Osteocyte Apoptosis, Bone Marrow Adiposity, and Fibrosis in the Irradiated Human Mandible

Abstract

Purpose: To assess the effect of radiation therapy on osteocyte apoptosis, osteocyte death, and bone marrow adipocytes in the human mandible and its contribution to the pathophysiology of radiation damage to the mandibular bone.

Methods and materials: Mandibular cancellous bone biopsies were taken from irradiated patients and nonirradiated controls. Immunohistochemical detection of cleaved caspase-3 was performed to visualize apoptotic osteocytes. The number of apoptotic osteocytes per bone area and per total amount of osteocytes, osteocytes per bone area, and empty lacunae per bone area were counted manually. The percentage fibrotic tissue and adipose tissue per bone marrow area, the percentage bone marrow of total area, and the mean adipocyte diameter (μm) was determined digitally from adjacent Goldner stained sections.

Results: Biopsies of 15 irradiated patients (12 men and 3 women) and 7 nonirradiated controls (5 men and 2 women) were assessed. In the study group a significant increase was seen in the number of empty lacunae, the percentage of adipose tissue of bone marrow area, and the adipocyte diameter. There was no significant difference in bone marrow fibrosis nor apoptotic osteocytes between the irradiated group and the controls.

Conclusions: Irradiation alone does not seem to induce excessive bone marrow fibrosis. The damage to bone mesenchymal stem cells leads to increased marrow adipogenesis and decreased osteoblastogenic potential. Early osteocyte death resulting in avital persisting bone matrix with severely impaired regenerative potential may contribute to the vulnerability of irradiated bone to infection and necrosis.

Figure 1

Histologic sections (× 200 magnification) of irradiated mandibular bone from an irradiated patient (D_max = 34 Gy). (A) Goldner trichrome stain. Osteocyte nuclei are stained dark purple. Arrowheads point toward empty lacunae, indicating osteocyte death. (B) Cleaved caspase-3 stain. Arrowheads point toward cleaved caspase-3 positive osteocytes, indicating osteocyte apoptosis.

Figure 2

(A) Number of empty lacunae per bone area (N.e.Lc/B.Ar) in the control and irradiated groups (Mann-Whitney U test; P = .007). (B) Percentage of adipose tissue of bone marrow area (Ad.T.Ar/Ma.Ar) in the control and irradiated groups (Mann-Whitney U test; P = .007). (C) Adipocyte diameter in the control and irradiated groups (Mann-Whitney U test; P = .005).

Figure 3

Histologic sections with Goldner trichrome stain of mandibular bone from an unirradiated control patient (A: × 40 magnification, B: × 100 magnification) and from an irradiated patient, D_max = 53 Gy (C: × 40 magnification, D: × 100 magnification). In both specimens, fibrotic areas as well as adipose tissue is present. The nonirradiated bone marrow has more abundant nuclei and the fibrosis is more localized. The irradiated bone marrow is hypocellular with smaller fibrotic patches scattered throughout the marrow space.