Nanomedicine : Where Have We Been and Where are We GoingWorldwide market for nanoscale devices was 406 million dollars in 2002. 2003. Congressional hearing on societal implications

Nanomedicine :Where Have We Been and Where are We Going ???

Thomas J. Webster, Ph.D.

Editor, International Journal of NanomedicineCo-director, Indo-U.S. Center for Biomaterials for

HealthcareAssociate Professor

Division of Engineering and Department of Orthopaedics

Brown University, Providence, RI

What is Nanomedicine ???

Nanomedicine: The use of materials in medicine whose components exhibit significantly changed properties by gaining control of structures at the atomic, molecular, and supramolecular levels.

1959Richard Feynman describes molecular machine building with atomic precision.

1974Norio Taniguchi uses term "nano-technology" in a paper on ion-sputter machining*.

1990First nanotechnology journal called Nanotechnology published.

1996First business oriented nanobio conference held by International Business Communications “Biological Approaches and Novel Applications for Molecular Nanotechnology” December 9-10, 1996, in San Diego, CA, USA.

2000President Clinton announces U.S. National Nanotechnology Initiative.

*Taniguchi, N (1974), On the Basic Concept of 'NanoTechnology’ Proc. ICPE, 5-10.

History of Nano-Technology

2002 Worldwide market for nanoscale devices was 406 million dollars in 2002.

2003Congressional hearing on societal implications of nanotechnology.

2006Reflecting the world-wide interest in nanomedicine, the first international journal in nanomedicine is created: International Journal of Nanomedicine.

History of Nano-Technology

Nano iPOD

Nanotechnology science fiction ?

Is this nanotechnology ?

• The National Science Foundation forecasts that the global market for nanotechnology-related products and services will reach $1 trillion by 20151.

• Nanomedicine will exhibit strong growth in all sectors until as far out as 2011, leading to multi-billion dollar revenues2.

1http://www.biz-lib.com; 2http://www.piribo.com

Projections for Nano-Technology

Del SolT-shirts

use nanotechnologyto change color

in the sun

InMat LLC uses nanotechnology to increase tennis ball

lifetime

But what about nano-technologyin medicine (nanomedicine) ???

Nanophase Grain Size

Conventional Grain Size

Closer Look at Nano-technologyin Biomedical Applications

6

4

2

0

microns

6 4 2 0microns

01.3

microns

0

6

6 4 20

4

2

0microns

microns

6microns1.3

0

Compared to conventionalmaterials, nanophase materials possess enhanced:

• processing,• catalytic,• optical, • mechanical,• electrical, and• surface

properties that may enhance existing biomedical implantapplications.

T. J. Webster, in Advances in Chemical Engineering Vol. 27, Academic Press, NY, pgs. 125-166, 2001.

Presenter

Presentation Notes

Tom Webster’s research at Purdue focusses on the use of nanophase materials in biomedical applications. Nanophase materials possess grain sizes less than 100 nm; in contrast, traditional or conventional materials are composed of grain size greater than 100 nm. The atomic force micrographs that appear on this page demonstrate differences in grain sizes of conventional versus nanophase titania formulations. By changing the grain size of a material into the nanometer regime, numerous properties of the material change. Enhanced properties of nanophase materials have been investigated for over a decade now, however, full appreciation of these materials in biomedical engineering applications have yet to be realized. Some of the improved properties of nanophase materials include processing conditions, catalytic, mechanical, electrical and surface properties. For example, ceramics may be processed through sintering (or heat treating) at much lower temperatures than larger grain size materials. This means that these materials can be processed for biomedical applications at much less of an expense to implant manufacturer’s. In addition, by decreasing grain sizes of materials, one can obtain significantly higher surface areas. Since many applications concerning drug delivery are dependent on the amount of surface area available for enzymatic activity, an increase in surface area could result in more efficient pharmaceutical drugs.

Objective

The objective of the following studies was to determine whether tissue regeneration is altered on biologically-inspired nano-structured surfaces compared to the nano-smooth surfaces we are implanting today.

Nanomedicine Laboratory

Nano-technology

Tissue Engineering

Better Understanding of Cell FunctionsLeading to Increased Tissue Regeneration

Catalytic

Biosensors

Building Constructs

Electronic Applications

OrthopedicDental

Vascular

Cartilage

BladderNervousSystem

PART I: BONE Nanoparticulate Ceramics

American Ceramic Society Bulletin, 82(6): pp. 1 – 8, 2003.

