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INTRODUCTION TO BIOADHESION CHRISTINE ORTIZ, Associate Professor Department of Materials Science and Engineering, MIT WWW :

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Presentation on theme: "INTRODUCTION TO BIOADHESION CHRISTINE ORTIZ, Associate Professor Department of Materials Science and Engineering, MIT WWW :"— Presentation transcript:

1 INTRODUCTION TO BIOADHESION CHRISTINE ORTIZ, Associate Professor Department of Materials Science and Engineering, MIT WWW : http://web.mit.edu/cortiz/www c D. Breger, used w/permission, http://www.ldeo.columbia.edu/micro/images.section/pages/bloodclot.html

2 BIOADHESION : DEFINITION Bioadhesion may be defined as the state in which two materials, at least one of which is biological in nature, are held together for extended periods of time by interfacial forces. In the context of their medical and pharmaceutical use, the term bioadhesion refers to the adhesion of synthetic and biological macromolecules to a biological tissue. The biological substrate may be cells, bone, dentine, or the mucus coating the surface of a tissue. If adhesive attachment is to a mucus coating, the phenomenon is sometimes referred to as mucoadhesion. Many examples of bioadhesion exist in nature, including such diverse events as cell-to-cell adhesion within a living tissue, barnacles binding to rocks, and bacteria binding to tooth enamel. In health care, bioadhesives were first used as wound dressings, skin adhesives, and denture fixatives. Over the last two decades, bioadhesives have been of interest within the pharmaceutical sciences for their potential to optimize drug delivery. Such drug delivery may be optimized at the site of action (e.g., on the cornea or within the oral cavity) or at the absorption site (e.g., in the small intestine or nasal cavity). Bioadhesives may also be used as therapeutic agents in their own right, to coat and protect damaged tissues (gastric ulcers or lesions of the oral mucosa) or to act as lubricating agents (in the oral cavity, eye, and vagina). Skin adhesives, tissue sealants, and dental and bone adhesives and cements are also defined as bioadhesives. This article first focuses on the types of muco/bioadhesives currently used in the pharmaceutical sciences, from first-generation hydrophilic polymers to second-generation polymers and lectins. The nature of bioadhesive interactions, types of bioadhesive formulations developed, and regions of the human body to which they may be administered are also considered. Other types of medical bioadhesives, such as those used in wound management, surgery, and dentistry, are also discussed.

3 Blood and Blood Vessels 40% cells in plasma or serum (pH7.4, IS=0.15 M) which contains 6-8% proteins (over 3,000 different types) in HOH, including : -58% albumins -38% globulins -4% fibrinogens

4 http://www.artegraft.com/ Synthetic Vascular Grafts or Prosthesis : prosthetic tube that acts either a permanent or resorbable artificial replacement for a segment of a damaged blood vessel (e.g. from athersclerosis,aneurysms, organ transplant, cancer, arteriovenous fistula, diabetes) : $200 million market worldwide http://www.atriummedical.com/ http://www.vascutek.com/

5 expanded polytetrafluoroethylene (Gore-Tex, ePTFE) -fibrillated, open cell, microporous (pore size 0.5-30  m), 70% air, nonbiodegradable, chemically stable, used for 26 yrs, hydrophobic/nonpolar, flexible polyethylene terephthalate (Dacron, PET) -multifilamentous yarn fabricated by weaving/knitting, amphiphilic, smaller pores than ePTFE polyurethane derivatives bovine collagen -fibrous, hydrophilic Zhang, et al. J. Biomed. Mtls. Res.60(3), 2002, 502. Vascular Graft Materials www.vascutek.com

6 adsorbs BLOOD PRESSURE+ ATTRACTIVE FORCES denatures PLATELETS! http://www.rinshoken.or.jp/org/CR/photo-e.htm Solid-Liquid Interface blood plasma proteins BLOOD FLOW BIOMATERIAL SURFACE BLOOD CLOT! - acute occlusive thrombosis - infection / inflammation - neointimal hyperplasia D. Gregory http://medphoto.wellcome.ac.uk

