccal region.