6 microns

4,520 nm (conventional)Titania

39 nm (nanophase) Titania

Atomic Force Micrographs of Nanophase and Conventional Titania

64

20

4

2

0

0

1.3

microns

microns

microns 0

1.36

4

2

0

microns

64

20microns

6 microns

Webster TJ, Siegel RW, Bizios R, “Osteoblast adhesion on nanophase ceramics,” Biomaterials 20:1221, 1999.

Extensive in vitro studies have demonstrated greater bone cell functions on nano-ceramics

COATINGSNanospherical Ceramics

Working in collaboration with Spire Biomedical, using their IonTiteTM

technology, we have coated traditional orthopedic implant materials with nanospherical ceramics to increase bone growth.

Uncoated Ta Scaffolds

Conventional HA Coated Ta Nano HA Coated Ta Ta Scaffold

COATINGSNanospherical Ceramics

Uncoated Ta Scaffolds

Conventional HA Coated TaNano HA Coated Ta

Implantation Time = 2 weeks; Animal Model = rat calvaria

2 weeks 4 weeks

Bone Bonding NanOss™ HAConventional HATitaniumStainless SteelPolymers

2 weeks8 -12 weeks10-14 weeks12-16 weeksDoes not bond

Angstrom Medica:Bilateral Canine Osseointegration

Distal and Proximal Femur

What About Infection ?

Nanospherical Ceramics:Resisting Bacteria Infection

While increasing bone cell functions, nanomaterials reduce bacteria functions.

Positive Control (Wrought Ti)

Conventional ZnO Nanophase ZnOMag. = 400X

S. Epidermis

Conventional ZnO

Nanophase ZnO

Don’t Like Nanoparticles ?

PART II: BONEAnodized Titanium

Sketch map of anodization system

PROCEDURES:

Pretreatment: chemical polishing using HF/HNO3 mixture

Anodization: 0.5 or 1.5%HF

Voltage: 20V

Time: 20 min

Rinse and dry

Clean: acetone and ethanol

Sterilize

Increased Osteoblast Functions on Anodized Ti

Unanodized Ti

Anodized Ti 1.5% HF treatment

Nanotube

Bar = 1 micron

Anodized Ti0.5% HF treatment

Nano-particles

Ti Screw Insertion

Anodized Titanium Amputee Rat Model

Rat Walking on Anodized Titanium Implant as Soon as 3 Days After Surgery

Lack of Bone Growth with Unanodized TiAfter 28 days of Implantation

Increased Bone Growth with Anodized TiAfter 28 days of Implantation

Tetracycline StainInjection at 4 Days from End of 28 Day Experiment

Unanodized Ti Anodized Nanotubular Ti

No Infection Observed Around Anodized Ti Implant After 28 Days

Unanodized Ti Anodized Nanotubular Ti

Biosensors for Monitoring Orthopedic Tissue Growth

OH

O

CHC

CH3

Om C

O

CH2O Hn

H

HN

HN

N

TitaniumAnodized Ti w/CNT growth

Existing Bone

+New Bone

Growth

Polypyrrole

Poly(L-lactic acid-co-glycolic acid)

Nano-technology

Tissue Engineering



Catalytic

Biosensors

Building Constructs


OrthopedicDental

Vascular

Cartilage


PART III: Central Nervous System

• Chronic probes monitor and apply electrical signals

• Glial scar tissue increases probe impedance

www.cnf.cornell.edu/2001cnfra/20012.pdf

www.engin.umich.edu/facility/cnct/probeback.html

Figure: TEM of Individual Carbon Nano-dimensional Fibers

Bar = 100 nm.

pyrolytic outer core

no pyrolytic outer core

Design of Carbon Nanofibers for Neural Implants

J. L. McKenzie, M. C. Waid, R. Shi, T. J. Webster, “Cytocompatibility of astrocytes on carbon nanofibers,” Biomaterials , available on-line, 2008.

• Decreased functions of astrocytes on carbon nanofibers.

• Decreased functions of astrocytes on polymers containing carbon nanofibers.

• Increased functions of neurons (i.e., neurite extension) on carbon nanofibers.

• Increased functions of neurons (i.e., neurite extension) on polymers containing carbon nanofibers.

