Multispectral oximetry of murine tendon microvasculature with inflammation

Abstract

We report a novel multispectral imaging technique for localised measurement of vascular oxygen saturation (SO) . Annular back-illumination is generated using a Schwarzchild-design reflective objective. Analysis of multispectral data is performed using a calibration-free oximetry algorithm. This technique is applied to oximetry in mice to measure SO in microvasculature supplying inflamed tendon tissue in the hind leg. Average SO for controls was 94.8 ± 7.0 % (N = 6), and 84.0 ± 13.5 % for mice with inflamed tendon tissue (N = 6). We believe this to be the first localised measurement of hypoxia in tendon microvasculature due to inflammation. Quantification of localised SO is important for the study of inflammatory diseases such as rheumatoid arthritis, where hypoxia is thought to play a role in pathogenesis.

Multi-spectral vascular oximetry of rat dorsal spinal cord

“…We described a visible-light multi-spectral microscope system for longitudinal oximetry studies in small animals. This device will continue to be improved and continue tested in the field, that includes modifications at the mathematical model establish by [7,11] for oximetry calculations, avoiding some errors when using two oximetry wavelengths [12,13] for multi imaging vasculature studies. Enabling a low-resource settings tool that will lead to better understanding of the progression of degenerative diseases based on the oxygenation levels, especially during the absence of oxygenation or hypoxia stages [14] .…”

Measurement of absorption in scattering media using objective laser speckle: application to blood oximetry

“…The measured spectrum is determined only by the measurand and geometry and is therefore insensitive to systematic errors introduced by contrast reduction in the instrumentation, or within the sample, which plague measurement based on the contrast of image features. This is particularly important since these contrast reductions can be very difficult to determine and calibrate particularly for biological measurements [12]. This is pertinent for oximetry in the eye, which has been restricted to oximetry of the retinal vasculature [8,30,32,42] and for which the occular media can reduce contrast by 50% or more.…”

“…Oximetry of vascular blood has been widely applied using calibrated variations in image contrast at two wavelengths [30] or by fitting the free parameters of a physical model to hyper-or multi-spectral images [31,32]. An accurate model of vascular contrast requires calibration of contrast reduction due to light scatter by the instrument, ocular media (in the case of retinal imaging) or by overlying biological tissue and some progress has been made in exploiting the additional data contained within multi-spectral images to calibrate these effects [12]. Our technique for measurement of chromophore concentration (such as is required for oximetry) has no requirement for structure within the object field and so is suitable, for example, for measuring oxygenation in the unresolved capillary bed of tissue.…”

“…Such intensity-based measurements are prone to significant uncertainty, which can lead to even larger relative errors in determination of c i . For example, in the measurement of the ratio of the relative concentration of vascular oxyhemoglobin in the retina [5][6][7][8][9][10][11], or within tissue [12], it is common to infer the ratio I t /I o from the contrast of images of the vasculature. For direct imaging of tissue with high-quality optical instruments, light scattering within the instrument can significantly reduce contrast, even for uncontaminated optics, reducing the validity of the Beer-Lambert law.…”

“…For direct imaging of tissue with high-quality optical instruments, light scattering within the instrument can significantly reduce contrast, even for uncontaminated optics, reducing the validity of the Beer-Lambert law. Contrast can be further reduced to very low levels by scattering due to overlying tissue, although it is possible to account for these effects by incorporation of scattering as a free parameter in a physical model for multispectral image formation of vasculature [12].…”