Immunolocalization of stress proteins and extracellular matrix proteins inthe rat tibia

Stress proteins (heat shock proteins [hsps]) serve a number of protective f unctions, including protection from apoptosis and acting as chaperones duri ng protein biosynthesis. For example, hsp 27 has been defined as a chaperon e for the G3 domain of aggrecan, while hsp 47 is the chaperone for type I c ollagen. Separate cytoprotective roles for hsp 27 and hsp 70 have been demo nstrated. The aim of this study was to define the expression of hsps in ost eoblastic and chondrocytic cells of the growing rat long bone in relationsh ip to the immunohistochemical localization of aggrecan, type I collagen and the presence of fragmented DNA that defines apoptotic events, Tibiae were harvested from Fisher 344 rats (n=6) and fixed in 10% buffered formalin. Sa mples were decalcified in 10% EDTA, bisected, and processed for histologic examination. Sections (5 mm) were immunohistochemically stained using a str eptavidin-biotin detection method, Go-localization of hsps with apoptosis w as achieved using the TUNEL procedure, In the rat tibia growth plate, aggre can was generally distributed throughout cartilage and chondrocytes. Howeve r, hsp 27 expression was observed only in the lower hypertrophic chondrocyt es, hsp27 was present in osteoblasts lining newly formed bone, hsp 47 stain ing was also prominent within these osteoblasts where collagen type I immun olocalization occurred. The inducible form of hsp 70 was localized to the o steoblastic cells lining new bone in the primary spongiosa. In cartilage, D NA fragmentation was restricted to the hypertrophic, hsp27-positive, chondr ocytes. In contrast, DNA fragmentation was not co-localized with hsp27-posi tive osteoblastic cells of the primary spongiosa, although occasional apopt otic cells were identified. These results indicate that apoptosis is a mech anism by which hypertrophic chondrocytes are eliminated from cartilage prio r to calcification, but that other mechanisms are also likely to be involve d. They also suggest that hsps have cytoprotective and biosynthetic functio ns within osteoblasts and chondrocytes, but apoptotic signals may override these effects in some instances, resulting in apoptosis, (C) 2000 Harcourt Publishers Ltd.