Increasing Bovine Growth Plate Chondrocyte Proliferation Using Capacitive Coupled Electrical Fields (IMPORTANT)

Me: This post will be my way to combine 4 scientific article all written by Dr. Brighton in trying to see if removed bovine growth plate cartilage chondrocytes can be still be made to proliferate. I wanted to see whether the chondrocytes can possibly be regrown back into cartilage through just cartilage proliferation and possibly adding the right amount and type of growth factor concoction. 

Analysis: In a very recent post I had written I had compared Brighton’s two articles which he in 1983 had shown that capacitative couled elctrical fields applied on epiphyseal plates lead to increased longitudinal growth but later in 1986 another article said that the experiment at 5 V peak to peak didn’t lead to any statistical increases. My conclusion for that post was that it was very strange and I couldn’t resolve why the two experiments reuslts seemed to contradict each other. 

Brighton and his collaborators took these chondrocytes from growth plates found in newborn calf costochondral junctions and put them in a monolayer. The monolayer was subjected to a capacitvely coupled electrical field for 2 days and the additon of compounds like thymidine, proline, and phosphate were also added to see what would be the result on these compounds in terms of uptake. The signals which were done for a long time had little effect but the signals that were done only for a short time had the highest effect. From the 2nd article,…

“It is concluded that short-term exposure of growth plate chondrocytes to an appropriate capacitively coupled field stimulates cAMP production, but longer-term application of the electrical field is ineffective.”

This seems to agree with the old articles. in the 2nd study done, those rabbits had their legs subjected to electrical fields constantly for 6 weeks. In the first study, those rabbits only had their plates subjected for 2-4 days.

From the 1st article…”We conclude that the biologic response of cells in vitro is signal specific, and that the total amount of electrical energy required to stimulate the growth plate chondrocyte to increased proliferation is very small since the total time the 0.25% duty cycle signal was only 3.6 min of a 24-h period.”

From the 3rd article, Brighton concludes that…“articular cartilage chondrocytes grown in pellet form can be stimulated to increase glycosaminoglycan synthesis or to increase cell proliferation by an appropriate capacitively coupled electrical field. The importance of the serum concentration in the medium in evaluation of biosynthesis in vitro is noted.” What is very important to note is that the higher voltage did nothing but the lower voltages had a dramatic affect on the thymidine and sulfate uptake 

From the 4th article, it shows that the researchers had taken bovine chondrocytes from growth plates and made pellets from them. They applied the same type of capacitively coupled electrical field and and showed almost the same results as in article 3. It was noted that anything past 30 minutes of application seemed to have a diminishing effect on proliferation. The cAMP levels increased significantly from the 200-1000 V peak to peak signals. 

Here are the big takeaways from it. If you use the electrical field for too long, it ruins the effects. If you use the field at too high of a voltage, it also ruins the effect. You want to keep any application and use of the signal generator at a short and low intensity level for maximum effect.


From source link HERE

J Orthop Res. 1989;7(5):759-65.

Proliferative and synthetic response of bovine growth plate chondrocytes to various capacitively coupled electrical fields.

Brighton CT, Jensen L, Pollack SR, Tolin BS, Clark CC.

Source

McKay Laboratory of Orthopaedic Surgery Research, University of Pennsylvania School of Medicine, Philadelphia 19104.

Abstract

In vitro monolayer cultures of growth plate chondrocytes isolated from newborn calf costochondral junctions were subjected to capacitively coupled electrical fields for 48 h. In part A, the electrical signal was a 60-kHz sine wave applied at different voltages so as to produce electrical fields at the pericellular level of 7, 20, 50, and 126 mV/cm. Incorporations of [3H] thymidine and [35S] sulfate were assayed to determine the effect of the above fields on cells proliferation and matrix synthesis, respectively. Proliferation was increased by 47% in the 20 mV/cm field whereas the same field decreased [35S] sulfate incorporation by 21%. These changes were significant at p less than 0.05 in both instances. In part B, the 20 mV/cm field was applied in a pulsed fashion to produce daily duty cycles of 100, 25, 2, and 0.25%. Incorporation of [3H]thymidine, [35S]sulfate, and [14C]proline per DNA were assayed. Results indicated that the 100, 25, and 0.25% percent duty cycles showed significantly (p less than 0.01-0.05) increased proliferation, whereas the 0.25% signal (5 ms on/495 ms off for 6 h/day) significantly decreased [14C]proline incorporation. We conclude that the biologic response of cells in vitro is signal specific, and that the total amount of electrical energy required to stimulate the growth plate chondrocyte to increased proliferation is very small since the total time the 0.25% duty cycle signal was only 3.6 min of a 24-h period.

