Histopathological Reactions of Calcium Phosphate Cement - Chitosan Composites

Experimental calcium phosphate cement (CPC)-chitosan composite bone graft materials were recently developed. Although the composite is highly biocompatible, the in vivo resorption properties of the composites have not be been characterized.  Objective: To evaluate resorption characteristics of CPC-chitosan composites in a rat model. Materials: Powder phase of the composite was a CPC consisted of tetracalcium phosphate and dicalcium phosphate anhydrous. The liquid phase consisted of 10 mass% of 100kDa chitosan with degree of deacetylation (DDA) of approximately 70, 80, or 90 dissolved in 1mol/L acetic acid.　Powder to liquid (P/L) ratio of the CPC–chitosan composite was approximately 3.  Methods: CPC-chitosan composites were implanted subcutaneously in rats. Four weeks after operation the animals were sacrificed. Samples for histopathological study were stained with hematoxylin and eosin, and evaluated quantitatively using the NIH image analysis software. Results: All CPC-chitosan composites retained the original cylindrical shape and were encapsulated by thin fibrous connective tissues (FCT) with small amounts of infiltrated cells. The composite containing the DDA=70 chitosan was surrounded by extremely thin FCT, and the chitosan portion was mostly absorbed. The composites containing the DDA=80 or DDA=90 chitosan showed partial chitosan resorption but most of the chitosan area remained.  Quantitative analysis results showed that fraction of chitosan reamined (area of unabsorbed chitosan/original area of implant) for the three groups (mean±s.d.; n=3) were 0.02±0.01 < 0.87±0.06 » 0.95±0.05 (95% confidence level). Conclusion: The CPC-chitosan composites showed excellent biocompatibility and shape-integrity when implanted in subcutaneous tissues.  At 4 weeks post operation, there was a sharp contrast in the extent of resorption between the DDA=70 and the DDA=80 or DDA=90 chitosan. The results suggested that the CPC-chitosan (DDA=70) composite should be expected to resorb in bone grafting applications.  Supported by NIDCR grant DE11789, Nihon University, ADAHF, and NIST.