THE EFFECTS OF BISPHOSPHONATES ON THE RESORPTION CYCLE OF ISOLATED OSTEOCLASTS

Binding sites for wheat germ agglutinin (WGA)lectin have been shown to become revealed in the demineralized resorption lacunae that osteoclasts excavate on bone substrate. Peroxidase-conjugated WGA-lectin, which binds to bone matrix glycoconjugates and proteoglycans, was used in pit formation assays to assess the activity of isolated osteoclasts cultured on either 3-amino-1,1-hydroxypropylidene-bisphosphonate (APD)-or dichloromethylene bisphosphonate (Cl(2)MBP)-covered bone slices. Immunofluorescence and histochemical techniques were also used to study the effects of bone-bound bisphosphonates on isolated rat osteoclasts. Neither APD nor Cl(2)MBP interfered with the special organization of actin or vinculin in osteoclasts when the cells were initializing their resorption cycle. After 24 hours of culture, the number of resorbing osteoclasts increased strongly on control slices, but remained low on either APD- or Cl(2)MBP-treated slices. At this time, the actin and vinculin rings in osteoclasts also started to exhibit abnormal, more diffuse staining. Both bisphosphonates studied resulted in signs of cytotoxicity: the number of osteoclasts decreased on APD- or Cl(2)MBP-covered bone during the course of the study and those remaining attached exhibited severe cytoplasmic retractions. The total areas of resorption remained at significantly lower levels in both experimental groups studied, and this was due to decreases in both the number and sizes of individual resorption pits. The size of the most extensive lacunae detected on the Cl(2)MBP slices did not exceed 5 x 10(3) mu m(2), whereas on the control slices, resorption pits bigger than 15 x 10(3) mu m(2) were frequently discovered.