Measuring mucosal blood supply in vivo with a polarization-gating probe

Abstract

There has been significant interest in developing depth-selective optical interrogation of biological tissue in general and of superficial (e.g., mucosal) tissue in particular. We report an in vivo polarization-gating fiber-optic probe that obtains backscattering spectroscopic measurements from a range of near-surface depths (100-200 microm). The design and testing was performed with polarized light Monte Carlo simulations and in tissue model experiments. We used the probe to investigate mucosal changes in early carcinogenesis. Measurements performed in the colonic mucosa of 125 human subjects provide the first in vivo evidence that mucosal blood supply is increased early in carcinogenesis, not only in precancerous adenomatous lesions, but also in the histologically normal-appearing tissue surrounding these lesions. This effect was primarily limited to the mucosal microcirculation and was not present in the larger blood vessels located deeper in colonic tissue.

Fig. 1

Conceptualization of polarized light Monte Carlo numerical experiment. An infinitely narrow illumination fiber results in an illumination area with normal incidence onto the tissue surface. The collection area is identical to the illumination area and is controlled by the NA of the fibers and the focal length, f, of the lens.

Fig. 2

Experimental setup. A fiber optic probe designed to collect backscattering angles of approximately 10°–20° with a 700µm diameter illumination/collection area. The probe consists of an input fiber which is coupled to a Xenon lamp, and two output fibers that are connected to an imaging spectrometer shown in (a). (b) is a magnified view of the probe tip, which comes in contact with the sample.

Fig. 3

Schematic of probe tip showing the fibers, sheet polarizers, GRIN lens, and cover glass. The bottom left is a cross-section view showing the position of the orthogonally oriented polarizers with respect to the source fiber (S), co-polarized collection fiber ( ∥ ) and cross-polarized collection channel (⊥). The bottom right shows the location and size of illumination area, also corresponding to the location and size of the collection areas of the collecting fibers.

Fig. 4

Experimental verification of penetration depth. (a) shows representative data from a saturation curve experiment where the intensity is recorded as a function of the sample thickness, normalized by the maximum value (i.e. cumulative probability distribution). The intensity distribution as a function of depth in (b) is calculated by taking the derivative of the intensity vs. sample thickness curve, normalized by the total area. (c) represents average penetration depths for each of the polarization gating signals taken from samples with a range of optical properties (μ_s =35 – 111cm⁻¹, μ_a = 0, g = 0.8 – 0.92).

Fig. 5

Polarized light Monte Carlo results of penetration depth for each polarization signal (scaled by ls). (a) Average penetration depth for varying collection angles for each polarization gating signal (R/ls* = 0.5). (b) Average penetration depth for varying illumination/collection spot sizes for 9 +/− 9° backscattering collection angle (medium μ_s = 100cm⁻¹, μ_a = 0, g = 0.87).

Fig. 6

Calibration Curves obtained for delta-polarized (a), co-polarized (b), and cross-polarized (c) signals from the probe in phantoms with varying hemoglobin, and scattering coefficient. The region within the horizontal and vertical dashed lines indicates concentrations that are typical for colon tissue (experimentally determined). The anisotropy factor was kept constant for these measurements (g=0.88 at λ=540nm).

Fig. 7

Example spectra obtained from polarization gating probe in tissue. The model fit is shown as a solid line.

Fig. 8

Probe measurements of Hb content from adenomatous lesions and uninvolved mucosa away from the lesion as compared to control patients with no dysplasia. There is a large difference between the Hb content of normal tissue from patients with no dysplasia (white bars) and adenomatous tissue (dark gray bars) for all three polarization gating signals (p<0.01). There is also a significant difference between the Hb content from control patients and uninvolved mucosa measurements from colonic segments with adenomas. This difference in normal-appearing tissue progressively diminishes for probe signals with larger penetration depths (i.e. p=0.03 for 95µm depth, p=0.24 for 185µm depth).

Fig. 9

Comparison of Hb content from patients with varying significance of pre-cancerous lesions measured 10–30cm away from the lesion. Patients with Advanced Adenomas have the highest increase in Hb content away from the lesion. This increase in Hb progressively diminishes for polarization signals with larger penetration depths.