DENTIN

BIOMODIFICATION
AGENTS
1
2 CONTENTS
 Introduction

 Structure of dentin

 Definition of dentin biomodification

 Reason for dentin biomodification

 Extracellular matrix components relevant to dentin biomodification

 Types of dentin biomodification agents

 Clinical applications of dentin biomodification

 Conclusion

 References
3 INTRODUCTION

 Dental caries is prevalent all around the world, and resin composites are widely
used as filling materials
 However, longevity of resinous restorations is not satisfactory with failure rate
ranging from 15% to 50%
 In addition, resin-dentin bonds are less durable than resin-enamel bonds, because
of the heterogeneity of the structure and composition of dentin.
 The failure of resin-dentin bonding results in microleakage, staining, recurrent
caries, and postoperative sensitivity, and the interaction of these situations can
further accelerate the degradation of the bond.
4

 Even though adhesive systems have significantly improved, the bonded interface
remains the weakest area of resinous restorations.
 Improving the chemical and mechanical stability of the collagen fibrils within the
hybrid layer may be of clinical importance to enhance resin-dentin bond
durability.
 This may be achieved by stabilization of dentin collagen with biocompatible
cross-linking agents to increase mechanical properties and decrease enzymatic
degradation with matrix metalloproteinase inhibitors (MMPI).
5 STRUCTURAL COMPONENTS

• Dentin - a complex mineralized tissue forms

main part of tooth structure DENTIN

• Composed of inorganic components

(hydroxyapatite crystals) & organic
components (mostly type I collagen fibers) PERITUBUL INTERTUBU
AR DENTIN LAR DENTIN
6 PERITUBULAR DENTIN

 Dentin that immediately surrounds dentinal

tubules

 5%- 12% more mineralized than inter

tubular dentin

 95% volume minerals

 Contains little collagen

 Appears to be enriched in non collagenous

matrix proteins – DSP
7 INTERTUBULAR DENTIN

 Forms main body of the dentin

 Primary secretary product

 Less mineralized than peritubular dentin

 30% volume of minerals

 Fibrils are arranged roughly at right angles to

dentinal tubules
8 DEFINITION

Biomodification of dentin is a biomimetic strategy therapy to mechanically

strengthen the existing collagen network and also control biodegradation rates of

extracellular matrix (ECM) components.

9
WHY DENTIN BIOMODIFICATION ?

• Current restorative procedures depends on the infiltration of synthetic polymers of

the adhesive system into partially or totally demineralized collagen fibers that make
up the dentin organic matrix, which is called Hybrid layer.

• Factors affecting bond strength :-

• excess hydrophilic monomers incorporated in the adhesive systems,

• high water concentration in the bonding procedure,

• inadequate infiltration of the monomers into the demineralized

collagens.
10

• Water in hybrid layer causes hydrolysis of monomers and unprotected collagen

fibres.

• Endogenous proteases such as MMPs (2, 3, 8, 9 and 20) and cathepsins targets
non-infiltrated & exposed collagen lesions.
11 Other factors leading to resin dentin failure:
• Discrepancy between adhesive resin infiltration and dentin demineralization depth
• Degradation of exposed collagen
• Degradation of the adhesive resin

Biomodification of dentin substrate involves pretreating it with MMPIs and collagen

cross‑linkers (CCL) as dentin biomodifiers improve mechanical properties by promoting a
more stabilized and durable adhesive interface
12 Extracellular matrix components relevant to dentin biomodification

1. Type I collagen:-

 most abundant of all collagen types.

 coiled-coil trimer molecule.

 repeated sequence of

amino acids Gly- proline and hydroxyproline)

Russo AK et al. Dentin Biomodification: Strategies, Renewable Resources and Clinical Applications.
Dent Mater 2014
13 • Inter-molecular cross-linking - basis for the stability, tensile strength &
viscoelasticity of the collagen fibrils

Collagen
cross linking

endogenous exogenous

Mediated by Induced by nonenzymatic

reactions
- chemical agents
- physical method
Enzymatic non-enzymatic
reaction reactions
14 2. Proteoglycans :-

 Main group of non-collagenous proteins in both pre-dentin and dentin.

