Describes about the biosynthesis,degradation, regulation of collagen and its importance in periodontium.

1. BIOSYNTHESIS OF COLLAGENOUS
PROTEIN
SHEETHALAN.M.S.R

2. CONTENTS
INTRODUCTION
STRUCTURE OF COLLAGEN
TYPES OF COLLAGEN
TISSUE DISTRIBUTION OF COLLAGEN
DISTRIBUTION OF COLLAGEN TYPES IN PERIODONTIUM
BIOSYNTHESIS OF COLLAGEN
REGULATION OF COLLAGEN BIO SYNTHESIS
DEGRADATION & REMODELLING
SYSTEMIC DISEASES ASSOCIATED WITH COLLAGEN ALTERATION
ALTERATION OF COLLAGEN IN PERIODONTAL DISEASE
USES OF EXOGENOUS COLLAGEN
CONCLUSION

3. INTRODUCTION
 Collagen is protein structures distributed throughout the
animal kingdom.
 The word collagen is derived from Greek roots Kolla (glue)
and gene.
 In French the word collagene designates glue producing
constituents because collagenous tissues were used as sources
of glue and gelatin.
 Collagen constitutes about one third of the total protein in the
body.
 Collagen, in the form of elongated fibrils, is mostly found
in fibrous tissues such as tendon, ligament and skin, and is
also abundant in cornea, cartilage, bone, blood vessels, the
gut, and intervertebral disc.

4. INTRODUCTION
 Collagen constitutes one to two percent of muscle tissue, and
accounts for 6% of the weight of muscles.
 Collagen forms the major structural component it also alters the
cell shape, differentiation and the activities of extracellular matrix
forming an important group of multifunctional connective tissue
protein.
 Collagen is an important component of tissues like periodontal
ligament and tendon where mechanical forces need to be
transmitted without loss.

5. STRUCTURE OF COLLAGEN

6. • The structure of triple helix was first described in detail
by Ramachandran and Kartha (1955).
• The whole molecule is approximately 300 nm long and 1.5 nm
in diameter.
• Collagen consist of triple helical structure formed by three
polypeptide chains called  chain.
• The  chains are left handed helices that wrap around each
other to form right handed, rope like triple helical rod.
The three left-handed helices are twisted together into a right-
handed coiled coil, forming a triple helix or super helix.

8. STRUCTURE OF COLLAGEN
 The triple helix may be a continuous stretch, or interrupted by non
collagenous segments.(Ramachandran&Ramakrishnan,Piez 1976)
 Glycine occupies every third position within the repeated amino acid
sequence in the triple helical domain
Sequence often follows the pattern Gly-Pro-X or Gly-X-Hyp.
 Glycine is essential because any larger amino acid cannot fit in the
center of triple helix

9.  Hydroxyproline and Hydroxylysine are unique amino acids .
 The collagen molecule is stabilized by a number of lysine
derived intra and inter molecular cross links.

10. TYPES OF COLLAGEN
 19 different types of collagen have been described by Prockkop and
Kivirrikko 1995 ; van der Rest and Garrone 1991)
 It ranges from type I to type XIX.
 They are divided into three groups depending on their ability to form fibrils.
1.Fibril forming collagens.
2.FACIT collagens.
3.Non fibrillar collagens

11. Fibril forming collagens:
-Easily recognised forms of collagen.
- They form banded fibrils.
- Type I, II, III, V, and XI collagens belong to this group.
- The triple helical structure of these molecules contains
uninterrupted stretch of 338 to 343 Gly-X-Y triplets in each alpha
chain.
- The molecular weight of these collagens are of 95,000 kda.
-The molecule mesaures 15x 3000 A’ (Ramachandran and Rama
krishnan 1976;Piez 1976)

12. FACIT collagens:
- They are fibril associated collagens with interrupted triple
helices
- In these proteins the collagenous domains are interrupted by
non collagenous sequences and are found associated with the
surface of fibril forming collagens.
- It contains type IX, XII, XIV and XVI.
- The type IX, XII and XIV collagens are unique in containing
glycosaminoglycan components covalently linked to protein.

13. Non fibrillar collagens
-All other types forms non fibrillar collagens that do not form fibrils.
-Network forming collagens –Types IV, VIII and X.
-Beaded –Type-VI
-Anchoring fibrils-Type-VII
-Cuticle collagens-in invertebrates.
-These collagens form a sheets or protein membranes that encloses
tissues and organs.

14.  In addition to the above collagen groups at least ten
noncollagenous proteins incorporating short ,triple helical
collagen domains have been described
 This group of collagen domain containing non matrix proteins
includes the lung surfactant protein , acetyl choline esterases ,
and mannose binding protein .
 These collagen are not considered true collagen as they do not
form part of the extracellular matrix

15. TISSUE DISRIBUTION OF COLLAGEN
Type I
- in most tissues including bone, tendon, ligament, skin etc.,
-Accounts for 65% to 95% of the total collagen.
-Provides tensile strength
Type II, IX, X, XI
-mostly associated with cartilage and vitreous humour.
-Type IX is associated with type II and helps the cartilage to function at low
friction.

16. TISSUE DISRIBUTION OF COLLAGEN
 Type III
- mostly in fetal and immature tissues.
-Always associated with type I and found in many tissues.
-More fibrillar and extensible than type I and traditionally called as reticular
fibers.
-Second major species and the proportion ranges from 5 to 30% in adult
tissues.

17. TISSUE DISRIBUTION OF COLLAGEN
 Type IV
- in basement membrane forming chicken wire network.
-It has interrupted non collagenous sequences at the ends and also in the
middle of the structure allowing it to be susceptible to all proteolytic enzymes.
-They are stable to the action of collagenases.
-They also bind to angiogenic and osteoblastic factors presenting them in
immobilized status to initiate and sustain angiogenesis and osteogenesis.

