Bone morphogenetic proteins (BMPs) promote the differentiation of osteoprogenitor cells, and also induce osteogenesis in bone marrow stromal cells (MSC) from rats and mice. However, compared to results with animal models, BMPs are relatively inefficient in inducing human MSC to undergo osteogenesis, and are much less effective in promoting bone formation in human clinical trials. Previous studies indicated that, while human MSC respond to dexamethasone with elevated levels of the osteoblast marker alkaline phosphatase, most isolates of human MSC fail to show alkaline phosphatase induction in response to BMP-2, BMP-4, or BMP-7. Several other genes known to be induced by BMPs are appropriately regulated; thus, human MSC are capable of some BMP-activated signaling. Analysis of the BMP receptors ALK-3 and ALK-6 indicated that, although ALK-6 mRNA was not expressed in human MSC, overexpressing a constitutively active ALK-6 receptor did not induce elevated alkaline phosphatase. Real-time RT-PCR was used to investigate expression of several osteoblast-related transcription factors in MSC after 6 days’ exposure to BMP2 or dexamethasone. Msx-2, a transcription factor that has been reported to inhibit differentiation of osteoprogenitor cells, showed 10-fold elevation in BMP-2-treated human MSC, but not in BMP-2-treated rat MSC. Overexpression of Msx-2 in human and rat MSC, however, did not alter alkaline phosphatase levels, which suggests that absence of BMP-stimulated alkaline phosphatase was not caused by the BMP-2-induced increase in Msx-2. Although Runx2 isoforms have been implicated in control of osteoblast differentiation, levels of this transcription factor were unaffected by BMP treatment. Expression of the FKHR transcription factor, which has been reported to regulate alkaline phosphatase transcription in mouse cells, showed a modest increase in response to BMP-2, but a much greater increase in dexamethasone-treated cells. We propose that BMP regulation of the bone/liver/kidney alkaline phosphatase gene is indirect, requiring expression of new transcription factor(s) that behave differently in rodent and human MSC.

In this large series of endoscopically resected IPs with extensive clinical follow-up, recurrences occurred an average of 23 months after the procedure. This emphasizes the importance of long-term endoscopic follow-up to detect recurrences in all patients. Endoscopic or endoscopic-assisted resection of IPs is a valid technique in this series with recurrence rates comparable with open approaches.

Air-liquid interface models using murine tracheal respiratory epithelium have revolutionized the in vitro study of pulmonary diseases. This model is often impractical because of the small number of respiratory epithelial cells that can be isolated from the mouse trachea. We describe a simple technique to harvest the murine nasal septum and grow the epithelial cells in an air-liquid interface. The degree of ciliation of mouse trachea, nasal septum, and their respective cultured epithelium at an air-liquid interface were compared by scanning electron microscopy (SEM). Immunocytochemistry for type IV beta-tubulin and zona occludens-1 (Zo-1) are performed to determine differentiation and confluence, respectively. To rule out contamination with olfactory epithelium (OE), immunocytochemistry for olfactory marker protein (OMP) was performed. Transepithelial resistance and potential measurements were determined using a modified vertical Ussing chamber SEM reveals approximately 90% ciliated respiratory epithelium in the nasal septum as compared with 35% in the mouse trachea. The septal air-liquid interface culture demonstrates comparable ciliated respiratory epithelium to the nasal septum. Immunocytochemistry demonstrates an intact monolayer and diffuse differentiated ciliated epithelium. These cultures exhibit a transepithelial resistance and potential confirming a confluent monolayer with electrically active airway epitheliumn containing both a sodium-absorptive pathway and a chloride-secretory pathway. To increase the yield of respiratory epithelial cells harvested from mice, we have found the nasal septum is a superior source when compared with the trachea. The nasal septum increases the yield of respiratory epithelial cells up to 8-fold.

Bioactive glass (BG) is an effective synthetic bone graft material. BG granules of narrow size range (300-355 mum) have the ability to form new bone tissue inside excavations produced by in vivo resorption. Previously, we demonstrated that BG stimulates the differentiation of cultured osteoblast precursors if the glass surface was biomimetically modified by the formation of bone-like apatite and adsorption of serum proteins. We now report that modified BG can also increase the rate at which multipotential rat bone marrow stromal cells (rMSC) will undergo osteogenesis. BG promoted rMSC osteogenesis both when cells were plated in contact with BG and when cells were not directly in contact with the BG. Alkaline phosphatase activity, a marker of bone cell differentiation, was used as an indicator for osteogenesis. Alkaline phosphatase activity of rMSCs exposed to osteoinducers such as ascorbate, dexamethasone, and BMP-2 was enhanced in the presence of BG. The stimulatory effect of BG was more pronounced in rMSC cultures with low basal alkaline phosphatase activity than in those with higher activity. The enhanced differentiation of rMSCs was associated with both a change in rMSC morphology and altered chemical composition of the cell culture media. rMSCs cultured on BG in the presence of BMP or dexamethasone exhibited a more rounded osteoblast-like appearance as compared with cells grown on tissue culture plastic. In the presence of BG, elevated levels of calcium and silicon in the culture medium were observed throughout the 7-day culture period, suggesting a continuous dissolution of surface-modified BG and resulting release of BG dissolution products. The data suggest that both surface- and solution-mediated events play a role in the osteogenic effect of BG.

To increase the yield of respiratory epithelial cells harvested from mice, we have found that the nasal septum is a far superior source when compared with the trachea. The greater surface area and increased concentration of ciliated epithelial cells has the potential to provide an eightfold increase in epithelial cells for the development of air-liquid interface cultures.