INVESTIGATING EFFECTS OF IGF-1 CISSUS QUADRANGULARIS AND FAGUS CRENATA ON THE PRODUCTION OF COLLAGEN IN MOUSE FIBROBLAST CELLS

Done by:Cheng Yong Jian Er Yuan ZhiAllison MeadowsGabi Guzman

CONTENTS Introduction Rational Objectives Hypothesis Materials Methods Timeline References

INTRODUCTIONTendinosis is a chronic form of tendinitis which occurs when mostly type-1 collagen that composes the tendon degenerates through repetitive motion and ageing (Summers, 2003)

Current treatments for tendon repair often contain compounds that stimulate collagen production (Haukipuro, 1991)

INTRODUCTIONCurrent treatments for tendon repair IGF-1 is a growth hormone that has been shown to be an in-vitro stimulant of type-1 collagen synthesis (Ivarsson, 1998)

C.Quadrangularis is said to cause an increase in collagen production and aid the healing of joint ailments (Jainu, M. and Mohan, KV., 2008)

INTRODUCTIONFagus Crenata is known to exhibits a superior effect of prevention of aging by promoting the production of collagen in the dermal fibroblast cells. (Y., Nakayama, N., Kojima, 1998)

Studies have also shown that F.Crenata have the effect of strongly promoting dermal fibroblast collagen biosynthesis (Takada et al, 2007)

RATIONAL We aim to investigate the effectiveness of

Cissus quadrangularis, IGF-I and Fagus crenata to stimulate the synthesis of collagen in mouse tendon fibroblast cells to develop new treatments for tendonitis.

OBJECTIVES Determine the effectiveness of IGF-1,

Cissus quadrangularis and Fagus Crenata on the stimulation of synthesis of collagen in mouse tendon fibroblast cells

HYPOTHESIS Our hypothesis is that:

IGF-1 increase production of collagen synthesis in mouse tendon fibroblast cells

and

Cissus quadrangularis and Fagus crenata increase production of collagen synthesis in mouse tendon fibroblast cells when used both separately and together with IGF-1

VARIABLE

•Amount and type of fibroblast cells treated•Environmental conditionsControlled

Variable•Treatment of fibroblasts cells•Type of herbs used (For AOS and HCI)•Concentration of substances added to

fibroblast cells

Independent Variable

•Amount of collagen produced by fibroblast cellsDependent

Variable

MATERIALS 3T6 Swiss Albino Nuclear Lystate Cissus quandrangularis Fagus Crenata IGF-1 Tissue Culture flask – 25cm2

CO2 incubator Dulbecco’s Modified Eagle Medium Variable volume micropipettes and pipette tips Vortex mixer Centrifuge machine Spectrophotometer Masson Trichrome staining kit Sircol Assay Kit

METHODSTests will be carried out in vitro with IGF-1 and C.quadrangularis (F.crenata for HCI) in DEM medium at different concentrations

Fibroblast cells would be separately treated with IGF-1, C.quadrangularis (F.crenata for HCI) and a combination of both. For the control, it will be left to grow in normal conditions

Mouse fibroblast cells will be cultured in a CO2 chamber at 37°C at a volume of 0.25ml/cm2 for a period of 5 days

METHODS

Masson Trichrome staining will be done on the cells and ImageJ will be used measure the stained collagen

Sircol collagen assay will be done to determine the exact concentration of the amount of collagen produced by the mouse fibroblast cells

Triplicates will be carried out and the results obtained would be compared with the control

TIMELINEDate AOS HCINov-Feb Determine the

concentration of substances to be experimented on

Read up more articles regarding

project

Feb-March Experiments in progress-Obtain 1st set of data

April-May Experiments in progress-Obtain 2nd set of data

June-July Experiments completed-Obtain 3rd set of data

9th July Experiments in progress

Preparation of Project’s Day Semi-

Finals21st August Preparation for Project’s Day Finals

presentation

REFERENCES Cissus Quadrangularis—The Best Kept Secret in

Bodybuilding. (2007). Retrieved September 28, 2009, from USPLabs Web site: http://www.usplabsdirect.com/supercissus.html

Erickson, L. (2002). Future treatments. Retrieved May 21, 2009, from http://www.tendinosis.org/future.html

Erickson, L. (2002). The Tendinosis Injury. Retrieved September 24, 2009, from http://www.tendinosis.org/injury.html

Takda, K., Suzuki, R., Guentert, M., Inomata, S., Hamada, C., and Sakiguchi, T., (June 14 2007) Anti-Aging composition and collagen production promoting composition

REFERENCES Borg, T. K., Ivarsson, M., McWhirter, A., Rubin, K.

(1998). Type I collagen synthesis in cultured human fibroblasts regulation by cell spreading, platelet-derived growth factor and interactions with collagen fibers. Matrix Biology. 16, 409-425.

Daha, M. R., Geest, R. N., Lam, S., VanKooten, C., Verhagen, N. A. M. (2004). Secretion of collagen type IV by human renal fibroblasts is increased by high glucose via a TGF-B-independent pathway. Nephrol Dial Transplant, 19, 1694-1701.

Leeson, C. R., Leeson, T. S., & Paparo, A. A. (1985). Connective tissue proper. Textbook of Histology. Philidelphia: W. B. Saunders Company.

REFERENCES Ghahary, A. , Houle , Kilani, T. , Scott, G. , Shen

and Tredget, E. , (2000), Vol. 17, No. 3, Pages 167-176. Mannose-6-Phosphate/IGF-II Receptors Mediate the Effects of IGF-1-Induced Latent Transforming Growth Factor β1 on Expression of Type I Collagen and Collagenase in Dermal Fibroblasts

Baroni, G., Benedetti, A., Casini, A., Folli, F., Gaglotti, G., Macarri, G., Marucci, L., Orlandoni, P., Perego, L., Ridolfi, F. and Sario, A. (1999) Insulin and Insulin-Like Growth Factor-1 Stimulate Proliferation and Type I Collagen Accumulation by Human Hepatic Stellate Cells: Differential Effects on Signal Transduction Pathways

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