7 Total Intersurface Force as a Function of Separation Distance :F(D) WHAT CONTROLS PROTEIN ADSORPTION? END- GRAFTED POLYMER “BRUSHES” ADSORBED POLYMER LAYERS BIOMATERIAL SURFACE Many different components, both attractive (e.g. hydrogen, ionic, van der Waals, hydrophobic, electrostatic) and repulsive (e.g. configurational entropy, excluded volume, osmotic, enthalpic, electrostatic, hydration), can lead to complex interaction profiles. D

8 chemically end-grafted PEO 50K “mushroom” L contour= 393 nm R F =8.7 nm F sodium phosphate buffer solution IS=0.01M pH=7.4 D lipid-bound HSA functionalized probe tip, R TIP ~65 nm (SEM) Au-coated silicon chip covalently immobilized HSA ~10 nm s = 62 ± 28 nm ~35-190 proteins in maximum interaction area (D=0) ~2.5 PEO chains in maximum interaction area (D=0) Si 3 N 4 Direct Measurement of Protein Interactions with Poly(ethylene oxide) (PEO) Macromolecules Rixman, et al. accepted, Langmuir 2003.

9 Chemical Attachment Scheme of Lipid-Bound HSA to Si 3 N 4 Probe Tip A. Vinkier; Heyvaert, I.; D'Hoore, A.; McKittrick, T.; C., V. H.; Engelborghs, Y.; Hellemans, I. Ultramicroscopy 1995, 57, 337. S. O. Vansteenkiste; Corneillie, S. I.; Schacht, E. H.; Chen, X.; Davies, M. C.; Moens, M.; Van Vaeck, L. Langmuir 2000, 16, 3330. Fluorescence micrograph of HSA-functionalized cantilever (courtesy of Irvine Lab-DMSE) probe tip location

10 (*Steve Santoso (MIT-Biology) http://pymol.sourceforge.net) Human Serum Albumin (HSA) M. O. Dayhoff Atlas of Protein Sequence and Structure; National Biomedical Foundation: Washington DC, 1972. S. Azegami; Tsuboi, A.; Izumi, T.; Hirata, M.; Dubin, P. L.; Wang, B.; E., K. Langmuir 1999, 15, 940-947. II  The smallest and most abundant blood protein in the human body, HSA accounts for 55% of the total protein content in blood plasma  3-D structure consists of 3 homologous subdomains, each containing 5 principal domains and 6 helices.  Subdomains form hydrophobic channels placing basic and hydrophobic residues at the ends while the surface remains predominantly hydrophilic  L contour = 225 nm  Isoelectric point=4.7  116 total acidic groups (98 carboxyl and 18 phenolic -OH) and 100 total basic groups (60 amino, 16 imidazolyl, 24 guanidyl). I(N) II III(C)

11 “HEART SHAPED” STRUCTURE OF CRYSTALLIZED HSA (Curry, S., H. Mandelkow, et al. Brookhaven Protein Databank.) C  8 nm PROPOSED ELLIPSOIDAL STRUCTURE OF HSA IN SOLUTION (Haynes, et al. (1994). Coll. Surf. B. : Biointerfaces 2: 517.)  14 nm -9e-8e+2e I (N) III (C) II  4 nm charge residue map - red, + bluehydrophilic-hydrophobic map

12 AFM Images of End-Grafted (Mono-Thiol) PEO 50K Chains on Polygranular Gold Substrate (*contact mode, solvent=PBS buffer solution, IS=0.15, pH=5.6) 100 nm 50 nm polygranular Au Au-PEO 50K distance between polymer chains= =62  26.8 nm 2 =2.610 -4 nm -2

