Perillyl alcohol inhibits TCR-mediated [Ca(2+)](i) signaling, alters cell shape and motility, and induces apoptosis in T lymphocytes.

Perillyl alcohol (POH) inhibits isoprenylation and has shown anticancer and chemopreventive properties in rodent models. The mechanism that underlies the anticancer activity of POH and other isoprenylation inhibitors is unknown but has been postulated to involve decreased levels of isoprenylated Ras and Ras-related proteins. Previously we demonstrated that POH effectively inhibits human T cell proliferation in vitro and can prevent acute and chronic rejection in a rat cardiac transplant model.

Characterization of cellular optoporation with distance.

We have developed and characterized cellular optoporation with visible wavelengths of light using standard uncoated glass cover slips as the absorptive media. A frequency-doubled Nd:YAG laser pulse was focused at the interface of the glass surface and aqueous buffer, creating a stress wave and transiently permeabilizing nearby cells. Following optoporation of adherent cells, three spatial zones were present which were distinguished by the viability of the cells and the loading efficiency (or number of extracellular molecules loaded).

An indwelling brachytherapy balloon catheter: potential use as an intracranial light applicator for photodynamic therapy.

An indwelling balloon applicator developed for postoperative intracavity afterloading brachytherapy was evaluated for photodynamic therapy (PDT). Following tumor resection, the applicator is positioned in the center of the resultant cavity and the balloon is inflated with a scattering solution. The liquid-filled balloon stabilizes the resection cavity ensuring a constant and simple geometric shape during treatment.

Minimally-invasive debulking of ovarian cancer in the rat pelvis by means of photodynamic therapy using the pegylated photosensitizer PEG-m-THPC.

Interstitial photodynamic therapy (PDT) using the pegylated photosensitizer PEG-m-THPC was evaluated as a minimally-invasive procedure to selectively debulk unrespectable pelvic ovarian cancer (NuTu-19) in immunocompetent rats. To assess tumour selectivity, PEG-m-THPC at dosages of 0.3, 3.

Quantitative near-infrared spectroscopy of cervical dysplasia in vivo.

The aims of this study were: (i) to quantify near-infrared optical properties of normal cervical tissues and high-grade squamous intra-epithelial lesions (H-SIL); (ii) to assess the feasibility of differentiating normal cervical tissues from H-SIL on the basis of these properties; and (iii) to determine how cervical tissue optical properties change following photodynamic therapy (PDT) of H-SIL in vivo. Using the frequency domain photon migration technique, non-invasive measurements of normal and dysplastic ecto-cervical tissue optical properties, i.e.

Highly selective targeting of ovarian cancer with the photosensitizer PEG-m-THPC in a rat model.

Photodynamic therapy (PDT) uses light to activate a photosensitizer that has been absorbed or retained preferentially by cancer cells after systemic administration. The first pegylated photosensitizer, tetrakis-(m-methoxypolyethylene glycol) derivative of 7,8-dihydro-5,10,15,20-tetrakis(3-hydroxyphenyl)-21,23-[H]-porphyrin (PEG-m-THPC), was evaluated to target selectively unresectable pelvic ovarian cancer bulks. Our goals were two-fold: (1) to establish an ovarian cancer model suitable for the development of debulking techniques and (2) to characterize the pharmacokinetics and tumor selectivity of PEG-m-THPC by fluorescence microscopy.

Mapping the sensitivity of T cells with an optical trap: polarity and minimal number of receptors for Ca(2+) signaling.

Contact with antigen-presenting cells (APCs) initiates an activation cascade within T lymphocytes, including a rise in cytosolic calcium, lymphokine production, and cell division. Although T cell-APC physical contact is required for an immune response, little is known about the patterns of cellular interactions and their relation to activation. Calcium imaging combined with an optical trap enabled the T cell contact requirements and polarity to be investigated at the single-cell level.

Reflectance measurements of layered media with diffuse photon-density waves: a potential tool for evaluating deep burns and subcutaneous lesions.

