IJCS | Volume 32, Nº2, May/June 2019

248 Negrão et al. Effect of herbicide 2,4D inhalation on cardiac remodeling Int J Cardiovasc Sci. 2019;32(3)247-252 Original Article by 93%, with Brazil being the leader in consumption worldwide, a position previously occupied by the United States. 7 The Ministry of Health considers that there are more than 100,000 chemical substances capable of causing chemical, physical, morphological and pathophysiological damage to the health of populations. 8-12 Among those who suffer the most from the exaggerated use of agrochemicals are rural workers, when these substances are utilized incorrectly. 6,9 In 1990, the World Health Organization (WHO) estimated that approximately 3,000,000 cases of pesticide poisoning occurred per year in the world, with 220,000 deaths per year. 13 Only in 2009, more than 1 billion liters of agrochemicals were utilized in Brazil; it is the same as if each one of the Brazilian individuals consumed an average of five liters of poison per year. 14 In Brazil, pesticide poisoning is a public health problem and a matter of compulsory notification. 15 The toxicity can be fast or slow and it can take from months to years for symptoms to appear in the worker exposed to the pesticide. Among the harmful effects are allergic reactions, skin and eye irritation, neurological, respiratory, hematopoietic, hepatic, nephrological, gastrointestinal, pancreatic, and cardiovascular alterations, as well as several toxic reactions. 9-12 The effects of several types of pesticides have already been well established, resulting in cardiotoxicity with mitochondrial alterations, oxidative stress, hypotrophy, and molecular alterations of gene expression related to cardiac calcium. 16-19 On e o f t h e h e r b i c i d e s f r e q u e n t l y u s e d to fight weeds in sugarcane plantations is 2,4D (2,4-dichlorophenoxyacetic acid), 6 which has harmful effects when ingested, absorbed by the skin and/or inhaled. This herbicide can cause peripheral neuritis and transient diabetes during exposure to it, as well as loss of appetite, skin irritation, nausea, gastrointestinal tract irritation, vomiting, chest and abdominal pain, exhaustion, weakness, muscle twitching, mental confusion, seizures and even coma. If 2,4 D is used at high concentrations, it may cause degenerative liver and kidney lesions. 20 However, to the best of our knowledge, there have been no studies evaluating the cardiotoxic effects of 2,4 D using a nebulizing spray containing a sodium chloride (NaCl) solution during a 72h-period. The aim of this study was to analyze the cardiac morphological effects on the left ventricle (LV) of mice submitted to daily nebulizing sprays containing the 2,4D herbicide for 15 minutes. It is hypothesized that exposure to high concentrations of 2,4D herbicide will lead to cardiac remodeling. Methods Ethical aspects This was an experimental cross-sectional study submitted to theAnimal Ethics Committee of Universidade do Oeste Paulista and evaluated by the Animal Use Ethics Commission (CEUA, from the Portuguese Comissão De Ética do Uso De Animais ), and approved under protocol n. 3331 to be performed according to the Guide for the Care and Use of Laboratory Animals of the Institutes of Health (USA). Sample characterization For the experiment, 15 male Swiss mice (30-40 g) were provided by the Animal Center of Universidade do Oeste Paulista , where they were housed in collective plastic cages (5 animals per cage) measuring 30x16x19 centimeters, under a mean temperature of 22 ± 2ºC, with 12-hour cycles of light and dark, i.e., light from 7AM to 7PM and dark from 7PM to 7AM. To perform the experiment, the 15 animals were randomly divided into three groups, using the simple randomization method (random sequence performed through Excel): - CG: control group (n = 5) exposed to nebulization with sodium chloride solution (NaCl) for 15 minutes/ day, for 3 consecutive days. - LCG: low concentration group (n = 5) exposed to nebulization of 2,4D herbicide with 3.71 x 10-3 grams of active ingredient per hectare (g.i.a./ha) for 15 minutes/ day, for 3 consecutive days. - HCG: high concentration group (n = 5) exposed to nebulization of 2,4D herbicide with 9.28 x 10-3 grams of active ingredient per hectare (g.i.a/ha) for 15 minutes/ day, for 3 consecutive days. 2.4D Herbicide Exposure Protocol The experimental protocol used two boxes of 2.4D (32x24x32 cm), each attached to a Pulmosonic Star ®9 ultrasonic nebulizer, and the following concentrations were administered after being diluted in 10 mL of 0.9% sodium chloride: - CG: 10 mL of 0.9% sodium chloride; - LCG: Solution consisting of 3.71 x 10-3 grams of active