Time-resolved diffusing wave spectroscopy applied to dynamic heterogeneity imaging.

We report what is to our knowledge the first observation of a time-resolved diffusing wave spectroscopy (DWS) signal recorded by transillumination through a thick turbid medium: the DWS signal is measured for a fixed photon transit time, which opens the possibility of improving the spatial resolution. This technique could find biomedical applications, especially in mammography.

Time-resolved diffusing wave spectroscopy beyond 300 transport mean free paths.

We presented theoretical and experimental demonstrations of the possibilities of performing time-resolved diffusing wave spectroscopy: We successfully registered field fluctuations for selected photon path lengths that can surpass 300 transport mean free paths. Such performance opens new possibilities for biomedical optics applications.

Time-resolved measurements from speckle interferometry.

We present time-resolved measurements by speckle interferometry of the light scattered by a liquid medium. Measurements were performed by use of reflectance geometry and are compared with results obtained in the same conditions with a femtosecond laser and a streak camera. The setup was also tested in vivo on the forearm of a human volunteer to demonstrate the potential utility of such a setup for biomedical applications.

Synthesis and binding properties of photoactivable biotin-conjugated verapamil derivatives for the study of P-170 glycoprotein.

The design and synthesis of two photoactivable biotin-labeled analogues of verapamil (6 and 7) is reported. Preliminary evaluation of the biological profile of 6 (EDP 137) and 7 (EDP 141) shows that they have comparable affinities to that of verapamil for P-170, the protein responsible for multidrug resistance (MDR). Since both appear to bind irreversibly to the protein and the presence of biotin in their structure makes them easily detectable by avidin, they promise to be of great help in studying the protein and its mechanism of action.

Correlation between the kinetics of anthracycline uptake and the resistance factor in cancer cells expressing the multidrug resistance protein or the P-glycoprotein.

Multidrug resistance (MDR) in model systems is known to be conferred by two different integral proteins, the 170-kDa P-glycoprotein (Pgp) and the 190-kDa multidrug resistance-associated protein (MRP1). One possible pharmacological approach to overcome drug resistance is the use of specific inhibitors, which enhance the cytotoxicity of known antineoplastic agents. However, while many compounds have been proven to be very efficient in inhibiting Pgp activity only some of them are able to inhibit MRP1. The other likely approach is based on the design and synthesis of new non-cross-resistant drugs with physicochemical properties favoring the uptake of the drug by the resistant cells. The intracellular drug retention influences its cytotoxic effect. The level of the intracellular drug content is a function of the amount of drug transported inside the cell (influx) and the amount of drug expelled from the cell (efflux). In this work, the kinetics of drug uptake and the kinetics of active efflux of several anthracycline derivatives in both Pgp expressing K562/Adr cells and MRP1 expressing GLC4/Adr cells was determined. Our data have shown that in both cell lines there is no correlation between the resistance factor and the kinetics of drug efflux by these pumping systems. However, a very good correlation between the resistance factor and the kinetics of drug uptake has been established in both cell lines: the resistance factor decreases when the kinetics of drug uptake increases. This work has clearly shown that when the rate of transmembrane transport of anthracycline is high enough, the efflux mediated by the protein transporter is not able to pace with it. The protein transporter essentially operates in a futile cycle and the resistance factor is tending to one. It does not mean, however, that when the resistance factor is close to one the anthracycline is not transported by the pump.

The role of laser-induced autofluorescence spectroscopy in bladder tumor detection. Dependence on the excitation wavelength.

