I used Nd;YAG laser with operating in CW and Short pulse laser with1064 nm wavelength on the skin of mouse: By studing the expression of heat shock proteins 70 and 47 the results shown that with CW tissue get more damage however with pulse we found less damage in the laser area. Article Expression of heat shock proteins 70 and 47 in tissues follo...
Now, I am applying the same laser on the fibroblast cells to see the difference effect of lasers on tissue and cell. The method I used is MTT assay which it shown that with Pulse the cell activity is lower than using CW. (The excel file is attached here).
The results for tissue and cell are opposite of each other.
More information:
1- the power for both lasers are around 2W.
2- Irradiation time 1 minute on cells.
3- Cells were plated in 96-well plate (10,000 cells/well)
4- control was non irradiated.
Could you please help me about what is the hypothesis behind this ?
If the "short" pulses were adequately short, the laser energy might be stress-confined, meaning that a majority of the energy from the laser was coupled photomechanically or photoacoustically. In long pulse or CW mode, most laser energy is transferred through photothermal coupling - turning into heat and hence increasing the temperature in the skin.
In the in vivo scenario, when the fibroblasts were located within the dermal matrix, they might have experienced attenuated (weak) acoustic/stress waves from the short laser pulses, while experienced more heat generation (thus temperature increase) in the CW mode - 1064 nm penetrates deep into the skin because of its relatively low absorption coefficient.
When you transfer the experiment to an MTT assay in an in vitro environment, the fibroblasts were now directly being irradiated with the short laser pulses. Mechanical stress induced directly at the surface of the cells as a result of absorption could disrupt the cell membranes resulting in injuries, which could be the reason you see the opposite effects. In fact, cell membrane disruption through photomechanical/photoacoustic mechanism with pico- and nano-second lasers have previously been studied for drug delivery and gene transfection. Mechanical disruption and damage induced by direct irradiation in the MTT assay may outweigh the thermal injury induced by a CW mode laser, hence you noticed an opposite effect compared to in vivo outcome.
Just curious: when you said you noticed higher HSP70 and HSP47 expression and tissue damage in your in vivo study with the laser in CW mode, I assume you were referring to the fibroblasts in the dermis (since HSP 47 is a chaperone for pro-collagen synthesis specific to fibroblasts). Have you looked at HSP70 expression of the keratinocytes in situ, and also run an MTT assay with keratinocytes?
Kin
The influence of laser is very effective paramete in this your field
Laser influence = E / spot area
regards
Raad
At 150 ns and at 1064 nm, wherever there is optical absorption, you would have stress confinement in most biological tissues, where most of the laser energy was absorbed and thermoelastic stress induced before it was able to propagate (relax, as in "acoustic relaxation") beyond the immediate zone of absorption. This could produce local cavitation and shockwaves that damage cells and tissue.
Addendum: However, note that the absorption coefficient at 1064 nm in tissue is relatively low, but scattering is decent (not as high as in the UV-VIS) but still dominated by Mie scattering. So another potential photoacoustic damage could be, if you had focused the laser beam, attributed to optical breakdown within the tissue. That would generate plasma expansion which could also produce cavitation and shockwaves. Believe it or not, sometimes when this happens, you could hear it!
Hi Neda,
In pulsed mode the exposure time was 15s, and in CW was the duration the same?
I recommend Kin F Chen's answers. Also the observations are pretty interesting. My best wishes to Neda Perchami.
- Badges
- Science topic
- Similar topics
- Mathematics