The basic principles of a non-contact, near-infrared technique for the mapping of layered tissues are discussed theoretically and verified experimentally. The propagation properties of diffuse photon-density waves in tissues depend on the optical properties of the tissue. When a layered medium is irradiated by amplitude modulated light, the difference in optical properties between the layers is evident in the phase and amplitude of the diffuse reflection coefficient, which is a result of the interference of the partial waves propagating in the different layers.

Cell Viability and DNA Denaturation Measurements by Two-Photon Fluorescence Excitation in CW Al:GaAs Diode Laser Optical Traps.

Cell viability and DNA denaturation are measured through two-photon fluorescence excitation using a single diode laser beam as the trapping and exciting source simultaneously. Two-photon fluorescence emission spectra are presented for CHO cells and T lymphocytes loaded with various fluorescent probes. This single beam method is demonstrated to be a safe tool to monitor the viability of optically trapped cells, even under intense 809 nm diode laser illumination (∼106 W/cm2).

Intrauterine light probe for photodynamic ablation therapy.

Background: Endometrial ablation by means of photodynamic therapy is currently being evaluated as an outpatient treatment for dysfunctional uterine bleeding. Photodynamic therapy requires the activation of a photosensitizer by laser light. We describe a new device specifically designed to provide light delivery to the uterus for endometrial photodynamic therapy.

Laser-micropipet combination for single-cell analysis.

Due to its potential for exquisite mass detection limits and resolving power, capillary electrophoresis is used for biochemical measurements on single cells; however, accurate measurements of many physiological parameters require sampling strategies that are considerably faster than those presently available. We have developed a laser-based technique to lyse single, adherent, mammalian cells on millisecond time scales. The cellular contents are then introduced into a capillary where electrophoretic separation and detection are performed.

Uptake of the photosensitizer benzoporphyrin derivative in human endometrium after topical application in vivo.

Study Objective: To determine both the time leading to maximum endometrial drug uptake and distribution of the photosensitizer benzoporphyrin derivative-monoacid ring A (BPD-MA) after intrauterine instillation (Canadian Task Force classification ).

Design: Assessment of histology specimens (Canadian Task Force classification I).

Setting: University-based facility.

In vivo detection of metastatic ovarian cancer by means of 5-aminolevulinic acid-induced fluorescence in a rat model.

Study Objectives: To determine the feasibility of macroscopic visualization of small ovarian cancer metastases in vivo by fluorescence after intravenous administration of 5-aminolevulinic acid (ALA); to assess the time after drug injection when fluorescence of small metastases is maximum; and to correlate macroscopic in vivo fluorescence with both microscopic ex vivo fluorescence and histologic findings.

Design: Controlled animal study (Canadian Task Force classification I).

Setting: University-based facility.

Laser scissors and tweezers.

In summary, we described the use of laser scissors and tweezers from three perspectives: (a) the historical background from which these two techniques evolved, (b) an understanding and lack of understanding of the mechanisms of interaction with the biological systems, and (c) the applications of the scissors and tweezers alone and in combination. As the technology improves and we gain a better understanding of how these two tools operate they will become even more useful in probing cell structure and function, as well as practically manipulating cells in genetics, oncology, and developmental biology.

In vivo fluorescence detection of ovarian cancer in the NuTu-19 epithelial ovarian cancer animal model using 5-aminolevulinic acid (ALA).

The purpose of this study was to determine whether in vivo fluorescence detection of protoporphyrin IX (PpIX) could be used to identify intraperitoneal micrometastases of epithelial ovarian carcinoma after application of 5-aminolevulinic acid (ALA). ALA was applied intraperitoneal at different concentrations (25, 50, and 100 mg/kg) and iv (100 mg/kg) to immunocompetent Fischer 344 rats bearing a syngeneic epithelial ovarian carcinoma. At different time intervals after ALA administration (1.

Non-invasive measurements of breast tissue optical properties using frequency-domain photon migration.

A multiwavelength, high bandwidth (1 GHz) frequency-domain photon migration (FDPM) instrument has been developed for quantitative, non-invasive measurements of tissue optical and physiological properties. The instrument produces 300 kHz to 1 GHz photon density waves (PDWs) in optically turbid media using a network analyser, an avalanche photodiode detector and four amplitude-modulated diode lasers (674 nm, 811 nm, 849 nm, and 956 nm). The frequency of PDW phase and amplitude is measured and compared to analytically derived model functions in order to calculate absorption, mu a, and reduced scattering, mu s, parameters.

