MODULASI EKSPRESI CD68, INOS, DAN CD163 SERTA BIOKOMPATIBILITAS SISTEMIK OLEH BIOMATERIAL HIBRIDA TRIKALSIUM SILIKAT DAN KALSIUM KARBONAT BIOGENIK (ACHATINA FULICA) PADA PULPA GIGI TIKUS TERINFLAMASI: STUDI IN VIVO

Abstract Objectives: To evaluate the in vivo immunomodulatory efficacy and systemic biocompatibility of a hybrid biomaterial comprising 75% tricalcium silicate (TCS) and 25% biogenic calcium carbonate (CaCO₃) derived from Achatina fulica shells in inflamed dental pulp. Materials and Methods: This experimental in vivo study utilized 48 male Wistar rats. Pulp inflammation was mechanically induced prior to biomaterial application. Subjects were randomly allocated into three main groups (n=16/group): a negative control (saline), a positive control (100% TCS), or an experimental group (75% TCS + 25% biogenic CaCO₃). Euthanasia was subsequently performed in stages on days 1, 3, 7, and 14 (n= 4/timepoint subgroup). Biomaterial characterization utilized Fourier-transform infrared spectroscopy (FTIR) and scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM-EDX). Tissue responses were evaluated via immunohistochemical H-scores for CD68 (pan-macrophage), iNOS (M1 phenotype), and CD163 (M2 phenotype). Local pulp biocompatibility was further evaluated using adjunctive parameters of systemic enzyme safety observation (hepatorenal profiles). Results: FTIR confirmed the high purity of biogenic CaCO3, and the hybrid biomaterial demonstrated stable calcium retention (23.7%) while effectively buffering elemental dilution. In vivo, the hybrid composition significantly suppressed pan-macrophage infiltration and early M1 pro-inflammatory polarization (iNOS) by day 3, reaching basal levels by day 14 (p<0.005). Conversely, it induced a notable upregulation of the M2 reparative marker (CD163), peaking at day 14 (p<0.005). Furthermore, the hybrid formulation proved to be safe systemically, successfully maintaining hepatorenal parameters (transaminases, BUN, and creatinine) at normal physiological baselines, in contrast with the transient elevations observed with pure TCS. Conclusions: The integration of biogenic calcium carbonate into a tricalcium silicate matrix accelerates the phenotypic transition of macrophages from a pro-inflammatory M1 to a pro-resolving M2 profile. This hybrid material mitigates initial alkalinity-induced stress, aids in the resolution of acute pulpitis, and exhibits a favorable safety profile by preserving vital organ homeostasis, presenting a translatable approach for vital pulp therapy.