The study aimed to test the effects of commercial deer velvet antler (DVA) and insulin-like growth factor 1 (IGF-1) on cell proliferation using 3T3 mouse fibroblast cells. Scratch assays were performed with DVA, IGF-1, and their combination in media with and without serum. The assays found increased cell proliferation at higher concentrations of the substances. However, results from follow-up MTT assays to confirm the effects were inconclusive. Further scratch assays and MTT assays are needed to fully support the hypothesis that DVA enhances cell proliferation similarly to IGF-1.

1. Methods
Abstract
The Effects of Commercialized Performance Enhancing Deer Velvet Antler on
Cell Proliferation in 3T3 Mouse Fibroblasts
Emily Eichelberger, Chelsea Easter, Sydney Shearer, Timothy Shoff, and Irene M. Wolf, PhD
Department of Biology, Saint Francis University, Loretto, Pennsylvania 15940
Many athletes seek supplements that accelerate
the recovery process, such as insulin-like growth
factor 1 (IGF-1). In humans, IGF-1 is stimulated by
growth hormones and synthesized primarily by the
liver, but also produced by bone and muscle tissue. It
is important in skeletal growth by inducing
chondrocyte proliferation and has effects on
connective tissue proliferation. IGF-1 is an anabolic
steroid and banned by the World Anti-Doping Agency
(WADA), yet it is still used by many athletes.
Commercial companies claim that a product created
from Deer Velvet Antler (DVA) mimics IGF-1.
According to an article on BornFitness.com,
individuals aiming for higher performance and
accelerated recovery rates have begun using DVA.
However, very little laboratory research has been
conducted to support these claims. We hypothesized
that IGF-1 and commercialized DVA will enhance cell
proliferation in 3T3 Mouse fibroblast cells. Data
utilizing scratch assays reveal an increase in cell
proliferation when treating the cells with
commercialized DVA and DVA plus synthetic IGF-1
in serum. Interestingly, the data indicates that smaller
concentrations of DVA induces greater cell
proliferation. Conversely, all three drugs show
increased cell growth when grown in media without
serum. Future studies will be conducted using an MTT
assay to confirm the effects of DVA on cell
proliferation.
3T3 mouse fibroblast cells were housed in an
incubator held at 37°C and 5% CO2. The cells were
grown in Dulbecco’s Modified Eagle Medium
(DMEM) with high glucose, 10% bovine calf serum
(BCS), and 1% penicillin/streptomycin. The cells were
plated onto 35 mm petri dishes at 6x104 cells/mL with
serum. After 24 hours, the media was replaced with
media containing serum (BCS) in trial 1 and without
serum in trial 2. The desired concentrations of drugs
were added and the cells were then scratched with a
pipet tip. A picture was taken with an inverted
microscope immediately following the scratch (0 hrs),
the scratch was measured and the cells were placed in
the incubator for 24 hours. Pictures and measurements
were also taken at 24 and 48 hours.
The results show an increase in cell
proliferation when looking at human and mouse
IGF-1, DVA only, and DVA plus synthetic IGF-1.
In the scratch assay trials using human IGF-1 and
media with serum (BCS), both human IGF-1 and
DVA/IGF-1 showed higher cell proliferation rates
at higher concentrations. Interestingly, the DVA
induced higher growth rates at lower
concentrations. In the trials using mouse IGF-1 and
media without serum (BCS), all three drugs showed
increased cell proliferation rates given at higher
concentrations. The results from the MTT Assay
are inconclusive and need to be retested in other
trials. Overall, more scratch assay trials and MTT
Assays need to be performed to further support our
hypothesis. An AKT Assay will also be ran to
determine if DVA follows the same pathway as
IGF-1 to induce cell proliferation.
- Department of Biology at Saint Francis
University
- TriBeta National Biology Honors Society Grant
- Saint Francis University School of Sciences
Undergraduate Research Grant
Methods
Approximately 48 hours prior to treatment, the 3T3 cells were cultured in a 96-well plate at 5x104 cells/mL in a
37°C incubator and in 5% CO2. Different concentrations of IGF-1, DVA, and DVA + IGF-1 were used to treat the cells,
using DMEM with no serum. Treatments were in triplicate, and repeated three times for n=3. The plate was then
incubated for 48 hours. Following the 48 hours, media was refreshed and the drugs were added. The plate was then
placed back into the incubator. After 24 hours, an MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide)
assay was performed in order to evaluate and determine cell proliferation activity. 10 microliters of MTT reagent was
added to each well plate and was placed back in the incubator to allow the reaction to occur. Contents of the wells were
removed and 100 microliters of Crystal Dissolving Solution was added to each well. The final absorbance levels were
measured at 570 nm using a spectrophotometer. All results will be subjected to statistical analysis.
Figure 2: MTT Assay A) DVA, B) DVA/IGF-1,
C) Mouse IGF-1. Darker colors indicate
higher levels of cell proliferation. Mouse IGF-
1 show higher growth rates at higher
concentrations. The results of this assay were
inconclusive.
Figure 1: Human IGF-1 Scratch Assay containing Serum.
Conclusion
Acknowledgements
Results
Graph 1: Scratch Assay with Serum (Trial 1) Scratch
Assay reveals increased cell proliferation at higher
concentrations with Human IGF-1 and DVA/IGF-1
plus.
Graph 2: Scratch Assay without Serum (Trial 2) Scratch
Assay reveals increased cell proliferation at higher
concentrations with Mouse IGF-1, DVA, and
DVA/IGF-1 Plus.
C
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