13 Poly(ethylene oxide) (PEO) In Aqueous Solution (Prog. Polym. Sci. 20, 1995, 1043) hydrophilic & water soluble @RT low  <0.5, high  2 =30-60 cm 3 mol/g 2 (large excluded volume),  W(A) =60 o intramolecular H- bond bridges between -O- groups and HOH maintains some hydrophobic character high flexibility, low  =1.38-1.95 high mobility, fast  c  =15-100 ps locally (7/2) helical supramolecular structure (tgt axial repeat = 0.278 nm) low van der Waals attraction neutral t 0.278 nm g t t tt t g t t t (tgt) Nature 416, 409 - 413 (2002)

14 DETERMINATION OF SURFACE INTERACTION AREA AND CONTACT AREA R TIP D MAX r R TIP -D MAX surface interaction (tip and substrate not in contact) PROBE TIP D MAX <100 nm, R TIP <100 nm A TIP (D=0) = 3000-17,000 nm 2  ~40-180 proteins for a monolayer A CONTACT < 3 nm 2 (tip and substrate in contact  negligible substrate deformation) SUBSTRATE aqueous solution F MAX F MAX /protein<40pN Rixman, et al. accepted, Langmuir 2003.

15 “APPROACH” (COMPRESSION OR LOADING)

16 F Au R F (PEO) magnitude of force much larger than predicted by theory Rixman, et al. submitted, Langmuir 2003. AVERAGE APPROACH CURVE : HSA PROBE TIP VERSUS PEO (SUBTRACTED AU INTERACTION) PBS, IS=0.01M, pH=7.4

17 HSA versus PEO : Effect of NaCl IS Approach CONCLUSION: Electrostatic double layer and configurational entropy are outweighed by another interaction which increases with IS →possibly due to water interphase layer R F (PEO) ● Salt screening : electrostatic double layer force expected↓ with ↑IS ● NaCl reduces the goodness of solvent for PEO (Armstrong, et al. 2001) : configurational entropy force expected↓ with ↑IS Rixman, et al. 2003 unpublished data

18 HSA versus PEO : Effect of Solvent on Approach Isopropanol has been shown to block hydrophobic interaction forces (Jiang, et al 2002) R F (PEO) Rixman, et al. 2003 unpublished data

19 Poly(ethylene oxide) (PEO) : REPULSIVE INTERACTIONS IN WATER neutrality : won’t attract oppositely charged species hydrophilic/ water soluble : hydration enthalpic penalties for disruption of supramolecular structure H-bonding with water steric (large excluded volume) electrostatic double layer forces - - - - - - - - - - - - - - - high flexibility & mobility : no local steric or charge

20 “RETRACT” (TENSION OR UNLOADING)

21 Quantities Used to Evaluate Nanoscale Adhesion, /Radius, = average maximum attractive force and corresponding separation distance within a dataset recorded for each point of pull-off and averaged over an entire data set, /protein=effective adhesive interaction energy per unit area : BCP Theory (  =1.4), JKR (  =1.5), DMT Theory (  =2) :, /A SUBSTRATE =energy dissipated during loading- unloading cycle Limitation : can’t use for curves exhibiting large adhesive forces followed by large cantilever instability regions (weak cantilever).

22 INDIVIDUAL APPROACH AND RETRACT CURVES, HSA PROBE TIP VERSUS PEO-AU SURFACE, PBS, IS=0.01M, pH=7.4 F 76% of total experiments Au reversible decompression of the (net) repulsive surface interaction and no adhesion Au Rixman, et al. submitted, Langmuir 2003.

23 adhesive binding force unknown desorption interaction profile long-range adhesion due to stretching of individual PEO chain F Au extension of individual PEO chain (net) repulsive surface interaction nonspecific adsorption tether nonhysteretic repulsion 17% of total experiments INDIVIDUAL APPROACH AND RETRACT CURVES : HSA PROBE TIP VERSUS PEO-AU SURFACE, PBS, IS=0.01M, pH=7.4 F RUPTURE (Au-S)  2-3 nN Rixman, et al. submitted, Langmuir 2003.

