Here's a more indepth article, with reknowned expert, Dr. Tiina Karu.
I dug this up 2 years ago, the format may not be as tidy as I'd like, but here it is:Prof. Tiina Karu's
Cellular Mechanisms of Low-Power Laser Therapy
) 1. What is photobiomodulation (low-power laser therapy?)
More than 30 year ago the first publications about low-power laser therapy or
photobiomodulation (at that time called laser biostimulation) appeared. Since then
approximately 2000 studies have been published on this topic (analysis of these publications
can be found in ). Medical treatment with coherent light sources (lasers) or noncoherent
light (Light Emitting Diodes, LED's) has passed through its childhood and early maturity.
Photobiomodulation is being used by physiotherapists (to treat a wide variety of acute and
chronic muscosceletal aches and pains), dentists (to treat inflamed oral tissues, and to heal
diverse ulcerations), dermatologists (to treat oedema, indolent ulcers, burns, dermatitis),
rheumatologists (relief of pain, treatment of chronic inflammations and autoimmune diseases),
and by other specialists (e.g., for treatment of middle and inner ear diseases, nerve
regeneration). Photobiomodulation is also used in veterinary medicine (especially in
racehorse training centers) and in sports medicine and rehabilitation clinics (to reduce swelling
and hematoma, relief of pain and improvement of mobility and for treatment of acute soft
tissue injuries). Lasers and LED's are applied directly to respective areas (e.g., wounds, sites
of injuries) or to various points on the body (acupuncture points, muscle trigger points). For
details of clinical applications and techniques used, the books [ 1-3] are recommended. 2. What light sources (lasers, LED's) can be used?
The field of photobiomodulation is characterized by variety of methodologies and use of
various light sources (lasers, LED's) with different parameters (wavelength, output power,
continuous wave or pulsed operation modes, pulse parameters). These parameters are
usually given in manufacturers manuals.
The GaAlAs diodes are used both in diode lasers and LED's, the difference is whether the
device contains the resonator (as the laser does) or not (LED). In latter years, longer
wavelengths (-800-900 nm) and higher output powers (to 100 mW) are preferred in
therapeutic devices. Should a medical doctor use a laser or a diode?
The answer is - it depends on what one
irradiates, in other words, how deep tissue layers must be irradiated. By light interaction with a
biotissue, coherent properties of laser light are not manifested at the molecular level. The
absorption of low-intensity laser light by biological systems is of a purely noncoherent (i.e.,
photobiological) nature. On the cellular level, the biological responses are determined by
absorption of light with photoacceptor molecules (see the section 3 below). Coherent
properties of laser light are not important when cellular monolayers, thin layers of cell
suspension as well as thin layers of tissue surface are irradiated (Fig. 1). In these cases, the coherent and noncoherent light (i.e., both lasers and LED's) with the same wavelength,
intensity and dose provides the same biological response.
Some additional (therapeutical)
effects from the coherent and polarized radiation (lasers) can occur in deeper layers of bulk
tissue only and they are connected with random interference of light waves. An interested
reader is guided to the ref.  for more details. Here we illustrate this situation by Fig. 1.
Large volumes of tissue can be irradiated by laser sources only because the length of
longitudinal coherence Lcoh is too small for noncoherent radiation sources . 3. Enhancement of cellular metabolism via activation of respiratory chain: a universal
photobiological action mechanism
A photobiological reaction involves the absorption of a specific wavelength of light by the
functioning photoacceptor molecule. The photobiological nature of photobiomodulation
means that some molecule (photoacceptor) must first absorb the light used for the irradiation.
After promotion of electronically excited states, primary molecular processes from these states
can lead to a measurable biological effect (via secondary biochemical reaction, or
photosignal transduction cascade, or cellular signaling) at the cellular level. The question
is, which molecule is the photoacceptor.
Fig. 1. Depth (On in which the beam coherency is manifested, and coherence length Lcoh in
various irradiated systems: (A) monolayer of cells, (B) optically thin suspension of cells, (C)
surface layer of tissue and bulk tissue. Lcoh, - length of temporal (longitudinal) coherence of
laser light, hw) marks the radiation.
