Tratamento da adenite vesicular (vesiculite) em touros com aplicação de células tronco mesenquimais alogênicas

Abstract

This study aimed to evaluate the effects of applying allogeneic mesenchymal stem cells (MSCs) directly to inflamed vesicular glands in the treatment of vesiculitis. Twelve bulls that presented acute and chronic vesiculitis with two or more recurrences were selected at Semen Collection and Processing Centers (SCPC), based on the presence of a thick yellowish/greenish opaque liquid produced in infected tissue, consisting of dead white blood cells and bacteria with tissue debris and serum. in the ejaculate, pyocytes on the motility and vigor evaluation slide, reagent to the California test Mastitis Test - CMT (one cross and above) and presence of leukocytes on a slide stained by Rapid Panoptico with more than 5 polymorphonuclear cells (PMN) for field. Ultrasound examination of the vesicular glands was performed, however, the clinical signs were definitive for the diagnosis. The proposed method of intraglandular injection of MSCs was application through the ischiorectal fossae with a long needle measuring 30 to 35cm and a guide measuring 25 to 30cm in length directly into the affected vesicular gland. The MSCs were cultured and frozen in the Bio Cell Terapia Celular® laboratory (Brasília, DF, Brazil) and thawed and prepared by washing and centrifugation for intraglandular injection on the day of application. A total of 3x106 MSCs were injected for vesicular gland. The variables analyzed were: ejaculate volume (mL), motility in % and vigor from 1 to 5 in light microscopy, concentration (millions of sperm/mL), number of total sperm in the ejaculate, sperm morphology evaluated in contrast phase with 1000 times magnification. CMT using 0.5 mL of semen plus 2.5 mL of reagent (classification from 0 to 3 crosses), ultrasound of the vesicular glands evaluated on a scale of 1 to 5, with 1 being normal, 2 - with hyperechogenic areas, 3 - with hyperechogenic areas and thickened walls, 4 - with hyperechogenic areas, thickened walls and cysts, 5 - with hyperechogenic areas, thickened walls, cysts and presence of fibrosis and growth of bacterial colonies in sperm culture from frozen semen when possible. All variables were measured before and after treatment. Data were evaluated for normality of residuals using the Shapiro-Wilk test. When the normality of the test was significant (P ˂ 0.05) the data were transformed or outliers were removed and reevaluated. The “T-Test” was applied to identify statistical differences between variables before treatment and after treatment. Furthermore, the binomial test was performed to compare 2 proportions using the chi-square test of the R program. The probability of P ≤ 0.05 was considered a significant difference. Data were presented as mean ± standard error of the mean (S.E.M.). Improvements were observed in initial motility from 60.09 ± 4.8 to 69.89 ± 4.6 (P < 0.05), as well as in post-thaw motility (PD) from 26.26 ± 6.77 to 42. 5 ± 5.99 (P < 0.05). The number of doses produced increased significantly after treatment with MSCs, from 95.61 ± 23.31 units to 337.84 ± 67.75 units (P < 0.05) for ejaculate. The number of piocytes observed for field went from 5.83 ±0.48 to zero, demonstrating the recovery of the inflamed vesicular glands. Based on the results presented, it was concluded that the application of 3x106 MSCs in the vesicular glands of bulls with vesiculitis is safe and efficient, as it improved several parameters evaluated in this research, mainly the production of semen doses for ejaculate.