Jerome C. Mizeret

Clinical evaluation of a method for detecting superficial transitional cell carcinoma of the bladder by light-induced fluorescence of protoporphyrin IX following topical application of 5-aminolevulinic acid: Preliminary results

In bladder cancer, conventional white light endoscopic examination of the bladder does not provide adequate information about the presence of “flat” urothelial lesions such as carcinoma in situ. In the present investigation, we examine a new technique for the photodetection of such lesions by the imaging of protoporphyrin IX (PpIX) fluorescence following topical application of 5‐aminolevulinic acid (ALA).

Instrumentation for real-time fluorescence lifetime imaging in endoscopy

The fluorescence lifetime of living tissues is, in certain cases, related to their pathologic state and is therefore of interest for cancer detection. Measuring fluorescence lifetime in vivo during an endoscopic examination has thus been a challenging objective for several years. The present article deals with the development and first clinical trails of an instrumentation producing fluorescence lifetime images in real time. The acquisition of such fast phenomenon (nanosecond time scale) on an image has been made possible by using the homodyne detection approach, in which the excitation light and the detection gain are modulated in a phase-coherent way. Based on images acquired at different phase between the excitation and detection modulation, the fluorescence lifetime is calculated for each pixel of the image. Different configurations of excitation modulation characteristics (pulse train versus sine-wave amplitude modulation) have been investigated and compared using Fourier transforms. Interestingly, a...

Endoscopic tissue characterization by frequency-domain fluorescence lifetime imaging (FD-FLIM).

Tissue characterization by endoscopic fluorescence imaging of endogenous or exogenous fluorochromes is a promising method for early cancer detection. However, the steady-state fluorescence contrast between healthy tissue and lesions such as early-stage carcinomas is generally poor. The authors propose to improve this contrast by using the additional information contained in the fluorescence lifetime (FLT). The FLT of several fluorochromes is sensitive to their physico-chemical environment.The FLT can be measured by frequency-domain methods. The excitation light from a continuous wave (CW) laser is modulated in amplitude at radio-frequencies by an electro-optic modulator, and delivered to the tissue via an optical fibre. The endoscopie site is imaged by an endoscope on to an optical device. The gain of the fluorescence image detector is also modulated at the same frequency for homodyning. The tissue fluorescence image is recorded at several phases between the excitation and the detection modulations during an acquisition cycle. With these images, an image processor calculates the apparent FLT for each pixel and constructs a lifetime image of the endoscopie site. This process is performed at quasi-video frequencies.The influence of various physical parameters (modulation frequency, number of images by cycle, shot noise, tissue optical properties etc.) has been investigated by analytical analysis, simulation methods and experimentation.Preliminary results obtained on human tissues are also presented to illustrate the potentiality of the method.

Clinical assessment of fluorescence cystoscopy during transurethral bladder resection in superficial bladder cancer

The prognosis of superficial bladder cancer in terms of recurrence and disease progression is related to bladder tumor multiplicity and the presence of concomitant “plane” tumors such as high-grade dysplasia and carcinoma in situ. This study in 33 patients aimed to demonstrate the role of fluorescence cystoscopy in transurethral resection of superficial bladder cancer. The method is based on the detection of protoporphyrin-IX-induced fluorescence in urothelial cancer cells by topical administration of 5-aminolevulinic acid. The sensitivity and the specificity of this procedure on apparently normal mucosa in superficial bladder cancer are estimated to be 82.9% and 81.3%, respectively. Thus, fluorescence cytoscopy is a simple and reliable method for mapping the bladder mucosa, especially in the case of multifocal bladder disease, and it facilitates the screening of occult dysplasia.

Frequency-domain fluorescence lifetime imaging for endoscopic clinical cancer photodetection: Apparatus design and preliminary results

We describe a new fluorescence imaging device for clinical cancer photodetection in hollow organs in which the tumor/normal tissue contrast is derived from the fluorescence lifetime of endogenous or exogenous fluorochromes. This fluorescence lifetime contrast gives information about the physicochemical properties of the environment which are different between normal and certain diseased tissues. The excitation light from a CW laser is modulated in amplitude at a radio frequency by an electrooptical modulator and delivered by an optical fiber through an endoscope to the hollow organ. The image of the tissue collected by the endoscope is separated in two spectral windows, one being the backscattered excitation light and the other the fluorescence of the fluorochrome. Each image is then focused on the photocathode of image intensifiers (II) whose optical gain is modulated at the same frequency as the excitation intensity, resulting in homodyne phase-sensitive images. By acquiring stationary phase-sensitive frames at different phases between the excitation and the detection, it is possible to calculate in quasi-real time the apparent fluorescence lifetime of the corresponding tissue region for each pixel. A result obtained by investigating the endogenous fluorochromes present in the mucous membrane of an excised human bladder is presented to illustrate this method and most of the optical parameters which are of major importance for this photodetection modality have been evaluated.