Function:-
 play role in dentin mineralization and structural integrity of collagen fibers
 control tissue hydration and molecular diffusivity

Proteoglycans

Chondroitin/keratan sulfate Small leucine-rich

family proteoglycans (slrps)
15 3) Endogenous proteases

1) Matrix metalloproteinases (MMPs)

• Calcium and zinc dependent endopeptidases

• Capable to destruct all types of extracellular matrix (ECM) and basement

membrane (BM) proteins

• In pulp dentin complex : MMP -2, -3, -8, -9

Russo AK et al. Dentin Biomodification: Strategies, Renewable Resources and Clinical Applications.
Dent Mater 2014
16 MMP family has 23 members. This family is divided into 6 groups which are as follows:

Play a role in
 collagen matrix breakdown during dental caries and periodontal diseases.
 resin-dentin bond interface degradation.
 pulp inflammation and cancer
17 2)Cathepsins
 Become active at acidic pH

 Most are endopeptidases, with some exceptions like cathepsin B that can also act as a
carboxypeptidase

Functions-
• ECM degradation in physiological and pathological processes like bone remodeling,
inflammation, rheumatoid arthritis, diabetes, multiple sclerosis and cancer .
• caries progression

Russo AK et al. Dentin Biomodification: Strategies, Renewable Resources and Clinical Applications.
Dent Mater 2014
18

Endogenous proteases ,MMPs and cathepsins are active in intact and carious dentin and

show decreased activity with age in sound dentin

19 DENTIN BIOMODIFICATION AGENTS

Based on sources

Freitas BF et al. OHDM 2019

20 Based on action
MMP INHIBITORS CROSS – LINKING AGENTS
Chlorhexidine (CHX) Riboflavin (RB)
Ethylenediaminetetraacetic Carbodiimide
Acid (EDTA)

Galardin Proanthocyanidin
Quaternary Genipine
ammonium salts

Tetracycline (TC) and Polyphenols

analogues

Epigallocatechin-
3-gallate (EGCG)

Proanthocyanidin , carbodiimide & gluteradehyde – both effects

21
PHYSICAL METHODS

 Also called the Photo-oxidative method

 Uses Ultraviolet light (365 nm ) + Riboflavin

 Mode of action:-

UVA :- breaks down the intrinsic bonds of

collagen & generates oxygen free radicals

Riboflavin:- Hydroxyl functional clusters in

RF attack proline or lysine in the collagen.

In addition, RF acts as photosensitizer for UVA,

promoting the formation of new cross-links
22

bridging of amine groups(N H) of glycine of one chain with carbonyl groups (C O)of
hydroxyproline and proline in adjacent chains

Increased resistance to collagen degradation mediated by collagenase, in addition to

significant improvement in tensile strength

Freitas BF et al. OHDM 2019

23
 Application:- incorporate RF into adhesive

 Concentration of RF :- 1% & 3%

1% - increase microtensile bond strength

3% - improve immediate bond strength

 Toxicity :- less toxic

 Alternative to UVA:- visible blue light (clinically more applicable)

24 CHEMICAL METHODS

1) Synthetic agents :-

1. Glutaraldehyde

• Has the ability to cross links with amino

groups

of collagen

• Mode of action:- increase in tensile strength and

elasticity with decrease the rate of collagen

degradation

Freitas BF et al. OHDM 2019

25
• Application :- after acid etching as a primer

• application time :- 60 sec

• concentration :- 5% by weight

• Nano-leakage :- seen in immediate period

• toxicity :- highly toxics

26 2. Carbodiimide (EDC)

• Has been used as an alternative to glutaraldehyde

• Application time :- 1 to 4 hours; clinically

unacceptable

studies have shown that even 1 min pre-treatment is

adequate

• toxicity :- less toxic

• Its potential is non-specific by the activation of the

carboxylic acid groupings of glutamic and aspartic acids to
form an O-acylsourea intermediate. After, reacts with the
amino groups of lysine and hydroxylysine to form an amide
27
Drawback :-
 limited cross linking ability
 inhibitory effect on endogenous & exogenous enzymatic activities

Freitas BF et al. OHDM 2019

28
II. Galardin :-

• Specific MMP inhibitor (MMP 1,2,3,8.9)

• Mode of action :- chelation of Zn ion on catalytic domain of MMP

• Concentration:- 0.02 mM

• Drawbacks :- decreased bond strength

nanoleakage

Freitas BF et al. OHDM 2019

29 III)Chlorhexidine

• first used as a dentin disinfectant & re-wetting agent prior to

application of the adhesive resin

• Most commonly used

• Concentration :-

for etch & rinse method - 0.2%

for self etch method- 0.1%

• Application time :- 60 sec

• Application:- topically in an aqueous solution , prior to

application of adhesive
30
Mode of action

• MMP inhibitor (MMP 2,8,9) & strong inhibitor of cysteine cathepsins activity

• Even at low concentrations (0.2%), chlorhexidine showed preservation of

bond strength and reduced interface degradation, expressed by the reduction

of nanoinfiltration.

