Biotherapic of Trypanosoma cruzi 17x controlled histopathological alterations in mice infected by this protozoon
Authors: Sandri Patrícia Flora , Falkowski Gislaine Janaina Sanchez , JÃƒÆ’Ã‚Âºnior AnÃƒÆ’Ã‚Â©lio Dias Nascimento , Spack Miguel , Moreira Neide Martins , Toledo Max Jean de Ornelas , Filho Benício Abreu , Gabriel Maristela , AraÃƒÆ’Ã‚Âºjo Silvana Marque de 
Published: December 2021

Introduction: about 10 million people worldwide suffer from ChagasÃƒÂ¢Ã¢â€šÂ¬Ã¢â€žÂ¢ disease [1]. The World Health Organization (WHO) has explicitly acknowledged the significance of this condition and supports the use of Complementary and Alternative Medicine by health systems integrated with conventional treatments. Even so, one century after its discovery it still represents a global challenge [1,2]. Biotherapics are ultradiluted medicines and the infection of mice by Trypanosoma cruzi is an excellent model to understand their effect [3,4]. At 8 weeks, mice are physiologically more developed than at age 4 weeks, including a more competent immune system [5].

Aim: the aim of this study was to assess the effect of biotherapic of T. cruzi in dilution 17x on liver and spleen tissue of mice of different ages infected by this protozoon.

Method: in a blind, randomized controlled trial 12 male Swiss mice aged 4 and 8 weeks, infected by 1,400 blood trypomastigotes T. cruzi Y strain were divided into groups control ÃƒÂ¢Ã¢â€šÂ¬Ã¢â‚¬Å“ treated with 7% hydroalcoholic solution (CI-4=3 animals or CI-8=3 animals) and treated with biotherapic 17x (BIOT-4=3 animals or BIOT-8=8 animals). Treatment (0.2 ml biotherapic/day/animal, per gavage) started after infection was verified (4th day) and animals were sacrificed on the 3rd day of treatment. For histopathological exam, the liver and spleen were removed and fixated in 4% paraformaldehyde for 24 hours and then processed for inclusion in paraffin. Semi-serial 7mm cuts were made and subjected to hematoxylin-eosin stain. It was performed a quantitative analysis of the number of nests of amastigotes and inflammatory foci in the liver. Slides were observed under microscope Olympus BX41 (Tokyo, Japan) and images captured with camera Qcolor3 (Olympus) coupled to the microscope. In the spleen it was counted the number of nests of amastigotes and the number of foreign-body giant cells. In each organ, 20 microscopic fields/cut were counted under power 40x totaling 120 fields/animal with microscope Olympus CBA (Tokyo, Japan). To analyze data it was used software Statistica 8.0. For data not exhibiting normal distribution it was used Kruskal-WallisÃƒÂ¢Ã¢â€šÂ¬Ã¢â€žÂ¢ test at 5% significance and ANOVA for the ones with normal distribution. Chi-square test was used to compare percentages. Biotherapic 17d was prepared by adding 0.9 ml of blood with T. cruzi (10E+7 trypomastigotes/ml) to 9.1 ml of distilled water in laminar flow. Following dilutions were prepared in 86% hydroalcoholic solutions up to 16x. Dilution 17x was prepared with 7% hydroalcoholic solution [6]. It was performed microbiological control and in vivo biological risk of the biotherapic. Results showed a number of colony forming units proper for use (>1CFU/ml). Intraperitoenal inoculation of the biotherapic did not cause infection in animals. This study was approved by the Ethics Committee for Animal Experimentation/UEM protocol 030/2008.

Results and Discussion: in the liver, animals of group BIOT-8 exhibited less nests of amastigotes (p