When considering the cellular effects, this question can be answered by action spectra. Any
graph representing a photoresponse as a function of wavelength, wave number, frequency,
or photon energy, is called action spectrum. Action spectra have a highest importance for
identifying the photoacceptor inasmuch as the action spectrum of a biological response
resembles the absorption spectrum of the photoacceptor molecule. Existence of a structured
action spectrum is strong evidence that the phenomenon under study is a photobiological
one (i.e., primary photoacceptors and cellular signaling pathways exist). Fig. 2 represents
some examples of action spectra for eukaryotic cells: two of them (A, B) consider the
processes occurring in cell nucleus, and one spectrum (C) is for cell membrane. Fig. 2D shows
the absorption spectrum of the monolayer of the same cells.
Fig. 2. Action spectra of: (A) DNA and (B) RNA synthesis rate in HeLa cells; (C) plasma
membrane adhesion of HeLa cells for red-to-near IR radiation; (D) absorption spectrum of air-
dried monolayer of HeLa cells for the same spectral region. Original data can be found in ref. .
The spectra in Fig. 2 represent the red-to-near infrared (IR) region only, i.e. the region that is
most important for photobiomodulation. The action spectra for full visibleto-near IR region can
be found in . In  one can find action spectra for various cellular responses for other
eukaryotic and prokaryotic cells as well.
Two conclusions can be drawn from action spectra in Fig. 2. First, the similarity of the action
spectra for different cellular responses suggests that the primary photoacceptor is the same
for all these responses. Second, the existence of the action spectra for biochemical
processes occurring in various cellular organelles (nucleus, Fig. 2A, B and plasma membrane,
Fig. 2C) assume the existence of cellular signaling pathways inside of a cell between the
photoacceptor and the nucleus as well as between the photoacceptor and cell membrane.
Action spectra also indicate, which wavelengths are the best for irradiation: maximal biological
responses are occurring when irradiated at 620, 680, 760 and 820-830 nm (maxima of the
spectra in Fig. 2). Skipping over the story of identifying the photoacceptor (described in )
let us conclude that photoacceptor for eukaryotic cells in red-to-near IR region is believed to
be the terminal enzyme of the respiratory chain cytochrome c oxidase (located in cell
mitochondrion). To be more exact, it is a mixed valence (partially reduced) form of this
enzyme, which has not yet been identified. In the violet-to-blue spectral region, flavoproteins
(e.g., NADHdehydrogenase in the beginning of the respiratory chain) are also among the
photoacceptors as well terminal oxidases. An important point has to be emphasized
. When the excitable cells (e.g., neurons,
cardiomyocites) are irradiated with monochromatic visible light, the photoacceptors are also
believed to be components of respiratory chain. Some of the experimental evidence
concerning excitable cells is shortly summarized in Fig. 3. It is quite clear from experimental
data (reviewed in ) that irradiation can cause physiological and morphological changes in
nonpigmental excitable cells via absorption in mitochondria. Later, similar irradiation
experiments were performed with neurons in connection with low-power laser therapy. It was
shown in 80's that He-Ne laser radiation alters the firing pattern of nerves; it was also found
that transcutaneous irradiation with HeNe laser mimicked the effect of peripheral stimulation
of a behavioral reflex. These findings were found to be connected with pain therapy (review ) So, what happens when the molecule of photoacceptor absorbs photons?
Answer - electronic excitation followed by photochemical reactions occurring from lower excitation
states (first singlet and triplet). It is also known that electronic excitation of absorbing centers
alters their redox properties. Until yet, five primary reactions have been discussed in literature
(Fig. 4). Two of them are connected with alteration of redox properties and two mechanisms
involve generation of reactive oxygen species (ROE). Also, induction of local transient (very
short time) heating of absorbing chromophors is possible. Details of these mechanisms can
be found in [4, 5].
There is no ground to believe that only one of the reactions shown in Fig. 4 occurs when a
cell is irradiated and excited electronic states are promoted. The question is, which
mechanism is decisive. It is not excluded that all mechanisms shown in Fig. 4 lead to a similar
result, to a modulation of redox state of the mitochondria (a shift to more oxidized direction).
However, depending on the light dose and intensity used, some mechanism(s) can prevail
The next question is, the following if photoacceptors are located in the mitochondria, then
how the primary reactions occurring under irradiation in the respiratory chain (Fig. 4) are
connected with DNA and RNA synthesis in the nucleus (the action spectra in Fig. 2A, B) or
with changes in plasma membrane (Fig. 2C)? The principal answer is that between these
events there are secondary (dark) reactions (cellular signaling cascades or photosignal
transduction and amplification, Fig. 5).