• The inhibition mechanism of chlorhexidine occurs by inhibition of proteases

because of its chelating property with calcium ions

Freitas BF et al. OHDM 2019

31
• Toxicity :- less toxic

Draw backs :-

 increase water entrapment in smear layer

 incorporation in adhesive – CHX may diffuse out of dentin matrix

 low substantivity when applied on dentin

32
IV. Quaternary Ammonium Salts :-

Benzalkonium chloride (BAC) :-

• MMP inhibitor

• Mode of application:- incorporated in 37% phosphoric acid

• binds strongly to demineralized dentin even after rinsing

• Concentration:- 1%

• Application time:- 15 sec

Nagpal R et al. Dentin biomodifiers to stabilize the bonded interface. Mod Res Dent 2020
33 V. EDTA:-

• Concentration:- 17%

• EDTA acts as a chelating agent, reacting with calcium ions from dentin
hydroxyapatite and forms soluble calcium salts.

• As EDTA is an effective Zn2+ and Ca2+ chelator, it might inhibit MMP activity

• Draw back:- removed from dentin by extensive rinsing

Nagpal R et al. Dentin biomodifiers to stabilize the bonded interface. Mod Res Dent 2020
34 2) Naturals agents:-

I. Proanthocyanidin(PA) :-

• Most effective MMP inhibitor (MMP 2&9)

• Sources :- Grape seed , cinnamon , apple , cocoa beans, etc

• Application time :- varied 10 min to 1 hour (not clinically practical)

• PA could effectively cross link collagen in time period as short as 10 sec

Nagpar R etal. Proanthocyanidin: A natural dentin biomodifier in adhesive dentistry.

Journal of Restorative Dentistry 2016
35
Mode of action:- 3 mechanisms

1. Protease resistance - via irreversible conformational changes of proteases

which are co-participating in collagen biodegradation

2. Indirectly interfere with protease production & activation by modulating host immune
response

3. Increase density of collagen network by inducing exogenous cross links & decreases
collagenase absorption induced by reduced swelling

Nagpar R etal. Proanthocyanidin: A natural dentin biomodifier in adhesive dentistry.

Journal of Restorative Dentistry 2016
 Grape seed extract Cocoa seed extract

36

% of PA 95% 45%

Favourable solvent Ethanol based adhesive Acetone based adhesive

Application time 10 min 10 min

Micro tensile bond Higher immediate bond More stable at long time
strength strength

Castellan CS et al. Effect of Dentin Biomodification Using Naturally Derived Collagen Cross-
Linkers: One-Year Bond Strength Study. Int J Dent 2013
37
Advantages:
• The application of PACs in human dentin mimics different levels cross-linking
of collagen by non-enzymatic interactions resulting in improved biomechanics
and biostability
• In addition, they increase the tensile strength
• Increased resistance to biodegradation
• Increased modulus of elasticity
• The ability to connect in proline-rich proteins favors immediate adhesion and
stabilization and to collagen fibers, ever in demineralized dentin
• Less toxic

Freitas BF et al. OHDM 2019

38

Draw back:-

• hampers the monomer conversion

• alters the polymerization kinetics in all adhesives regardless of the

photoinitiators
39
II. Genipin :-

• Mode of action:- intra molecular cross linking

• Concentration:- 0.5%

• Toxicity :- less toxic

Draw back :- slow cross linking reaction

deep blue staining of soft tissue

40

Cardol and cardanol

• Derived from the liquid cashew nut (LCC, Anacardiumoccidentale)
• Its chemical structures are phenols, which are capable of cross-links through
hydrogen bonds chain responsible for a hydrophobic characteristic.
• High resistance against water biodegradation
• Studies showed that they yield overall best dentin biomodification outcomes
when applied for one minute without staining the dentin collagen
41
The aroeira extract (Myracrodruon urundeuva)
• It is an anti-inflammatory and antimicrobial agent.
• When tested in dentin showed improvements in properties mechanical
properties of dentin and degradation

Freitas BF et al. OHDM 2019

42
Green tea (Camellia sinensis)
• The main polyphenol found in green tea (Camellia sinensis) and inhibitor of
MMP (2 and 9) is epigallocatechin 3-gallate (EGCG).
• Beyond the ability to cross-link, it has an antimicrobial potential against Gram-
positive and Gramnegative bacteria.
• The application of 0.1% EGCG as a dentin pretreatment associated with an
adhesive conventional 2-step design resulted in the maintenance of the bond
strength after 6 months of water storage
43

• Chitosan showed a significant increase in the resistance to degradation of

collagenase after dentin is coated with nanoparticles of chitosan (CSnp).

