Chewing-like as a possible behavior of persistent orofacial pain in the experimental temporomandibular dysfunction / Mastigação (chewing-like) como possível comportamento de dor orofacial persistente na disfunção temporomandibular experimental.

BACKGROUND AND OBJECTIVES: Stimulations with formalin in the orofacial region can be related to transient or subacute nociceptive activity and behavioral changes. The evaluation of behavioral changes induced by persistent or chronic irritating nociceptive substance has not yet been described.METHOD: Complete Freund's Adjuvant (CFA) was injected in the temporomandibular joint (TMJ) region of rats and analyzed comparing it to the groups treated with saline and 2.5% formalin. In addition, behaviors such as grooming, freezing, rest/sleeping and chewing-like were electronically observed and quantified.RESULTS: It was shown that the chewing-like behavior was significantly increased and that it was inhibited by indometacin (5 mg/kg) and morphine (4 mg/kg). CONCLUSION: These results suggest that chewing-like may be a possible behavior of persistent or chronic orofacial pain, and may be a tool for clinical-pharmacological studies.

JUSTIFICATIVA E OBJETIVOS: Estímulos com formalina na região orofacial podem estar relacionados com a atividade nociceptiva e as alterações comportamentais transitórias ou subagudas. A avaliação de comportamentos sob ação de substância irritante nociceptiva persistente e crônica ainda não foi descrita.MÉTODO: Foi feita injeção de adjuvante completo de Freund (ACF) na região da articulação temporomandibular (ATM) de ratos e foi analisada comparando-a com os grupos tratados com salina e formalina a 2,5%. Além disso, foram observados e quantificados eletronicamente os comportamentos grooming, freezing, rest/sleeping e chewing-like (mastigação). RESULTADOS: Observou-se que o comportamento mastigação (chewing-like) estava significativamente aumentado e que ele foi inibido pela indometacina (5 mg/kg) e morfina (4 mg/kg).CONCLUSÃO: Esses resultados sugerem ser o chewing-like um possível comportamento de dor orofacial persistente, oferecendo-se como instrumento para análise clínico-farmacológica

Producción de anticuerpos policlonales IgG contra una proteína con actividad de óxido nítrico sintetasa de Toxoplasma gondii recombinante (NOS-Tg-r) y marcación inmunológica en taquizoítoso / Production of Polyclonal Antibodies against Toxoplasma gondii Recombinant protein with Nitric Oxide Synthase activity and immunologic marking in Tachyzoites

La enzima óxido nítrico sintetasa ha sido estudiada en mamíferos; en los últimos años se ha descrito que existe también en protozoos, pero se desconocen aspectos importantes de su función. Se logró producir anticuerpos policlonales contra la proteína recombinante con actividad de óxido nítrico sintetasa (NOS-Tg-r) de Toxoplasma gondii y realizar marcación inmunológica en taquizoítos. Se usaron dos conejos Nueva Zelanda (Oryctolagus cuniculus)que se inmunizaron por vía intramuscular con NOS-Tg-r, y dos tipos de adyuvantes, hidróxido de aluminio y adyuvante de Freund. Se comprobó la presencia de anticuerpos policlonales con la técnica de ensayo inmunoenzimático indirecto. Los resultados obtenidos mostraron que a NOS-Tg-r con adyuvante de Freund indujo mayor respuesta inmune que la de la NOS-Tg-r con hidróxido aluminio p 0,005). Para verificar si había reacción cruzada, se realizó una prueba ELISA utilizando como antígenos: metaloproteasa de T. gondii recombinante, cisteína proteasa 5 de Entamoeba histolytica recombinante, albúmina al 2%, hidróxido de aluminio y adyuvante de Freund. Los valores obtenidos con sueros preinmunes y contra proteínas alternas no superaron el punto de corte (0,069), lo cual indica que los anticuerpos policlonales obtenidos son específicos para NOS-Tg-r. Se realizó marcación inmunológica en taquizoítos de Toxoplasma gondii con inmunofluorescencia indirecta que mostró una marcación difusa a nivel de citoplasma y confirmó la presencia de esta proteína en los taquizoítos.

The nitric oxide synthase (NOS)is an enzyme well described on mammals but little is known about the role of these enzymes on pathogenic parasites. We produced polyclonal antibodies against a recombinant NOS enzyme from Toxoplasma gondii nd e lso er formed n mmunol locali zation of the enzyme on tachyzoites. We used two New Zealand rabbits (Oryctolagus cuniculus) to perform intramuscular immunization and we used two types of adjuvant: aluminum hydroxide and Freund ’s adjuvant. We tested the generation of polyclonal antibodies by an indirect ELISA assay. The results showed that levels of antibodies were higher in rabbits immunized with Freund ’s adjuvant than with aluminum hydroxide (p =0.005). In order to test cross reactions, we used a recombinant Toxoplasma metallooprotease, a recombinant cysteine protease from Entamoeba histolytica, albumin 2%, hydroxide aluminium and Freund ’s adjuvant as antigens on indirect ELISA assays for the polyclonal serum antibodies. No serum showed absorbances higher than the cutoff level with these antigens, indicating that the polyclonal antibodies were specific for recombinant Toxoplasma NOS. Additionally, we performed immunodetection of the enzyme on Toxoplasma gondii tachyzoites and we obtained a diffuse labeling of the parasite.

Experimental Autoimmune Uveitis induced by Bovine Iris and Ciliary body in Lewis Rat

This study was conducted to develop an animal model of uveitis resembled anterior uveitis in humans after immunization with iris-ciliary body antigen. Male Lewis rats were immunized with the buffer-and detergent insoluble bovine iris-ciliary body mixed with Complete Freund`s adjuvant (CFA) and Pertussis toxin(PTX). A soluble fraction derived from bovine melanin associated antigen(BMAA) after digestion with the proteolytic enzyme V8 protease was prepared and this soluble fraction of BMAA also induced an experimental autoimmune uveitis (EAU). On gel eletrophoresis for soluble fraction of BMAA, prominent bands between 29 kDa and 43 kDa were clealy observed. In this model, clinical anterior uveitis was induced around 2 weeks, peaked at 18 days and disappeared later than 4 weeks after immunization. Histopathological results of EAU disclosed an infiltration of inflammatory cells, mainly lymphocytes and macrophages, into iris and ciliary body as well as in part the choroid, not retina. In conclusion, we developed a model of EAU with Lewis rats after immunization with BMAA subcutaneously and confirmed the immune mediated inflammation was focused mainly on iris and ciliary body and in part on choroid as well as found that MAA might be soluble after V8 protease treatment.