Collagen FibrillogenesisCollagen Fibrillogenesis

Mary Beth BirdMary Beth BirdMentor: Ramana Pidaparti, PhD. Mentor: Ramana Pidaparti, PhD.

Mechanical Engineering DepartmentMechanical Engineering DepartmentJuly 23, 2007July 23, 2007

Using Voltage to Optimize Using Voltage to Optimize Protein GrowthProtein Growth

Goals of ProjectGoals of Project To trigger self-assembly mechanisms in collagenTo trigger self-assembly mechanisms in collagen To establish an experimental setup that will To establish an experimental setup that will

support fibrillogenesissupport fibrillogenesis To monitor and alter variables in experiment to To monitor and alter variables in experiment to

determine the best environmental conditions for determine the best environmental conditions for fibrillogenesis fibrillogenesis

To replicate experimental data in computational To replicate experimental data in computational modeling derived from research datamodeling derived from research data

Background on CollagenBackground on Collagen Most common form of protein in the human bodyMost common form of protein in the human body Triple helical molecule consisting of 3 alpha Triple helical molecule consisting of 3 alpha

chains of amino acidschains of amino acids There are 13 different kinds of collagen, Type I – There are 13 different kinds of collagen, Type I –

XIIIXIII Cross-linking during fibrillogenesis causes Cross-linking during fibrillogenesis causes

unique properties amongst the different typesunique properties amongst the different types Collagen can be found in all connective tissues; Collagen can be found in all connective tissues;

ie. bones, tissues, cartilage, ….ie. bones, tissues, cartilage, ….

The Original IdeaThe Original Idea Use metal substrate as a voltage conductor Use metal substrate as a voltage conductor

coated with SDS (Sodium Dodecylsulfate)coated with SDS (Sodium Dodecylsulfate) Layer with a 1-2cm sq of collagen in an acidic Layer with a 1-2cm sq of collagen in an acidic

solution solution Apply voltage across metal conductorApply voltage across metal conductor Monitor growth formation using electron Monitor growth formation using electron

microscopemicroscope Variables: pH, time, voltage, concentration of Variables: pH, time, voltage, concentration of

collagen solutioncollagen solution

Problems EncounteredProblems Encountered Type of substrateType of substrate Determining collagen Type and form to be usedDetermining collagen Type and form to be used Use of SDS (Sodium Dodecylsulfate, ionic surfactant)Use of SDS (Sodium Dodecylsulfate, ionic surfactant) Use of Hydroxylapatite (mineral)Use of Hydroxylapatite (mineral) Application of voltage using Corrosion Data ProgramApplication of voltage using Corrosion Data Program Physical experimental setupPhysical experimental setup Ability to monitor pH and temperatureAbility to monitor pH and temperature Method of examining collagen formationMethod of examining collagen formation Method of quantifying and understanding dataMethod of quantifying and understanding data Ability to confirm presence of fibrilsAbility to confirm presence of fibrils Prevention of corrosionPrevention of corrosion

Images from the LabImages from the Lab

ConcernsConcerns CorrosionCorrosion

How can we prevent corrosion?How can we prevent corrosion? Alter voltage? Alter pH using NaOH? Alter substrate?Alter voltage? Alter pH using NaOH? Alter substrate?

Substrate SurfaceSubstrate Surface How can we improve the surface of the substrate to How can we improve the surface of the substrate to

prevent false images?prevent false images? Analyzing ImagesAnalyzing Images

How do we confirm presence of fibrils?How do we confirm presence of fibrils? What is the best way to take images?What is the best way to take images? SEM, AFM, Optical MicroscopeSEM, AFM, Optical Microscope

Future of the ProjectFuture of the ProjectContinue to refine methods to decrease Continue to refine methods to decrease

the number of experimental errorsthe number of experimental errorsControl and alter experimental variables Control and alter experimental variables

ie. pH, temperature, voltageie. pH, temperature, voltageForm a better method to quantify and Form a better method to quantify and

examine dataexamine dataExpand project to include DNAExpand project to include DNA

AcknowledgementsAcknowledgementsProject Team Members: Project Team Members: - Sumeet Lall, HSURPSumeet Lall, HSURP- Dr. Ramana PidapartiDr. Ramana PidapartiMechanical Engineering Dept., VCUMechanical Engineering Dept., VCU

- Dr. Gary BowlinDr. Gary Bowlin Biomedical Engineering Dept., VCUBiomedical Engineering Dept., VCU- Dr. Peter C. MoonDr. Peter C. Moon School of Dentistry, VCUSchool of Dentistry, VCU- Eddie McCumiskey, Grad Student NanoMan LabEddie McCumiskey, Grad Student NanoMan LabMechanical Engineering Dept., VCUMechanical Engineering Dept., VCU- Dr. Stephen Fong’s LabDr. Stephen Fong’s LabChemical and Life Sciences Engineering Dept., VCUChemical and Life Sciences Engineering Dept., VCU

ReferencesReferences1. Encyclopedia of Human Biology. 1. Encyclopedia of Human Biology. 2nd ed. San Diego, CA: 2nd ed. San Diego, CA:

Academic Press; 1997. Academic Press; 1997.

2. Du C, Cui F, Zhang W, Feng Q, Zhu X, de Groot K. 2. Du C, Cui F, Zhang W, Feng Q, Zhu X, de Groot K. Formation of calcium phosphate/collagen composites Formation of calcium phosphate/collagen composites through mineralization of collagen matrix. through mineralization of collagen matrix. Journal of Journal of Biomedical Materials ResearchBiomedical Materials Research [serial online]. [serial online]. 2000;50:May 31, 2007. 2000;50:May 31, 2007.

3. Sini P, Denti A, Tira M, Balduini C. Role of decorin on 3. Sini P, Denti A, Tira M, Balduini C. Role of decorin on in in vitro vitro fibrillogenesis of type I collagen. fibrillogenesis of type I collagen. Glycoconjugate Glycoconjugate JournalJournal. 1997;14:871-874. . 1997;14:871-874.