24 INDIVIDUAL APPROACH AND RETRACT CURVES: HSA PROBE TIP VERSUS PEO-AU SURFACE : PBS, IS=0.01M, pH=7.4 F 7% of total experiments Au extension of 2 PEO chains Rixman, et al. submitted, Langmuir 2003.

25 adhesive binding force unknown desorption interaction profile long-range adhesion due to stretching of individual PEO chain nonhysteretic repulsion 17% of total experiments INDIVIDUAL APPROACH AND RETRACT CURVES : HSA PROBE TIP VERSUS PEO-AU SURFACE : PBS, IS=0.01M, pH=7.4 =0.16±0.18 nN =265±137nm /Radius= 2.46±2.76 mN/m not calculated (DMT, JKR, BCP theories not applicable) =1.31E3 k B T /A SUBSTRATE =0.5 mJ/m 2 one polymer chain Rixman, et al. submitted, Langmuir 2003.

26 strain-induced conformational transition (ttg  ttt) t 0.278 nm g t t tt t g t t t (tgt) INDIVIDUAL APPROACH AND RETRACT CURVES : HSA PROBE TIP VERSUS PEO-AU SURFACE : PBS, IS=0.01M, pH=7.4 (*first reported by Oesterhelt, et al. 1999) reduction in extensional force reversible on experimental time scales CREATION OF MOLECULAR ELASTICITY MASTER CURVE

27 F adhesion /Radius (mN/m) F adhesion (nN) /Radius (mN/m) (nm) (nN) ADHESION FORCES AND DISTANCES FOR INDIVIDUAL RETRACT CURVES, HSA PROBE TIP VERSUS VARIOUS SURFACES : PBS, IS=0.01M, pH=7.4

28 NH 2 OH H-bonding SUMMARY OF RESULTS : PROTEIN-PEO INTERACTIONS Large, long-range surface repulsion that can’t be explained by electrostatic and steric interactions alone (?  WATER) Elimination of surface adhesion (from ~1.35 nN) even at such low grafting densities At high compressions, long range adhesion ( =160 pN) and stretching with an individual PEO 50K chain allows the probing of short-range attractive contacts between surface functional groups and an individual PEO chain

29 ADVANTAGEOUS MOLECULAR ATTRIBUTES FOR MAXIMUM BIOCOMPATIBILITY 1) maximum hydrophilicity and water solubility, i.e. molecules capable of strong hydrogen bonding such that there exists an enthalpic penalty to dehydration and disruption of supramolecular structure imposed by incoming protein molecules 2) a net neutral charge so that the surface will not attract proteins of net opposite charge or regions on a protein surface of opposite charge via electrostatic interaction. 3) for macromolecular surfaces, higher molecular weight, long chains with a large degree of backbone flexibility to produce maximum steric repulsion 4) Nontoxic HOW DO BLOOD VESSEL INTERIOR (LUMEN) SURFACES CONTROL NONSPECIFIC ADSORPTION?

30 Glycocalyx : External, Porous, Dynamic, Densely Carbohydrate Rich Region of Cell Membrane That Play a Role in Cell-Cell Recognition and Also Prevents Non-Specific Interactions, 500 nm thick (Vink, et al 1996 Circ. Res. 79, 581) Presumably, artificial biomaterial surfaces can be made more compatible if they are more similar in chemistry, morphology, and mechanical properties to the cell surface. http://www.d.umn.edu/~sdowning/Membranes/ Control of Nonspecific Adsorption In Blood Vessels

31 Glycocalyx-Mimetic Neutral Oligosaccharide Monolayers (Synthesized by Seeberger Lab, MIT-CHEM) chitobiose (CB) oligomannose-9 (Man-9) linear trimannoside (LT)

32 Glycocalyx-Mimetic Neutral Oligosaccharide Monolayers (Synthesized by Seeberger Lab, MIT-CHEM)

33 Plant Fibers cellulose


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