Figure 1.

(A) Experimental design. (A1) Random Allocation—Forty adult male Wistar rats, (Rattus norvegicus), aged 60 days old, weighing around 250 g were divided into two broad groups: CG—Control group (n = 20)—that received only water with liquid diet and EG—Experimental group (n = 20)—that received ethanol solution 25% (v/v) with liquid diet after adaptation period. (A2) Ethanol Adaptation and Dependence Induced—Animals were gradually drunk at progressive concentrations of ethanol solution (8–16–25% v/v). After 21 days of alcohol adaptation, the animals remained at 25% (v/v) until the surgical procedure for 90 days. (A3) Treatments—After surgical procedures—bilateral bone defect model in the parietals, four subgroups were preformatted according to treatment (blood clot vs. biomaterial) and clinical conditions (alcoholic vs. non-alcoholic): Animals that received only water with liquid diet: CCG (right parietal bone defect was filled with blood clot) and BCG (left parietal bone defect was filled with biomaterial); Animals that received ethanol solution 25% (v/v): CEG (left parietal bone defect was filled with blood clot); BEG (right parietal bone defect was filled with biomaterial). B) Experimental Periods—at 10, 20, 40 and 60 days the skulls of 5 animals/group were collected, totalizing 5 defects/period of each subgroup CCG, BCG, CEG and BEG. (C) Surgical Procedures–Bilateral Bone Defect Model–(C1) 5-mm left parietal osteotomy; (C2) Two bone defects in parietal bones; (C3) One defect filled with biomaterial and the contralateral only with blood clot; (C4) Periosteum suture with nylon 5-0.