Frequency-domain photon migration measurements of normal and malignant tissue optical properties in a human subject.

A 1-GHz multifrequency, multiwavelength frequency-domain photon migration instrument is used to measure quantitatively the optical absorption (mu(a)) and effective optical scattering (mu(s) ?) of normal and malignant tissues in a human subject. Large ellipsoidal (~10-cm major axis, ~6-cm minor axes) subcutaneous malignant lesions were compared with adjacent normal sites in the abdomen and back. Absorption coefficients recorded at 674, 811, 849, and 956 nm were used to calculate tissue hemoglobin concentration (oxyhemoglobin, deoxyhemoglobin, and total), water concentration, hemoglobin oxygen saturation, and blood volume fraction in vivo.

Comparison of Ho:YAG versus Nd:YAG thoracoscopic laser treatment of pulmonary bullae in a rabbit model.

Objective: To determine the relative efficacy and morbidity of Ho:YAG versus Nd:YAG laser treatment of bullous lung disease in an animal model.

Summary Background Data: Laser coagulation procedures for treatment of emphysematous pulmonary bullae and heterogeneous emphysema continue to evolve. The role of lasers in lung volume reduction surgery remains controversial due to issues of relative efficacy and morbidity.

Giant cell formation in cells exposed to 740 nm and 760 nm optical traps.

Background And Objective: Optical trapping is becoming a useful and widespread technique for the micromanipulation of cells and organelles. Giant cell formation following optical trapping was studied to detect the potential adverse effects.

Study Design/materials And Methods: The nuclei of preselected single CHO cells were exposed to 740 nm and 760 nm laser microbeam generated by a titanium-sapphire tunable laser at 88 and 176 mW and different time exposures.

Time-resolved and steady-state fluorescence measurements of beta-nicotinamide adenine dinucleotide-alcohol dehydrogenase complex during UVA exposure.

beta-nicotinamide adenine dinucleotide (NADH)-alcohol dehydrogenase complex was compared to either UVA irradiation (364 nm; 50 mW cm-2; 0-60 min) or heat in order to investigate complex stability. Prior to irradiation, frequency-domain fluorescence lifetime measurements indicated the presence of two principal components having short (subnanosecond) and long (nanosecond) fluorescence lifetimes reflecting free and bound NADH respectively. UVA exposure resulted in decreased NADH fluorescence intensity concomitant with decreased absorption at 337 nm.

Selective photosensitizer localization in the human endometrium after intrauterine application of 5-aminolevulinic acid.

Objective: Our purpose was twofold: to determine the distribution of the endogenous photosensitizer protoporphyrin IX in the uterus and to ascertain the time interval leading to maximal endometrial fluorescence after intrauterine instillation of 5-aminolevulinic acid.

Study Design: One milliliter of a 400 mg/ml 5-aminolevulinic acid-Hyskon solution was instilled into the uterine cavity of 27 women before hysterectomy. On frozen sections of uterine samples 5-aminolevulinic acid-induced fluorescence was measured with fluorescence microscopy.

Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa.

Photostress has to be considered during optical micromanipulation of gametes. Ultraviolet light, including low-energy UVA (32-400 nm) radiation, as well as high-intensity near infrared (NIR) laser radiation may induce cell damage. A total number of 580 light-exposed sperm cells were studied in single-cell photostress experiments.

Physiological monitoring of optically trapped cells: assessing the effects of confinement by 1064-nm laser tweezers using microfluorometry.

We report the results of microfluorometric measurements of physiological changes in optically trapped immotile Chinese hamster ovary cells (CHOs) and motile human sperm cells under continuous-wave (CW) and pulsed-mode trapping conditions at 1064 nm. The fluorescence spectra derived from the exogenous fluorescent probes laurdan, acridine orange, propidium iodide, and Snarf are used to assess the effects of optical confinement with respect to temperature, DNA structure, cell viability, and intracellular pH, respectively. In the latter three cases, fluorescence is excited via a two-photon process, using a CW laser trap as the fluorescence excitation source.