We assessed the ability of laser-induced autofluorescence spectroscopy to distinguish neoplastic urothelial bladder lesions from normal or nonspecific inflammatory mucosa. Three different pulsed laser excitation wavelengths were used successively: 308 nm (xenium chloride excimer laser), 337 nm (nitrogen laser) and 480 nm (coumarin dye laser). The excitation light was delivered by a specially devised multifiber catheter connected to a 1-mm core diameter silica monofiber introduced through the working channel of a standard cystoscope with saline irrigation. The captured fluorescence light was focused onto an optical multichannel analyzer detection system. Performance of this device was evaluated in 25 patients after obtaining consent and immediately before transurethral resection of a bladder tumor. Spectroscopic results were compared with histological findings. At 337- and 480-nm excitation wavelengths, the overall fluorescence intensity of bladder tumors was clearly decreased compared to normal urothelial mucosa regardless of tumor stage and grade. At the 308-nm excitation wavelength, the shape of the tumor spectra, including carcinoma in situ, was markedly different from that of normal or nonspecific inflammatory mucosa. No absolute intensity determinations were required in this situation, since a definite diagnosis could be established based on the fluorescence intensity ratio at 360 and 440 nm. This spectroscopic study could be particularly useful in designing a simplified autofluorescence imaging device for detection of occult urothelial neoplasms.

Influence of the emission-reception geometry in laser-induced fluorescence spectra from turbid media.

Routine clinical detection of precancerous lesions by laser-inducedautofluorescence was recently demonstrated in several medicalfields. This technique is based on the analysis of complex spectrawith overlapping broad structures. However, in biological tissues, scattering and absorption are wavelength dependent, and the observedfluorescence signals are distorted when the illumination and detectiongeometry varies, making comparison of results from different groupsdifficult. We study this phenomenon experimentally in human tissuein a simple experiment: A fiber is used for the excitation and anidentical fiber is used for reception of the signal; both fibers aremaintained in contact with the tissue. We study the distortion ofthe spectra as a function of the distance between the twofibers. For correction of the spectra we show that it is possibleto use a fast and accurate ab initio Monte Carlo simulationwhen the spectral variations of the optical properties of the mediumare known. The main advantage of this simulation is itsapplicability even for complex boundary conditions or when the sampleconsists of several layers.

Laser induced autofluorescence diagnosis of bladder tumors: dependence on the excitation wavelength.

PURPOSE: We assessed the ability of laser induced autofluorescence spectroscopy to distinguish neoplastic urothelial bladder lesions from normal or nonspecific inflammatory mucosa. MATERIALS AND METHODS: Three different pulsed laser excitation wavelengths were used successively: 308 nm. (xenium chloride excimer laser), 337 nm. (nitrogen laser) and 480 nm. (coumarin dye laser). The excitation light was delivered by a specially devised multifiber catheter connected to a 1 mm. core diameter silica monofiber introduced through the working channel of a standard cystoscope with saline irrigation. The captured fluorescence light was focused onto an optical multichannel analyzer detection system. Device performance was evaluated in 25 patients after obtaining consent and immediately before transurethral resection of a bladder tumor. Spectroscopic results were compared with histological findings. RESULTS: At 337 and 480 nm. excitation wavelengths the overall fluorescence intensity of bladder tumors was clearly decreased compared to normal urothelial mucosa regardless of tumor stage and grade. At the 308 nm. excitation wavelength the shape of the tumor spectra, including carcinoma in situ, was markedly different from that of normal or nonspecific inflammatory mucosa. No absolute intensity determinations were required in this situation, since a definite diagnosis could be established based on the fluorescence intensity ratio at 360 and 440 nm. CONCLUSIONS: This spectroscopic study could be particularly useful to design a simplified autofluorescence imaging device for detection of occult urothelial neoplasms.

Argon laser induced autofluorescence may distinguish between normal and tumor human urothelial cells: a microspectrofluorimetric study.

PURPOSE: To assess the ability of argon laser-induced autofluorescence spectroscopy (LIAFS) to discriminate normal from tumor human urothelial cells. MATERIALS AND METHODS: Emission spectra of single living cells excited at 488 nm. have been studied with confocal microspectrofluorimeter. RESULTS: Cellular autofluorescence appeared as a broad band with a maximum in the same "green" spectral range, 550 to 560 nm., probably corresponding to oxidized flavoprotein emission. However, the maximum autofluorescence intensity of normal urothelial cells was much higher, 10 times (p<0.0001) that of any of the tumor cell types tested. CONCLUSIONS: These results, suggesting a significantly reduced oxidized flavoprotein concentration in tumor urothelial cells, should prompt us to evaluate argon LIAFS as a potential tool to detect occult urothelial severe dysplasia and carcinoma in situ.