Effect of low intensity laser interaction with human skin fibroblast cells using fiber-optic nano-probes.

Over the past forty years, many efforts have been devoted to study low power laser light interactions with biological systems. Some of the investigations were performed in-vitro, on bulk cell populations. Our present work was undertaken to apply specially engineered fiber-optic based nano-probes for the precise delivery of laser light on to a single cell and to observe production of low power laser light induced reactive oxygen species (ROS). A normal human skin fibroblast (NHF) cell line was utilized in this investigation and the cells were irradiated under two different schemes of exposure: (1) an entire NHF cell population within a Petri dish using a fan beam methodology, and (2) through the precise delivery of laser energy on to a single NHF cell using fiber-optic nano-probe. Photobiostimulative studies were conducted through variation of laser intensity, exposure time, and the energy dose of exposure. Laser irradiation induced enhancement in the rate of cell proliferation was observed to be dependent on laser exposure parameters and the method of laser delivery. The total energy dose (fluence) had a greater influence on the enhancement in the rate of cellular proliferation than compared to laser intensity. The enhancement in the growth rate was observed to have a finite life-time of several days after the initial laser exposure. Fluorescent life-time imaging of ROS was performed during the nano-based single cell exposure method. The kinetics of ROS generation was found to depend strongly on the laser fluence and not on the laser intensity.

The electric field induced by light can explain cellular responses to electromagnetic energy: a hypothesis of mechanism.

When cells are irradiated with visible and near-infrared wavelengths a variety of stimulatory effects are observed in their metabolism. To explain the observed light effects, researchers try to identify the chromophores that are involved in the processes. However, the mechanism of light absorption by a chromophore does not explain many of the experimental observations and therefore the primary mechanism for cellular light responses remains unproven. In addition to the ability of photons to produce electronic excitation in chromophores, light induces a wave-like alternating electric field in a medium that is able to interact with polar structures and produce dipole transitions. These dipole transitions are analyzed in the present article at different cellular and biochemical levels, leading to the proposal that the primary mechanism for the observed light effects is related to the light-induced electric field.

Modification of the intrinsic fluorescence and the biochemical behavior of ATP after irradiation with visible and near-infrared laser light.

In this work, the effects of visible (655 nm) and near-infrared (830 nm) light on ATP in solution were examined. The addition of irradiated ATP to the hexokinase reaction caused significant differences in the reaction rates and in the Michaelis-Menten kinetic parameters, k(m) and v(max). Irradiated ATP cleavage by hexokinase occurred in less time. Changes were wavelength and dose dependent. Excitation of ATP with a 260 nm wavelength ultraviolet light induced a fluorescence emission that was decreased when Mg2+ was added due to ion binding of the phosphates, which are the structures that modify the fluorescence produced by the adenine dipoles. The irradiation of this ATP.Mg2+ solution using 655 and 830 nm light increased the fluorescence by a possible displacement of Mg2+ from the phosphates. In conclusion, visible and near-infrared light modifies the biochemical behavior of ATP in the hexokinase reaction and the fluorescence intensity of the molecule thus altering the Mg2+ binding strength to the oxygen atoms in the phosphate group.

Low power laser irradiation alters gene expression of olfactory ensheathing cells in vitro.

BACKGROUND AND OBJECTIVES: Both photobiomodulation (PBM) and olfactory ensheathing cells (OECs) transplantation improve recovery following spinal cord injury. However, neither the combination of these two therapies nor the effect of light on OECs has been reported. The purpose of this study was to determine the effect of light on OEC activity in vitro. MATERIALS AND METHODS: OECs were purified from adult rat olfactory bulbs and exposed to 810 nm light (150 mW; 0, 0.2, or 68 J/cm(2)). After 7-21 days in vitro, cells underwent immunocytochemistry or RNA extraction and RT-PCR. RESULTS: Analysis of immunolabeling revealed a significant decrease in fibronectin expression in the cultures receiving 68 J/cm(2). Analysis of gene expression revealed a significant (P < 0.05) increase in brain derived neurotrophic factor (BDNF), glial derived neurotrophic factor (GDNF), and collagen expression in the 0.2 J/cm(2) group in comparison to the non-irradiated and 68 J/cm(2) groups. OEC proliferation was also found to significantly increase in both light treated groups in comparison to the control group (P < 0.001). CONCLUSIONS: These results demonstrate that low and high dosages of PBM alter OEC activity, including upregulation of a number of neurotrophic growth factors and extracellular matrix proteins known to support neurite outgrowth. Therefore, the application of PBM in conjunction with OEC transplantation warrants consideration as a potential combination therapy for spinal cord injury.