Three regulation pathways are suggested in Fig. 4. The first one is the control of
photoacceptor over the level of intracellular ATP. It is known tat even small changes in ATP
level can alter cellular metabolism significantly. This regulation way is especially important by
irradiation of hypoxic, starving or otherways stressed cells. However, in many cases the
regulative role of redox homeostasis is proved to be more important than that of ATP. For
example, it is known that the susceptibility of cells to hypoxic injury depends more on the
capacity of cells to maintain the redox homeostasis and less on their capacity to maintain the
The second and third regulation pathways are mediated through the cellular redox state (Eh;
Fig. 4). This way involve redox-sensitive transcription factors (NF-KB and AP1, Fig. 4) or
cellular signaling homeostatic cascades from cytoplasma via cells membrane to the nucleus
(Fig. 4). As a whole, the scheme in Fig. 4 suggests a shift in overcell redox potential into more
oxidized direction. Modulation of cellular redox state affects gene expression namely via
transcription factors. It is important that in spite of some similar or even identical steps in
cellular signaling, the final cellular responses to the irradiation differ due to existence of
different modes of regulation of transcription factors. The mechanisms of regulation are not
understood well yet.
The magnitude of cellular responses depends on cellular redox potential (and its
physiological status, respectively) at the moment of irradiation. The cellular response is
stronger when the redox potential of the target cell is initially shifted to a more reduced state
(and intracellular pH, pH;, is lowered, as usually happens in injured cells). This explains why
the degrees of cellular responses can differ markedly in different experiments or in different
clinical cases, and why the effects are sometimes nonexistent.
One should emphasize that some biological limitations exist for photobiomodulation effects.
These are discussed in .4. Enhancement of cellular metabolism via activation of nonmitochondrial
photoacceptors. Indirect activation/suppression
The redox regulation mechanism cannot occur solely via respiratory chain (Section 3). Other
redox chains containing molecules, which absorb light in visible-to-near IR radiation, and are
some key structures that can regulate a metabolic pathway, can be photoacceptors for
photobiomodulation as well. One such example is NADPH-oxidase of phagocytic cells, which
is responsible for nonmitochondrial respiratory burst. This multicomponent enzyme system
located in the plasma membrane is a redox chain that generates reactive oxygen species
(ROS) as a response to the microbicidal or other types of activation. Irradiation with He-Ne
laser and diode lasers and LED's can activate this chain in various phagocytic cells. Many
worked examples can be found in . In phagocytes, the activation of respiratory chains in
mitochondria occurs as well, as NADHP-oxidase activation, but the latter is much stronger.
ROS, burst of which is induced by direct irradiation of phagocytes, can activate or inactivate
other cells, which were not irradiated directly. In this way, indirect activation or suppression of
metabolic pathways in non-irradiated cells occurs. Also, lymphokines and cytokines produced
by irradiated lymphocytes can influence metabolism of other cells. This situation is common by
irradiation on tissues. 5. Concluding Remarks
The photobiological action mechanism via activation of respiratory chain is a universal working
mechanism for various cells. Crucial events of this type of cell metabolism activation are
occurring due to a shift of cellular redox potential into more oxidized direction as well as due
to ATP extrasynthesis. Susceptibility to irradiation and capability for activation depend on
physiological status of irradiated cells: the cells, which overall redox potential is shifted to
more reduced state (example: some pathological conditions) are more sensitive to the
irradiation. The specificity of final photobiological response is determined not at the level of
primary reactions in the respiratory chain but at the transcription level during cellular signaling
cascades. In some cells, only partial activation of cell metabolism happens by this mechanism
(example: redox priming of lymphocytes).
All light-induced biological effects depend on the parameters of the irradiation (wavelength, Cited Literature
dose, intensity, irradiation time, and continuous wave or pulsed mode, pulse parameters).
According to action spectra, optimal wavelengths are 820-830, 760, 680, and 620 nn. Large
volumes and deeper layers of tissues can successfully irradiated by laser only (e.g. inner and
middle ear diseases, injured siatic or optical nerves, deep inflammations etc.). The LED's are
excellent for irradiation of surface injuries.
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4. Karu, T.I. (2002). Low power laser therapy. In: CRC Biomedical Photonics Handbook, T.
Vo-Dinh, Editor- in-Chief, CRC Press, Boca Raton (USA).
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