Second harmonic generation imaging microscopy studies of osteogenesis imperfecta

Abstract

We have used quantitative second harmonic generation (SHG) imaging microscopy to investigate the collagen matrix organization in the oim mouse model for human osteogenesis imperfecta (OI). OI is a heritable disease in which the type I collagen fibrils are either abnormally organized or small, resulting in a clinical presentation of recurrent bone fractures and other pathologies related to collagen-comprised tissues. Exploiting the exquisite sensitivity of SHG to supramolecular assembly, we investigated whether this approach can be utilized to differentiate normal and oim tissues. By comparing SHG intensity, fibrillar morphology, polarization anisotropy, and signal directionality, we show that statistically different results are obtained for the wild type (WT) and disease states in bone, tendon, and skin. All these optical signatures are consistent with the collagen matrix in the oim tissues being more disordered, and these results are further consistent with the known weaker mechanical properties of the oim mouse. While the current work shows the ability of SHG to differentiate normal and diseased states in a mouse model, we suggest that our results provide a framework for using SHG as a clinical diagnostic tool for human OI. We further suggest that the SHG metrics described could be applied to other connective tissue disorders that are characterized by abnormal collagen assembly.

Phase matching considerations in second harmonic generation from tissues: Effects on emission directionality, conversion efficiency and observed morphology

“…Experimentally we observed this trend and measured a ratio of approximately 2.5. (16) This demonstrates that phase matching considerations based on domain size and randomness can be used to make comparisons of the relative emission intensities in different tissues.…”

“…This is because many diseases are characterized by defects or changes in the assembly of collagen in the respective tissue and we thus suggest that the SHG imaging modality has great potential as a clinical diagnostic tool. For example, the connective tissue disorder Osteogenesis Imperfecta (OI) is characterized by abnormal collagen assembly and we have shown that SHG can reveal differences in the morphology not possible by other optical methods (16). Additionally, SHG has also shown early promise in imaging cancer since malignant tumors often have abnormal collagen assembly relative to normal tissue (9,17).…”

Precise, motion-free polarization control in Second Harmonic Generation microscopy using a liquid crystal modulator in the infinity space

“…However, there remains great potential in using detailed polarization analysis to classify normal and diseased tissues, such as cancer and connective tissue disorders, which often have different fibrillar properties. For example, we showed that the SHG anisotropy [40] and SHG-CD responses [33] in the connective tissue disorder Osteogenesis imperfecta (OI) were different than in normal tissues. We also recently showed that SHG polarization signatures could differentiate different collagen isoforms in self-assembled collagen gels which are models for stromal changes in ovarian cancer (Biophysical Journal, submitted).…”

From molecular structure to tissue architecture: collagen organization probed by SHG microscopy

“…In fact, SHG microscopy highlights morphologic changes in collagen structure, which indicate particular disease states, such as tumour invasiveness, as well as indicators of collagen remodelling in tumour stroma, which is playing a key-role in the tumour development from in-situ to invasive stage. Recently, SHG microscopy has been successfully applied to the study of altered physiological conditions of various kinds of tissues, including muscle [94], bones [95,96], and cartilages [97,98]. SHG can be easily combined with TPEF microscopy to realize a particularly powerful tool for connective tissue imaging.…”