• It also showed increase the microhardness of the root dentin layer

• The chitosan modified with methacrylate is proposed as a component of an etch-

and-rinse adhesive, a system to improve the durability of dental restorations.
44

In addition, the use of chitosan/riboflavin to modify collagen dentine, with the

defined proportions, stabilizes the fibrillar network of collagen enhances resin
infiltration and hybrid layer formation
45

Clinical applications of dentin

biomodification

Russo AK et al. Dentin Biomodification: Strategies, Renewable Resources and Clinical

Applications. Dent Mater 2014
46
1. Improving tissue biomechanics :-

• The mechanical properties of the denuded collagen fibrils is inferior to resin-

infiltrated collagen or mineralized collagen.

• Covalent inter- & intra-molecular cross-links are the basis for stability, tensile
strength, and viscoelasticity of the collagen fibrils.

• Biomodification agents are capable of doing that which will increase tensile
strength, viscoelasticity , durability of bond strength
47

• The mechanical properties of the underlying dentin and hybrid layer are significantly
improved (agent-mediated non-enzymatic collagen cross-linking)

• Long-term stability has also been reported and attributed to irreversible collagen
cross-linking and reduced dentin matrix biodegradation

• EDC (carbodiimide)/NHS(N-hydroxysuccinimide) have a significant long-term effect

on the resin–dentin bond strength
48
1) Tooth-biomaterials interfaces:-

 The most common cause of replacement of resin composite restorations is

secondary caries

 The hybrid layer is the weakest link at the adhesive interface bond.

 These denuded collagen matrices are also filled with water, which serves as a
functional media for the degradation of resin matrices and collagen

 Strengthening of type I collagen and reduced biodegradation are likely to reinforce

the dentin structure & in combination are promising strategies to develop a strong
and long lasting tooth-biomaterial interface.
49
2) Biodegradation–inhibition of proteases:-

• Low pH environment during restorative procedures -release and activate MMPs

and cathepsins -cleavage of exposed dentin collagen

• Collagenolytic and gelatinolytic activities occur in dentin and hybrid layers

• Dentin biomodification impair exogenous and endogenous proteolytic activity in a

concentration dependent manner.
50
Proroanthocyanidins (PACs):-

• induces remarkable resistance against degradation either from bacterial collagenase

or endogenous proteases such as MMPs

• Down regulation of endogenous proteases expression, protease inactivation/silencing

& protection of cleavage sites within collagen

• Potent inhibitory effects of EDC and Riboflavin/UVA on MMPs

51
2) Dental caries prevention and remineralization

• The PACs affinity for proline-rich proteins increases their applicability on root dentin
caries.

• They demonstrate a potent anti-bacterial effect

• PAC-rich grape seed extract- decreased the in vitro caries progression and
remineralization of root dentin .

• Due to:

• stabilization of dentin matrix for matrix remineralization

• formation of calcium-PAC complexes promoting mineral deposition

Russo AK et al. Dentin Biomodification: Strategies, Renewable Resources and Clinical Applications.
Dent Mater 2014
52
Green tea

• Protective function on dentin erosion and

abrasion

• reduced dentin wear, due to polyphenols

acting as inhibitors of MMP.

• showed better microhardness indicating

stabilization of collagen in dentine resulting in
functional remineralization.
53

• Biomodification agents did not impair adhesive polymerization.

• No significant difference in bond strength was seen among different groups

(Cardanol and PA) except for Cardol which demonstrated a positive influence on
intraradicular dentin bonding for glass-fiber post luting.
54 CONCLUSION

 Dentin biomodification is a bio-inspired strategy to enhance the properties of

dentin matrix with major impact on novel preventive and reparative/regenerative
dental therapies.

 It is essential to consider the mechanism and limitations of a wide range of

available strategies

 Preliminary in vitro studies have provided strong evidence of its application in

dentin–resin bonded interfaces and caries development/progression.

 PACs appear to be most promising due to their biocompatibility, high dentin

bioactivity and avail-ability as renewable resources.
55 REFERENCES:-

 Dentin biomodification: strategies, renewable resources and clinical

applications Ana K. Bedran-Russo a,∗, Guido F. Pauli b

 Dentin Biomodification Agents in Dentistry–A Critical Review Bárbara de

Fátima Barboza de Freitas

 Dentin Biomodification Potential Depends on Polyphenol SourceT.R.

Aguiar1 , C.M.P. Vidal1 , R.S. Phansalkar

 A self-assembly and higher order structure forming triple helical protein as

a novel biomaterial for cell proliferation† Meganathan Ilamaran, a Asuma
Janeena, a,c Sisila Valappil, a,c
56

 Dentin: Structure, Composition and Mineralization Michel Goldberg1 ,

Askok B. Kulkarni2 , Marian Young

 Matrix Metalloproteinases: From Structure to Function Marco

Fragai1,2 and Claudio Luchinat1,2

 Galloyl moieties enhance the dentin biomodification potential of

plant-derived catechins Cristina M.P. Vidal a , Thaiane R. Aguiar