Effect of Pulse-Burst Electromagnetic Field Stimulation on Osteoblast Cell Activities
Walter Hong-Shong Chang, Li-Ting Chen, Jui-Sheng Sun, and Feng-Huei Lin
Bioelectromagnetics 25:457-465, 2004.
SUMMARY:
The use of electrical stimulation for bone healing in modern medicine dates back to the 1950s and has since seen much clinical success, although the mechanism has yet to be understood. This paper assesses the stimulatory effect of pulsed electromagnetic field (PEMF) on bone cells and suggests a possible mechanism of this effect.
Osteoblast-like cells were harvested from newborn mice, cultured to confluence, and passaged once. The cultures were divided into "control" and "treated" 6-wells plates. Previous studies showed that bone cells grown in 1% calf serum medium displayed significant response to PEMF stimulation. Therefore, the harvested cells were initially grown in 10% calf serum, to facilitate attachment of osteoblasts, and then changed to 1% medium. The PEMF chamber was program to produce a burst frequency of 15 Hz and a magnetic field strength of 1G. "Treated" plates were subjected to the conditions in the PEMF chamber for 8 hrs a day. At 1, 3, 5 and 12 days after beginning PEMF stimulation, temporal effects on the cells were evaluated using various assays.
Parameters evaluated were cell proliferation, differentiation, mineralization, and gene expression. Cell proliferation were measured by mitochondria activity of bone cells using calorimetric assay which detects conversion of 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide. Differentiation was determined by the release of alkaline phosphatase (ALP) and lactate dehydrogenase (LDH), both of which were detected using available assay kit and ALP staining. Mineralization of osteo-nodules were meausred using von-Kossa staining. Gene expression analysis was done by isolating cell RNA, reverse transcription into cDNA, PCR amplification, and agarose electrophoresis.
Results showed increase proliferation in the PEMF treated culture by 34, 11.5, 13.3% after 3, 5, and 7 days of culture, relative to control culture. No LDH were detected in both treated and control cultures but ALP concentration decreased 11.4, 20.1, and 32.0% over 3, 5, and7 days of culture, relative to control. There was no difference in matrix mineralization in control and treated groups. Relative to the control, PEMF treated culture showed upregulation of osteoprotegrin mRNA, and downregulation of the receptor activator of NF-kB ligand (RANKL) mRNA.
From the results, it was infer that PEMF unequivocally increase the proliferation of bone cells. Yet, a decrease in ALP, an early differentiation marker, shows that PEMF didn't effect cell differentiation. The paper explained that differentiation and mineralization was delayed to make way for the increase proliferation. The upgregulation of osteoprotegrin and downregulation of RANKL, proteins involved in osteoblast growth cycle, may reveal that PEMF may have inhibitory effect on osteoblast maturation. It was concluded that PEMF have positive effect on proliferation but have no effect on cellular differentiation, maturation, and mineralization.
RELEVANCE
The movement of medicine has been to be as non-invasive as possible. Electrical stimulation has potential to be "blood-less" and possibly painless treatment. When it comes bone healing, the current process is slow: minor fractures and injuries can take weeks to months to heal. This paper shows that PEMF can expedite the healing process by increasing osteoblast proliferation without use of any extrinsic hormones. Additionally, this paper employs many assays, some of which we've used, that are educational and possibly useful in our 115 project.
7 comments:
I agree, non-invasive treatments are definitely becoming more prevalent. This sounds like promising bone regeneration therapy. Do you think the PEMF treatment would be useful even though it may stunt differentiation? How important is it for osteoblasts to be properly differentiated in bone regeneration? I imagine that there are a lot of signals which influence differentiation that are sent to bones regenerating in vivo. Were any in vivo studies done?
You mentioned that the osteoblasts may not have had time to differentiate due to their rapid proliferation. Did the researchers do any longer studies in which osteoblasts were given enough time after the PEMF treatment to differentiate?
Bone regeneration therapy sounds like a great idea with natural healing times taking weeks to months. I see that they were able to get cells that underwent increased proliferation, but were the reasearchers able to get a better idea as to why PEMF has this effect on the osteoblasts?
Also, is this increased proliferation due to PEMF limited to osteoblasts, specifically were any tests done to ensure that it will not have any adverse affects to the surrounding cells? Finally, could you also explain how the von-Kossa staining works for testing for mineralization.
I don't know much about the clinical use of PEMF, but do they really expose a patient's arm or leg to electromagnetic fields for 8hrs every day? Or did the researchers pick 8hrs as a maximum limit? I've read some papers in bone regeneration and they almost unequivocally say that growing bone quickly is not the same as growing bone well. In light of that, do you know if clinicians still use PEMF? It seems better to just functionalize an implant with BMPs (bone morphogenetic proteins). But you are right; the movement towards non-invasive medicine is important and this is a great example of that.
You said that mineralization in both the treated and control cells were the same. But, if the treated cells had less differentiation due to the PEMF, shouldn’t the control cells have more mineralization, because they were allowed to differentiate more and therefore allow for more mineralization?
And, do you think that we can use PEMF, if it is proven to be a working non-invasive procedure, on the people who have a decrease in osteoblast cells, such as the elderly, as to increase the number of osteoblast cells and therefore increase bone regeneration? What complications do you think could arise if this is used throughout the body to increase osteoblast proliferation in all bones?
If PEMF is used to expedite the healing process in a broken bone, what control mechanisms could you put in place to ensure that 1. only the healing region experiences osteoblast proliferation and 2. the proliferation occurs in a controlled manner? If a patient undergoes this therapy, will cells other than the targeted osteoblasts also experience an increased proliferation rate, or even some other side effect? Are the benefits of having a rapid proliferation greater than the possible downsides of the osteoblasts not differentiating at the expected rate? It seems that if you grow a lot of immature cells they may not react correctly in a manner that will speed up natural healing.
Cool stuff!
The fact that pulsed electromagnetic field can be used to heal the bone is new to me. I usually avoid magnetic field coming out from the microwave because I heard it’s bad for my brain. Next time, I might try to put my wrist in front of the microwave so I can heal some of my bone (I have a wrist problem and I’m not kidding!)
Does it have to be “pulsed” though? I might want to see the effect of magnetic field of microwave on the osteoblast cell proliferation and/or MCF-7 cells that we used for our project. This can be easy but interesting future experiment! Although magnetic field of microwave might too strong and kill all the cells immediately, if we can detect certain effect on MCF-7 cell, we can welcome pain free electrical treatment of breast cancer in the future (or NOT…=P)
Sorry…right now, I just tend to relate everything I read to our project…
We decided our data is “inconclusive” partially due to not enough data to conduct statistical analysis like ANOVA/ t-test because we only run two plates…SD between two number is…does not mean that much.
So, I wonder how many experiments they run to conclude that “PEMF have positive effect on proliferation but have no effect on cellular differentiation, maturation, and mineralization”?
Since right before the conclusion statement, you also mention that “the upgregulation of osteoprotegrin and downregulation of RANKL, proteins involved in osteoblast growth cycle, may reveal that PEMF may have inhibitory effect on osteoblast maturation,” what factors made them to conclude that PEMF have no effect on osteoblast maturation even they suspect it might have inhibitory effect on the maturation?
Thanks~~
I like this idea for stimulation of bone regrowth using a method that is found naturally in human physiology with bursts of electromagnetic waves. Since such signals are already found within the body, any damage to cells should seem to be limited. However, does the paper cover how extra impulses or greater magnitude of the electric signals effects surrounding tissue? Also, what does the paper mean by mineralization. Very interesting!
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