Quantitative analysis of forward and backward second-harmonic images of collagen fibers using Fourier transform second-harmonic-generation microscopy.
ABSTRACT Fourier transform second-harmonic generation (SHG) microscopy has been applied to quantitatively compare the information content between SHG images obtained from the forward and backward direction for three tissue types: porcine tendon, sclera, and ear cartilage. Both signal types yield consistent information on the preferred orientation of collagen fibers. For all specimens, the Fourier transform of the forward and backward SHG images produces several overlapping peaks in the magnitude spectrum at various depths into the tissues, indicating that some information present in the forward SHG images can be extracted from the backward SHG images. This study highlights the potential of backward SHG microscopy for medical diagnostics.
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ABSTRACT: We propose the use of second-harmonic generation (SHG) microscopy for imaging collagen fibers in porcine femoral cortical bone. The technique is compared with scanning electron microscopy (SEM). SHG microscopy is shown to have excellent potential for bone imaging primarily due its intrinsic specificity to collagen fibers, which results in high contrast images without the need for specimen staining. Furthermore, this technique's ability to quantitatively assess collagen fiber organization is evaluated through an exploratory examination of bone structure as a function of age, from very young to mature bone. In particular, four different age groups: 1 month, 3.5 months, 6 months, and 30 months, were studied. Specifically, we employ the recently developed Fourier transform-second harmonic generation (FT-SHG) imaging technique for the quantification of the structural changes, and observe that as the bone develops, there is an overall reduction in porosity, the number of osteons increases, and the collagen fibers become comparatively more organized. It is also observed that the variations in structure across the whole cross-section of the bone increase with age. The results of this work show that quantitative SHG microscopy can serve as a valuable tool for evaluating the structural organization of collagen fibers in ex vivo bone studies.Bone 12/2011; 50(3):643-50. DOI:10.1016/j.bone.2011.11.013 · 4.46 Impact Factor
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ABSTRACT: Keratoconus is an eye disorder wherein the cornea weakens due to structural and/or compositional anomalies. This weakened cornea is no longer able to preserve its normal shape against the intraocular pressure in the eye and therefore bulges outward, leading to a conical shape and subsequent distorted vision. Changes in structure and composition often manifest as a change in shape (or geometry) as well as in mechanical and optical properties. Thus, understanding the properties and structure of keratoconic corneas could help elucidate etiology and pathogenesis, to develop treatments, and to understand other diseases of the eye. In this review, we discuss the changes in structure, composition, and mechanical and optical properties of the cornea with keratoconus. Current treatments for keratoconus and a novel proposed treatment using two-photon excitation therapy are also discussed. The intended audiences are mechanical engineers, materials engineers, optical engineers, and bioengineers.04/2011; 4(3):223-36. DOI:10.1016/j.jmbbm.2010.09.014
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ABSTRACT: We propose an innovative invasiveless technique in the field of nonlinear optical imaging to facilitate monitoring of cell/scaffold combinations for tissue repair. By using a near infrared (NIR) femtosecond excitation, we were able to introduce a new index based on decay time response for fluorescence (F) and Second Harmonic Generation (SHG) obtained with Time Correlated Single Photon Counting (TCSPC) microscopy to monitor structural information on the state of the matrix collagen. Some human Mesenchymal Stem Cells (hMSCs) seeded in 3D scaffolds were tested with different culture times (from D7 to D56) to analyze the effect of Tumor Growth Factor beta 1 (TGF-β1) on type-2 collagen expression in the matrix. After 14 days in the presence of TGF-β1, our results showed an increase in the expression of type-2 collagen synthesized by hMSCs, and a change in collagen conformation, as an indication of its ability to be detected as a harmonophore by TCSPC-SHG without the need for an exogenous probe.Bio-medical materials and engineering 01/2010; 20(3):183-8. DOI:10.3233/BME-2010-0630 · 0.85 Impact Factor