Parameters such as inflammatory cell infiltration, osteoclast number, alveolar bone and cementum integrity were determined in a single-blind manner and graded, by scores varying from 0 to 3, based on the intensity of findings, as follows: Score 0: absence of or only discrete cellular infiltration, few osteoclasts, preserved alveolar process and cementum; Score 1: moderate cellular infiltration, presence of some osteoclasts, some but minor alveolar process resorption and intact cementum;

Score 2: accentuated cellular infiltration, large number of osteoclasts, accentuated degradation of the alveolar process and partial destruction of cementum; and Score 3: accentuated cellular infiltrate and total destruction of alveolar process and cementum.9 Blood samples were collected from the buy NVP-BKM120 Belnacasan datasheet orbital plexus of anaesthetised

animals (saline and ALD) before the experiment and on the 11th day. The BALP was evaluated using the thermoactivation method, by heating the sample at 56 °C for 10 min,10 since BALP is a thermosensible isoform of total alkaline phosphatase (TALP). BALP serum levels were obtained by the subtraction of heated alkaline phosphatase from TALP serum levels. The methodology used to evaluate the enzymes’ serum levels followed the manufacturers’ directions (Labtest®, Lagoa Santa-MG, Brazil). On the baseline and on the 11th day of the assay, blood samples were collected from the orbital plexuses of anaesthetised animals (saline and ALD). Liver function was evaluated through serum dosage of transaminases: aspartate aminotransferase (AST) and alanine aminotransferase (ALT). TALP serum levels were also evaluated. Specific kits were used, and methodology followed the manufacturer’s instructions (Labtest®, Lagoa Santa-MG, Brazil). The method used to analyse white blood cell counts, as well as its subpopulation (neutrophil and mononuclear cells), was as follows: 20 μl of blood, taken from the rat tail, was added to 380 μl of Turk solution. Total white blood cell counts Isotretinoin were performed using a Neubauer chamber and the differential counts were made using smears stained by

rapid Instant Prov Stain Set (Newprov Produtos para Laboratório; Pinhais-PR, Brazil). A leucogram of the groups of animals (saline and ALD) was performed before periodontitis induction, at the 6th hour and 2nd, 7th and 11th days after the ligature. Animals from saline and ALD groups had their body mass measured before periodontitis induction and after that, daily until the 11th day. Values were expressed as body mass variation (g) compared to the initial body mass. The data are presented as mean ± standard error of the mean (SEM) or median (and range), where appropriate. Analysis of variance (ANOVA), followed by Bonferroni’s test or Student’s t-test, were used to compare means, and Kruskal–Wallis and Dunn tests were used to compare medians. A p < 0.