PMID: 2760750     [PubMed – indexed for MEDLINE]

From source link 2 HERE
J Orthop Res. 1988;6(4):552-8.

Increased cAMP production after short-term capacitively coupled stimulation in bovine growth plate chondrocytes.

Brighton CT, Townsend PF.

Source

Department of Orthopaedic Surgery, University of Pennsylvania School of Medicine, Philadelphia 19104.

Abstract

Growth plate chondrocytes from newborn calf costochondral junctions grown in monolayer were subjected to a capacitive AC signal of 500 V peak to peak (P-P) at 60 kHz for 48 h and were analyzed for [3H]thymidine uptake. The stimulated chondrocytes showed a 130% greater uptake over unstimulated controls. Other newborn calf growth plate chondrocytes were stimulated at 500 V P-P at 60 kHz for 2.5, 5.0, 10.0, and 20.0 min and were analyzed for cAMP. Chondrocytes stimulated for 2.5 and 5.0 min showed a 142.8% (p less than 0.05) and 394.5% (p less than 0.01) increase over controls, respectively. The chondrocytes stimulated for 10.0 and 20.0 min showed no significant difference from the controls. It is concluded that short-term exposure of growth plate chondrocytes to an appropriate capacitively coupled field stimulates cAMP production, but longer-term application of the electrical field is ineffective.

PMID:  2837556     [PubMed – indexed for MEDLINE]

From source link HERE
J Orthop Res. 1984;2(1):15-22.

In vitro growth of bovine articular cartilage chondrocytes in various capacitively coupled electrical fields.

Brighton CT, Unger AS, Stambough JL.

Abstract

Isolated articular cartilage chondrocytes from 1- to 3-week-old male Holstein calf knee joints were formed into pellets containing 4 X 10(6) isolated cells and were grown in tissue culture medium (minimum essential medium/NCTC 135) containing either 1 or 10% newborn calf serum (NBCS) in plastic Petri dishes in 5% CO2 and air at 37 degrees C in saturation humidity. On the 4th postisolation day either [35S]sulfate or [3H]thymidine was added to the medium, and the pellets were exposed for 24 h to capacitively coupled electrical fields (10, 100, 250, and 1,000 V peak-to-peak, 60 kHz, sine wave signals). The pellets were then harvested, dialyzed, hydrolyzed, and assayed for DNA, protein, [35S]sulfate incorporation, and [3H]thymidine incorporation. Results indicated that at 250 V peak-to-peak there was a statistically significant increase in [35S]sulfate in 1% NBCS and a statistically significant increase in [3H]thymidine in 10% NBCS. At potentials above or below 250 V no changes were noted. Thus, articular cartilage chondrocytes grown in pellet form can be stimulated to increase glycosaminoglycan synthesis or to increase cell proliferation by an appropriate capacitively coupled electrical field. The importance of the serum concentration in the medium in evaluation of biosynthesis in vitro is noted.

PMID: 6491794      [PubMed – indexed for MEDLINE]

From source link HERE
J Orthop Res. 1988;6(2):265-71.

Capacitively coupled electrical stimulation of bovine growth plate chondrocytes grown in pellet form.

Armstrong PF, Brighton CT, Star AM.

Source

Department of Orthopaedic Surgery, University of Pennsylvania School of Medicine, Philadelphia.

Abstract

Pellets formed from isolated bovine growth plate chondrocytes were grown in various capacitively coupled electrical fields. The signals chosen were 0, 10, 100, 250, 500, 750, 1,000, and 1,500 V peak-to-peak, 60 kHz. The effect on cell proliferation and matrix production of these different voltages was determined by [3H]thymidine and [35S]sulfate uptake, respectively, Cyclic AMP assays were done to determine if increases in either thymidine or sulfate uptake were associated with changes in cAMP levels. Significantly increased cell proliferation occurred at 500, 750, and 1,000 V peak to peak. The calculated electric fields were 1.5 to 3.0 x 10(-2) V/cm. Proliferation was significantly inhibited at 1,500 V peak-to-peak with a calculated field of 4.5 x 10(-2) V/cm. Little if any change was seen in cAMP levels at 30 or 60 min following application of the appropriate electric signals.

PMID: 2830391       [PubMed – indexed for MEDLINE]

Leave a Reply

Your email address will not be published. Required fields are marked *