Stem Cells and Carbon Nanofibers for Treating Neural Damage

8WMCAO Reperfusion

start

Stroke Reperfusion0h 60min 9h

Stem cell transplant1W 2W 3W

Injection ofCNF +Stem cellsMRI

MRI, Nestin, BrdU stainingNestin/GFAP double stainingNestin/NeuN double staining

Yonsei

MR Analysis- SD rat: MCAO- Injection (50㎕, 31 gauge insulin syringe): SVZ stem cell (1.0 ×104 cells / mL)

+ Hydrophobic or Hydrophilic CNF

4 h 8 h

1 W 3 W

Yonsei

Presenter

Presentation Notes

In T2 MR images, after confirming the infarct area at 8hr after reperfusion, SVZ stem cells and hydrophobic or hydrophilic carbon nano fibers were injected to the injured brain.

Hydrophobic CNF 1w

Presenter

Presentation Notes

15. There are more hydrophobic CNF in the injured brain than hydrophilic CNF. Both of hydrophobic and hydrophilic CNF can help to hold the transplanted SVZ stem cells in the injured brain .

Hydrophobic CNF 3w

Nestin

GFAP CD11b

MAP2

Presenter

Presentation Notes

You can see the nestin-positive stem cells around the hydrophobic CNF and many of MAP2-positive neurons and GFAP-positive astrocytes were observed around hydrophobic CNF.

Animal Behavior Test

Treadmill

Treadmill testing

0

1

2

3

4

EC(1W) SVZ(3W) HL+SVZ(1W)

Step

s

Faster Return of Motor FunctionThrough the Use of Carbon Nanotubes

and Stem Cells

Yonsei

Stroke Stem Cells Stem Cells/(1 W) (3 W) Carbon Nanotubes

(1 W)

N = 9; Data = mean +/- SEM; * p < 0.01 (compared to all others)

**

Presenter

Presentation Notes

For functional recovery, Behavioral tests were performed on 1 week, 3 weeks, 5weeks and 8weeks after CNF and SVZ stem cell transplantation. Unfortunately, I couldn’t get all results of experimental groups now. So I’d like to show some behavioral test data which I can get since now. I am keep going to collect the data for functional recovery. This slide show the treadmill test. For this test, animal set on a treadmill and we count the steps between two lines as you see in the video records. Usually 1week after surgery, we cannot get any functional recovery with a experimental group. However, you can see that rats with SVZ cells (3weeks) and with HL CNF + SVZ cells (1week) had significant functional recovery at 3weeks and 1 week, respectively compared with rats in the experimental control group, and in case of rats transplanted with HL CNF + SVZ cells showed good score at only 1 week after surgery. These data indicates carbon nano fiber doesn’t affect the stem cells recovering the brain from functional deficit.

Nano-technology

Tissue Engineering



Catalytic

Biosensors

Building Constructs


OrthopedicDental

Vascular

Cartilage/Entheses


Part IV:Fibrocartilaginous Entheses

-Niyibizi et al (1994)

Zones of fibrocartilaginous entheses:

1. Pure dense fibrous connective tissue2. Uncalcified fibrocartilage (UF)3. Calcified fibrocartilage (CF)4. Bone / Implant

Ligament proper →

Upper fibrocartilage→

Lower fibrocartilage→Mineralized fibrocartilage→

Bone→

-Metal with porous surface

-Nanomaterial / Nano- surface roughness

- Calcified zone

- Bioactive uncalcified zone

- Cell modulating layer

Bioactive region

Mechanical Interlock to Implant (LPS)

Design of a Composite Device for Regeneration of Entheses (or Soft Tissue Attachment) to Metallic Orthopedic Implants

2. Soft Tissue Product

1. Coated Implant

Bar = 100 µmTop view

Sideview

Region: Metallic Interlock to Implant (LPS)

Metal with porous surface

Presenter

Presentation Notes

Note: Please refer January 9th progress report for notes.

Use of Small Intestine Submucosa

New Entheses Regeneration With Anodized Ti

Implant

Linear CollagenFibers

Fibrocartilage

Failed Entheses Regeneration With Non Anodized Ti

Implant

Loose Coating

Nano-technology

Tissue Engineering



Catalytic

Biosensors

Building Constructs


OrthopedicDental

Vascular

Cartilage


PART V: VASCULARMetals

0

500

1000

1500

2000

Plate Alone(polystyrene)

Wrought Ti Conventional Ti Nanophase Ti

Substrate

RAEC

Adh

esio

n (c

ells

/cm

2 ) LiveDeadTotal

Nano-Structured c.p. Ti IncreasesVascular Endothelial Cell Adhesion

Values are mean +/- SEM; n=3; * p < 0.01 (compared to cell adhesion on conventional and wrought Ti).

