Bone quality assessment of osteogenic cell cultures by Raman microscopy

The use of autologous stem/progenitor cells represents a promising approach to the repair of craniofacial bone defects. The calvarium is recognized as a viable source of stem/progenitor cells that can be transplanted in vitro to form bone. However, it is unclear if bone formed in cell culture is similar in quality to that found in native bone. In this study, the quality of bone mineral formed in osteogenic cell cultures were compared against calvarial bone from postnatal mice. Given the spectroscopic resemblance that exists between cell and collagen spectra, the feasibility of extracting information on cell activity and bone matrix quality were also examined. Stem/progenitor cells isolated from fetal mouse calvaria were cultured onto fused‐quartz slides under osteogenic differentiation conditions for 28 days. At specific time intervals, slides were removed and analyzed by Raman microscopy and mineral staining techniques. We show that bone formed in culture at Day 28 resembled calvarial bone from 1‐day‐old postnatal mice with comparable mineralization, mineral crystallinity, and collagen crosslinks ratios. In contrast, bone formed at Day 28 contained a lower degree of ordered collagen fibrils compared with 1‐day‐old postnatal bone. Taken together, bone formed in osteogenic cell culture exhibited progressive matrix maturation and mineralization but could not fully replicate the high degree of collagen fibril order found in native bone.In this Raman spectroscopic study, we examined the quality of bone formed in vitro by fetal mouse calvarial stem/progenitor cells under osteogenic differentiation conditions. We characterized bone mineral and matrix cell culture components and detected the presence of lipid and glycosaminoglycan‐like components. Bone formed in vitro at Day 28 was similar to 1‐day‐old postnatal mouse calvarial bone in terms of mineralization, mineral crystallinity, and collagen crosslink ratios, but differed in the degree of collagen fibril order.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/148347/1/jrs5521_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/148347/2/jrs5521.pd

Failure to Attract and Retain Clinician/Scientist Faculty Puts Our Profession at Risk

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/67925/2/10.1177_00220345990780100101.pd

Compatibility of Staining Protocols for Bone Tissue with Raman Imaging

We report the use of Raman microscopy to image mouse calvaria stained with hematoxylin, eosin and toluidine blue. Raman imaging of stained specimens allows for direct correlation of histological and spectral information. A line-focus 785 nm laser imaging system with specialized near-infrared (NIR) microscope objectives and CCD detector were used to collect approximately 100 × 450 µm Raman images. Principal components analysis, a multivariate analysis technique, was used to determine whether the histological stains cause spectral interference (band shifts or intensity changes) or result in thermal damage to the examined tissue. Image analysis revealed factors for tissue components and the embedding medium, glycol methacrylate, only. Thus, Raman imaging proved to be compatible with histological stains such as hematoxylin, eosin and toluidine blue.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/48009/1/223_2003_Article_38.pd