Akira Iwaki

Localization and quantification of proliferating cells during rat fracture repair: detection of proliferating cell nuclear antigen by immunohistochemistry.

Bilateral femurs of 12‐week‐old female Sprague‐Dawley rats were fractured, and the fractured femurs were harvested 36 h, 3, 7, 10, and 14 days after the fracture. Localization of cell proliferation in the fracture calluses was investigated using immunohistochemistry with antiproliferating cell nuclear antigen (PCNA) monoclonal antibodies. Thirty‐six hours after the fracture, many PCNA‐positive cells were observed in the whole callus. The change was not limited to mesenchymal cells at the fracture site where the inflammatory reaction had occurred, but extended in the periosteum along almost the entire femoral diaphysis where intramembranous ossification was initiated. On day 3, periosteal cells or premature osteoblasts in the newly formed trabecular bone during intramembranous ossification still displayed intense staining. On day 7, many premature chondrocytes and proliferating chondrocytes were PCNA positive. Endochondral ossification appeared on days 10 and 14, and the premature osteoblasts and endothelial cells in the endochondral ossification front were stained with anti‐PCNA antibodies. Quantification of PCNA‐positive cells was carried out using an image analysis computer system, obtaining a PCNA score for each cellular event. The highest score was observed in the periosteum early after the fracture near the fracture site. Immunohistochemistry using anti‐PCNA antibodies showed that the distribution of proliferating cells and the degree of cell proliferation varied according to the time lag after the fracture, suggesting the existence of local regulatory factors such as growth factors, and that significant cell proliferation was observed at the beginning of each cellular event.

Serum 1α,25-Dihydroxyvitamin D3 Accumulates into the Fracture Callus during Rat Femoral Fracture Healing

1,25-dihydroxyvitamin D3 (1,25(OH)2D3) is thought to be an important systemic factor in the fracture repair process, but the mechanism of action of 1,25(OH)2D3 has not been clearly defined. In this study, the role of 1,25(OH)2D3 in the fracture repair process was analyzed in a rat closed femoral fracture model. The plasma concentration of 1,25(OH)2D3 rapidly decreased on day 3 and continued to decrease to 10 days after fracture. We assessed whether this decrease was based on the accelerated degradation or retardation of the synthesis rate of 1,25(OH)2D3, from 25(OH)D3. After radiolabeled 3H-1,25(OH)2D3 or 3H-25(OH)D3 was injected i.v. into fractured or control (unfractured) rats, the concentrations of 25(OH)D3 and 1,25(OH)2D3 metabolites were measured by HPLC. The plasma concentrations of these radiolabeled metabolites in fractured group were similar to those in control rats early after operation. However, radioactivity in the femurs of fractured rats was higher than that of the control group. Furthermore, the radioactivity was concentrated in the callus of the fractured group analyzed by autoradiography. 1,25(OH)2D3 receptor gene expression was detected early after fracture and, additionally, both in the soft and hard callus on days 7 and 13 after fracture. These results showed that the rapid disappearance of 1,25(OH)2D3 in the early stages after fracture was not due to either increased degradation or decreased synthesis of 1,25(OH)2D3, but rather to increased consumption. Further, these results suggest the possibility that plasma 1,25(OH)2D3 becomes localized in the callus and may regulate cellular events in the process of fracture healing.

The Evaluation of Anticonvulsant Induced Bone Atrophic Changes in Children -Densitometric Analysis

For the purpose of mass screening of metalolic bone diseases, we evaluated long-term anticonvulsant induced bone atrophic changes.The diameter and osteodensity of the second metacarpal bone in hand films of 46 children in-patients of Beppu Seisien were measured by microdensitometry (MD method).Laboratory findings including gerum Ca, P, ALP were compared with the severity of bone atrophy.The following results were obtained. 16 cases (35%) were normal, 19 cases (41%) were in the initial stage of abnormality, 11 cases (24%) in grade I, no case with grade II or III of abnormality.The severity of bone atrophy correlated with serum Ca and ALP but the duration of anticonvulsant administration did not correlate with the severity of bone atrophy.Sixteen children with noted atrophy were treated with “1α-OH-D3” and after 15 months their bone atrophy was remeasured by the MD method. In almost all cases the bone atrophy was well improved showing the “1α-OH-D3” to be very effective.Baned on there results, it is suggested that roentgenologic and biochemical supervision of patients is required during long-term anticonvulsant therapy.

Clinical Evaluation of the Results of Posterior Decompression Surgery for OPLL in Cervical Spine

In the past, as a posterior decompression surgery for OPLL, Iaminectomy was perfomed. Howerver, postoperative follow-up was made difficult by such negative factors as instability of the operated part, abnormal alignment, scar formation, etc. Therefore, to counteract these negative results, a variety of Iaminoplasty are being tried in recent years. We have performed Iaminoplasty both by the Kirita and Hirabayashi methods at our hospital since 1983, and we hereby report our evaluation of the operative results and the postoperative complications.With the Hirabayashi method, we observed the limitation of the range of movement in the posterior flexion. We could observe no difference in the improvement rate of the clinical condition when using different operative techniques. Postoperatively, we had no case where OPLLs growth could be clearly observed.In case of the unilateral existence of ossifications with a high rate of stenosis, we thought it was necessary to practice sufficient caution at the time of operation in order to prevent complications.