**

*

Nano-technology

Tissue Engineering



Catalytic

Biosensors

Building Constructs


OrthopedicDental

Vascular

Cartilage


Bar = 1 µm

Control (Untreated) Sub-micron Structured

Nano-structured

Nano-structured PLGA Increases Bladder Tissue Regeneration

PU = polyurethane

Greater Rat Bladder Regeneration with Nano PLGA

Conventional PLGA:Poor bladder smooth muscle

infiltration after 8 weeks

Nano PLGA:Good replacement of all cell layers after 8 weeks

DiscussionOther research groups have observed altered cell functions on

nanometer compared to conventional topographies:

Increased endothelial cell spreading on 13, 35, and 95 nm islands created by polymer demixing of polystyrene and poly(4-bromostyrene)1. Increased neuron axon extension on quartz with surface features less than 100 nm2.Increased alignment of epithelial cells on 130 nm-wide and 9 micron-deep grooves using holographic photolithography3.

Nano-structured materials possess higher percentages of atoms at the surface, increased portions of surface defects, and greater numbers of material boundaries at the surface that may be influencing protein interactions important for cell function.

1Dalby et al. Biomat. 23:2945-2954 (2002). 2Torimitsu et al., ICCE/9: 795-796 (2002). 3Clark et al., J. Cell Sci. 99:73-77 (1991).

The Future of Nanomedicine

• The design of “raw” material nano-surface properties that can inhibit infection, limit chronic inflammation, and increase appropriate tissue growth (if needed).

• More in situ nanotechnology-derived sensors that sense and respond to events at the implant interface.

• The understanding of nanoparticle toxicity and the design of nanoparticles to limit toxicity in vivo.

• The collaboration of industry and the clinical arenas in nanomedicine.

Nanomedicine Lab

Not pictured: Rajesh Pareta, Batur Ercan, Yupeng Chen, Alyssa Ricker, Ariel Cohen, Aditi Dubey, Joe Carpenter, and Jong Youl Kim

http://openwetware.org/wiki/Image:Lab_member-low_size.jpg�

Angstrom Medica and Spire Biomedical

Argonide Corp. and Applied Sciences, Inc.

Coulter Foundation- Early Career Award

Department of Defense (DARPA)

DePuy Orthopedics (Johnson and Johnson)

Indiana 21st Century Fund

Nanophase Technologies, Corp.

National Science FoundationIntegrated Graduate Education and Research Training Fellowship (IGERT),Nanoscale Exploratory Research, REU

National Institute of HealthNanobiotechnology Initiative

Showalter Foundation

Whitaker Foundation

Acknowledgements

THANK YOU !!and please read the …….

International Journal of Nanomedicine

at: www.dovepress.com

http://www.dovepress.com/�

So why useNanotechnology inTissue Engineering

Applications ?

Bar is 1µm

*Goodman S.L. et al., Biomaterials. 1996 Nov;17(21):2087-95.

Cast Replica of Vascular Tissue Demonstrating Nanometer Roughness *

• Due to the presence of numerous nano-structures (i.e., proteins) in the body, cells are accustomed to interacting with surfaces that have a large degree of nanometer roughness.

• Despite this fact, many current syntheticmaterials used as tissue engineeringscaffolds possess conventional surface features only.

Our Tissues Are Nano-structured

AFM Scan of Bovine FemurCortical Bone

5 X 5 µm AFM Scan

Successful Tissue Engineering Materials Depend on Optimal Surface Properties

for Cell Function

Surface properties affecting protein conformation/bioactivity:Wettability; topography; etc.

Cellmembrane

Adhesive peptide sequence of protein(for example Arginine-Glycine-Aspartic Acid (RGD))

Cell

Integrin receptors

Proteins(for example : vitronectin, fibronectin, laminin, collagen, etc.)

Adapted and redrawn from Schakenraad, J.M . pp. 140-141, in Biomaterial Science (B. Ratner et al., eds.), Academic Press, Inc., San Diego, CA, 1996; T. J. Webster, in Advances in Chemical Engineering Vol. 27, Academic Press, NY, pgs. 125-166, 2001.

Documents

Nanomedicine : Where Have We Been and Where are We GoingWorldwide market for nanoscale devices was 406 million dollars in 2002. 2003. Congressional hearing on societal implications