Cylindrical fiberoptic light diffuser for medical applications

A cylindrical light diffuser has been developed mainly for medical applications, including photodynamic therapy (PDT), in particular interstitial PDT, PDT of the bronchi, or intravascular PDT.

Fluorescence excitation and emission spectra of ALA-induced protoporphyrin IX in normal and tumoral tissue of the human bladder

In vivo spectrofluorometric analysis represents a tool to obtain information about fluorophore distribution in tissue. Based on a Peltier-cooled CCD we designed a fluorescence excitation and emission spectrograph which allows to obtain tissue spectra endoscopically and in a clinical environment. Clinical studies were performed on patients with positive cytology or tumor recurrence in the urinary bladder. Patients received a 50 ml instillation of 3% ALA solution at pH 5.5 during 3 to 4 hours and underwent a normal white light cystoscopic examination together with light induced fluorescence photodetection at 5 to 8 hours after the beginning of the instillation. Local fluorescence measurements with a single fiber were performed before photodetection. These showed fluorescence ratios between tumor and normal tissue of 1.5 to 20 with the strongest ratios for exophytic papillary tumors. Fluorescence excitation between 380 nm and 450 nm revealed that the higher Protoporphyrin IX (PPIX) signal on tumor tissue is accompanied by a decrease of the autofluorescence at the emission wavelength of 500 nm.

Endoscopic frequency-domain fluorescence lifetime imaging for clinical cancer photodetection: apparatus design

We describe a new fluorescence imaging device for clinical cancer photodetection in hollow organs in which the image contrast is derived from the fluorescence lifetime of the fluorochrome at each point in a 2D image. Lifetime images are created from a series of images obtained from two gain-modulated image intensifiers. One of them (II-1) detects the light-induced tissue fluorescence, whereas the other (II-2) detects the backscattered fluorescence excitation light. This light is modulated at the same frequency as the detectors, resulting in homodyne phase-sensitive image. These stationary phase-sensitive images are collected using two CCD cameras, digitized and manipulated with a mathematical operator in real time. A series of such images, obtained with both image intensifiers at various phase shifts between their gain modulation and the modulation of the excitation light, is used to determine phase angle and/or the modulation of the fluorescence emission at each pixel. The reference values of these phase angles and modulations are obtained with II-2, whereas II-1 enables the measurement of the phase and modulation of the fluorescence. Phase and modulation are related to the fluorescence lifetime of the fluorochrome. An advantage of the experimental method proposed here is that pixel-to-pixel scanning is not required to obtain the fluorescence lifetime image, as the information from all pixels is obtained at the same time.

Clinical measurements of tissue optical properties in the esophagus

A noninvasive probe has been devised and clinically used to perform in vivo measurements of the optical properties of the esophageal wall. The absorption coefficient and the effective scattering coefficient are determined from the observation of the spatial distribution of the diffuse reflectance at the tissue surface. Results, obtained at 514 and 630 nm, i.e. wavelengths of interest in photodynamic therapy with actual clinically used photosensitizers are summarized. An efficient and easily-built isotropic microprobe is also presented which is suitable to measure in situ the radiant energy fluence rate distribution in tissue.

Endoscopic tissue fluorescence life-time imaging by frequency doamin light-induced fluorescence

An instrumentation is being developed to draw a fluorescence life-time map of tissue endoscopically. This fluorescence life-time of an endogenous or exogenous fluorochrome gives information about the physico-chemical environment which is thought to vary between normal and diseased tissue. The excitation light from a cw laser is modulated in amplitude at high frequencies by an electro-optic modulator and delivered to the endoscopic site through an optical fiber. The image of the tissue is spectrally split in two parts, the one being the backscattered excitation light, the other the fluorescence of the fluorochromes. Each image is focused on the photocathode of an image intensifier whose gain is modulated at the same frequency. By acquiring frames at different phases between the excitation and the emission, it is possible to calculate pixel by pixel the apparent fluorescence life-time of the corresponding tissue region.