Light promotes regeneration and functional recovery and alters the immune response after spinal cord injury.

BACKGROUND AND OBJECTIVES: Photobiomodulation (PBM) has been proposed as a potential therapy for spinal cord injury (SCI). We aimed to demonstrate that 810 nm light can penetrate deep into the body and promote neuronal regeneration and functional recovery. STUDY DESIGN/MATERIALS AND METHODS: Adult rats underwent a T9 dorsal hemisection, followed by treatment with an 810 nm, 150 mW diode laser (dosage = 1,589 J/cm2). Axonal regeneration and functional recovery were assessed using single and double label tract tracing and various locomotor tasks. The immune response within the spinal cord was also assessed. RESULTS: PBM, with 6% power penetration to the spinal cord depth, significantly increased axonal number and distance of regrowth (P < 0.001). PBM also returned aspects of function to baseline levels and significantly suppressed immune cell activation and cytokine/chemokine expression. CONCLUSION: Our results demonstrate that light, delivered transcutaneously, improves recovery after injury and suggests that light will be a useful treatment for human SCI.

Photobiomodulation improves cutaneous wound healing in an animal model of type II diabetes.

OBJECTIVE: We investigated the effects of photobiomodulation (PBM) on cutaneous wound healing in an animal model of type II diabetes, Psammomys obesus (Sand Rats). BACKGROUND DATA: 632-nm light has been established as the most effective wavelength for treatment of cutaneous wounds; however, the inconsistent efficacy of PBM may be due to inadequate treatment parameter selection. METHODS: Using 632-nm light, an initial series of experiments were done to establish optimal treatment parameters for this model. Following creation of bilateral full-thickness skin wounds, non-diabetic Sand Rats were treated with PBM of differing dosages. Wound healing was assessed according to wound closure and histological characteristics of healing. Optimal treatment parameters were then used to treat type II diabetic Sand Rats while a diabetic control group received no irradiation. In order to elucidate the mechanism behind an improvement in wound healing, expression of basic fibroblast growth factor (bFGF) was assessed. RESULTS: Significant improvement in wound healing histology and wound closure were found following treatment with 4 J/cm(2) (16 mW, 250-sec treatments for 4 consecutive days; p < 0.05). The 4 J/cm(2) dosage significantly improved histology and closure of wounds in the diabetic group in comparison to the non-irradiated diabetic group. Quantitative analysis of bFGF expression at 36 h post-injury revealed a threefold increase in the diabetic and non-diabetic Sand Rats after PBM. CONCLUSIONS: The results demonstrate that PBM at an energy density of 4 J/cm(2) is effective in improving the healing of cutaneous wounds in an animal model of type II diabetes, suggesting that PBM (632 nm, 4 J/cm(2)) would be effective in treating chronic cutaneous wounds in diabetic patients.

Dual-confocal fiber-optic method for absolute measurement of refractive index and thickness of optically transparent media.

We present a novel noncontact optical method for absolute measurement of refractive index and thickness of optically transparent media. The method is based on a simple dual-confocal fiber-optic sensor design. It includes two independent confocal channels consisting of two identical apertureless fiber-optic-type confocal microscopes constructed by use of a single 2x2 fiber coupler. A geometrical-ray model is used to obtain the analytical dependence between the sample's refractive index and its thickness. The measurement method provides high accuracy in spatially locating the specific imaging points that correspond to the backreflected intensity peaks of the confocal responses. Thus, a simultaneous measurement of the sample refractive index and thickness is achieved.