J. Bennett

The differentiation of adipocytic and osteogenic cells has been investigated in cultures of adult rat marrow stromal cells. Adipocytic differentiation was assessed using morphological criteria, changes in expression of procollagen mRNAs, consistent with a switch from the synthesis of predominantly fibrillar (types I and III) to basement membrane (type IV) collagen, and the induction of expression of aP2, a specific marker for differentiation of adipocytes. Osteogenic differentiation was assessed on the basis of changes in the abundance of the mRNAs for the bone/liver/kidney isozyme of alkaline phosphatase and the induction of mRNAs for bone sialoprotein and osteocalcin. In the presence of foetal calf serum and dexamethasone (10(-8) M) there was always differentiation of both adipocytic and osteogenic cells. When the steroid was present throughout primary and secondary culture the differentiation of osteogenic cells predominated. Conversely, when dexamethasone was present in secondary culture only, the differentiation of adipocytes predominated. When marrow stromal cells were cultured in the presence of dexamethasone in primary culture and dexamethasone and 1,25-dihydroxyvitamin D3 (1,25(OH)2D3; 10(-8) M) in secondary culture, the differentiation of adipocytes was inhibited whereas the differentiation of osteogenic cells was enhanced, as assessed by an increase in expression of osteocalcin mRNA. The results, therefore, demonstrate an inverse relationship between the differentiation of adipocytic and osteogenic cells in this culture system and are consistent with the possibility that the regulation of adipogenesis and osteogenesis can occur at the level of a common precursor in vivo.

Stromal colonies with fibroblastic morphology grown from rabbit marrow cells in culture supplemented with foetal calf serum. In this study the same marrow cells cultured with autologous rabbit plasma and hydrocortisone form colonies of a single lineage that express the adipocytic phenotype. A comparison of the potential for differentiation of cloned cell populations grown from fibroblastic and adipocytic colonies has been made using an in vivo diffusion chamber assay. The adipocytic colonies differentiated and grew to a limited size in medium with rabbit plasma and hydrocortisone, but attempts to isolate them and expand them in this medium failed. When the serum supplement was changed to foetal calf serum at day 10 the cells in the adipocytic colonies acquired a less differentiated morphology, there was a large increase in colony growth and cells were produced in sufficient numbers for the diffusion chamber assay. Thirty one fibroblastic colonies and twenty one adipocytic colonies were isolated either by limiting dilution or ring cloning and then expanded. Of these, eleven fibroblastic and eight adipocytic colonies provided enough cells (2 x 10(5) to 2 x 10(6] for implantation and culture in the chambers. Four of the eleven fibroblastic and three of the eight adipocytic colonies formed an osteogenic tissue in the chambers. It was concluded that cells that have differentiated in an adipocytic direction are able to revert to a more proliferative stage and subsequently to differentiate along the osteogenic pathway. Adipocytic and fibroblastic cells cultured in vitro from marrow have, with osteogenic cells, a common precursor in adult marrow.

Neonatal pig bone marrow stromal cells (PBMSC) were tested in vivo and in vitro to establish their use as a large-animal model for the study of skeletogenesis. When implanted in diffusion chambers in athymic mice for 6-8 weeks, both freshly isolated pig bone marrow and passage 2 PBMSC formed partially mineralized cartilage, bone-like material, and fibrous tissue. The cartilage showed metachromatic, perilacunar staining with toluidine blue and safronin O, alcian blue staining for chondroitin and keratan sulfate, and intense immunostaining for type II collagen. Osteocalcin was immunolocalized to the mineralized regions, consistent with the formation of bone. Alkaline phosphatase was primarily observed in cell layers at boundaries between tissue types. Unstimulated monolayer cultures of PBMSC produced type I but not type II collagen, responded to dexamethasone (10(-8) M) with a 1.7-fold increase in alkaline phosphatase activity, and were stimulated to divide by basic fibroblast growth factor (1.5-fold; EC50 1 ng/ml). Transforming growth factor beta (TGF-beta) blocked both dexamethasone-induced alkaline phosphatase expression (EC50, 1 ng/ml of TGF-beta) and the mitogenic effects of bFGF (EC50 0.06 ng/ml of TGF-beta). When incubated for 10-14 days in medium containing dexamethasone, beta-glycerophosphate and ascorbate PBMSC formed mineralized nodules. Calcification occurred in the middle of the aggregates and was associated with intensely alkaline phosphatase positive cells and a dense type I collagen-rich matrix. PBMSC also displayed colony-forming unit-fibroblastic activity, with approximately 1 in 80 of the plated cells formed colonies > 128 cells over 14-21 days. PBMSC therefore mimic the known activities of stromal cells from other species, including the human, suggesting that they are a valid model for skeletal research.

The nasopalatine cyst (NPC) was first described in 1914 and it is considered the most common non-odontogenic cyst. Most studies show a higher incidence of NPC among males than females, with a male/female ratio of 1.7:1. The majority of the cases described in people in their fifth decade involve Afro-Caribbeans, while those in their sixth decade are mainly caucasians. These cysts are normally asymptomatic, unless they are infected. The most commonly reported clinical symptom is swelling in the anterior part of the palate. The treatment of choice is enucleation. Even though it has been stated that NPCs may occur at any age no reports have been made on children less than 9 years old. Some reports support a predisposition in young Afro-Caribbeans, where NPCs appear to be more aggressive, larger and symptomatic. We present a case of a NPC in an 8-year-old caucasian female.