Banca de DEFESA: GUSTAVO MARINHO MIRANDA

Uma banca de DEFESA de MESTRADO foi cadastrada pelo programa.
STUDENT : GUSTAVO MARINHO MIRANDA
DATE: 16/12/2021
TIME: 09:00
LOCAL: IGM Sala Virtual 3
TITLE:

CHARACTERIZATION OF THE BIOCHEMICAL, HEMATOLOGICAL AND INFLAMMATORY PROFILE IN SICKLE MICE: 

CORRELATIONS WITH LIPID METABOLISM AND EICOSANOID PRODUCTION IN A SKIN ULCER MODEL

KEY WORDS:

Eicosanoids; Inflammation; Mouse transgenic; Sickle cell disease; Skin ulcer.

PAGES: 81
BIG AREA: Ciências Biológicas
AREA: Genética
SUMMARY:

ABSTRACT

INTRODUCTION: Sickle Cell Anemia (SCA) is an autosomal recessive hemoglobinopathy characterized by the mutation in the sixth position of the β-globin gene (GAG àGTG) that leads to the replacement of glutamic acid by valine, generating hemoglobin S (HbS). Its pathogenesis involves HbS polymerization, vaso-occlusion and sterile inflammation with subsequent complications and systemic changes. Sickle cell ulcers are complications that affect 8% to 10% of SCA patients. While it is well-established that sickle cell ulcer both results from genetic predisposition and induces inflammatory processes, the mechanisms involved, in particular, the participation of lipid mediators, remain to be better investigated.  OBJECTIVE: To characterize the hematological, biochemical, and inflammatory profile of transgenic sickle cell anemia mice and to evaluate the local and systemic alterations induced by cutaneous ulcer in these animals. MATERIALS AND METHODS:  After anesthesia, samples of whole blood, serum, organs and skin fragments of Townes B6:129 mice were collected in order to determine the baseline differences in biochemical, hematological and inflammatory parameters of healthy (AA), sickle cell trait (AS) and sickle cell anemia (SS) mice. In the murine model of sickle cell ulcer, four 4 mm skin lesions were induced on the back of the animals, and samples of these lesions were collected at the time-points 0, 24 and 72 h after ulcer induction. In these same evaluation periods, whole blood samples were separated for blood counts and leukogram analysis by semi-automated analysis, using URIT 5160 Vet equipment and blood smears with the Romanowsky method. Biochemical analyses of total cholesterol (TC), triglycerides, AST, ALT, urea and creatinine were performed in the serum by spectrophotometry by automated method in a smart vet 200+ equipment. The concentrations of PGE₂ and LTB₄ in the serum were evaluated by ELISA. The expression of genes associated with the production of eicosanoids such as COX-1, COX-2, 5-LO, PLA₂ and PLIN₂ in skin lesions was determined by RT-qPCR.  Histological analyses of inflammatory infiltrates in the skin were performed by optical microscopy using H&E-stained slides and quantifications were performed using the Imaje J program version 1.53. The differences between the groups and their respective time-points were evaluated by One-way ANOVA, Mann-Whitney and Kruskal-Wallis tests, using the Graphpad Prism program version 8.0. RESULTS: The characterization of baseline parameters showed that SS animals presented severe hemolytic anemia and leukocytosis, while the AS group showed no significant differences compared to the control group. High levels of AST were observed in AS and SS animals, compared to the AA group, while ALT was found to be increased in the SS group compared to both AA and AS animals, and creatinine levels were increased in the SS group compared to the AA group. In the SS group, the levels of PGE₂ in the serum were increased while LTB₄ levels were decreasedin comparison with AA and AS mice. Negative correlations between hemoglobin, red blood cells, hematocrit and PGE₂ were observed. The changes induced by the ulcer protocol in the blood counts of the SS group were not significant. However, we observed a reduction in total leukocytes within 24 h and an increase in 72 h in this group. On the other hand, the tissue leukocyte infiltrate increased in 24 h and a slight reduction in 72 h. In the SS group in 24 h, there was a reduction in total cholesterol (TC) and AST. In 72 h, we observed an increase in TC and a reduction of ALT and creatinine in the same group. Also in the SS group, tissue COX-2 expression was increased in 72 h, while systemic PGE₂ and LTB₄ did not present significant alterations. CONCLUSION: Transgenic sickle cell anemia mice present biochemical and hematological alterations that are correlated with an imbalance in lipid metabolism and eicosanoid production, contributing to a greater inflammatory state at the basal level. The induction of skin ulcers in this model leads to greater leukocyte recruitment in sick animals compared to healthy and sickle cell trait animals, which seems to be related to the biochemical, hematological, and inflammatory changes presented by these animals.

BANKING MEMBERS:
Externa à Instituição - PATRÍCIA TORRES BOZZA VIOLA - Fiocruz - RJ
Presidente - 005.343.193-60 - JAIME RIBEIRO FILHO - Fiocruz - RJ
Interna - 011.952.725-10 - NATALIA MACHADO TAVARES - UFBA