18. TISSUE DISRIBUTION OF COLLAGEN
Type V- Found in most tissues associated with type I.
Type VI - Forms beaded microfibrils and found in most tissues.
Type VII- Anchoring fibrils that attaches the anchoring plaque with the basement
membrane thus maintaining the integrity between dermis and epidermis.
Type VIII - In the descemet’s membrane and endothelium.
Type XII to XVI - In many tissues.

19. TISSUE DISRIBUTION OF COLLAGEN
Type XVII- In hemi desmosomes of the skin.
Type XIX - In rhabdo myosarcoma cells.

20. BIOSYNTHESIS OF COLLAGEN
Synthesis of pro  chains , their assembly to pro collagen and their conversion
into collagen fibers involves several well coordinated biosynthetic reactions
occurring in the nucleus, cytoplasm and the extracellular space.
The triple helical collagen molecule is not accessible to the modification
enzymes. Therefore the modifications can occur only on the nascent α
chains.
Due to these reasons, the collagen molecule is first synthesized as a large
precursor(pro-α chain) containing extra amino acids at both N and C
terminal ends

21. Gene expression:
-Collagen genes are large and range in size from 5kb to 130 kb.
-More than 30 genes has been described for the different collagen types.
Fibril Forming collagen genes:
Contains 42 exons for the major triple helical region.
The exons are composed of 54 bp and start with an intact codon for glycine
They are separated by introns that are 80 to 2000 nucleotides long .

22. Non Fibrillar Collagen gene:
-Organization of gene varies .
eg Type IV collagen gene the exons do not follow the 54 bp rule.
Type IX collagen gene the size of the exon ranges from 21 to 400 bp
Type X collagen gene the region that encode the tripple helical domain consists
of a single 2136 bp exon.

23. Collagen gene
↓
mRNA
↓
Pro α chains
↓
Collagen
↓
Collagen Fibers

24. In nucleus:
Collagen mRNAs are transcribed as precursors and undergo the
usual processing of
- Capping,
- Polyadenylation and
- Splicing.

25. TRANSCRIPTION
 The process of transcription involves the generation of a single stranded RNA using a DNA
template.
 It involves the three steps
1 Initiation
2 Elongation
3 Termination
 Occurs in both prokaryotes and eukaryotes
 Prokaryotes possess only one type of RNApolymerase
 Three classes of RNA are transcribed:
1.mRNA(messenger RNA)
2.rRNA(ribosomal RNA)
3.tRNA (transfer RNA) and
other small RNA species

26. Eukaryotes possess three RNA polymerases:
RNA polymerase I,II,III
 RNA polymerase I transcribes rRNA.
 RNA polymerase II transcribes mRNA and tRNA.
 RNA polymerase III transcribes other small RNAs.

27. TRANSCRIPTION:
 Starts with the binding of a molecular complex of RNA polymerase and other
accessory factors to the DNA segment.
 The binding locally separates DNA strands and exposes the template
sequence to be transcribed to RNA polymerase
 The transcription is initiated by the binding of σ subunit of RNA polymerase,
it is released after the polymerization of about 10 ribonucleotides.
 Then the core polymerase( made up of α,β,ω) subunits take over and
completes the transcription.

28. Initiation

29. CAPPING:
 After the synthesis of 30 nucleotides the RNA transcript is modified at its 5’ end by
capping.
 5’-end of the mRNA is capped 5’ to 5’ with a guanine nucleotide by the enzyme
guanyl transferase.
 Capping appears to be necessary for recognition of mRNA by the nuclear
transport apparatus.
 Capping contributes mRNA stability by protecting the molecule from
degradation by nucleases that attack RNA at 5’ end.
 The cap also plays an important role in positioning mRNA on ribosome for
initiation of translation

31. TERMINATION:
 The RNA-polymerase stops moving on the DNA template. The
RNA transcript falls off from the transcription complex.
 Newly made transcripts become immediately associated with
proteins to from heterogenous nuclear ribonuclear protein
particle.

32. Processing of transcription
products :
Transcription products undergo modifications like removal of introns ,addition of
bases or chemical modifications of certain nucleotides
POLYADENYLATION:
-50-250 adenine nucleotides are added to 3’ end of mRNA.
-Polyadenylation generates poly(A) tail and is carried by the enzyme poly(A)
polymerase in a two step reaction (Whale and Keller 1996).
-Stabilizes the mRNA, and plays an important role in transcription termination

33. SPLICING:
- The process of removal of introns ( non coding genes) is
known as splicing.
- Introns are the nucleotide sequences in mRNA which do not
code for protein.
- Exons of mRNA posses genetic code and are responsible for
protein synthesis

34.  Introns start with the sequence GU at their 5’junction with their preceeding
exons and end with AG at the 3’end site.
 Splicing requires the formation of a splicesome complex where the actual
splicing takes place
 Splicesome is formed by the association of small nuclear ribonuclear protein
particles with several protein splicing factors with the splice site in the
precussor RNA.
 Splicing offers a mechanism by which several isoforms of a single protein can
be produced by one mRNA

35. Transport of the mRNA to the
cytoplasm:
 After nuclear event, mRNA forms complex with proteins and then is
transported to the cytoplasm as mRNPs through nuclear pores.
 After reaching their destination, the mRNPs disassociate releasing their
protein components.
 The final location of mRNA may be perinuclear, near plasma membrane, or
in any other geographical location within the cytoplasm

36. Translation:
Conversion of a messenger RNA sequence into the amino acid
sequence of a polypeptide (i.e., protein synthesis).
Translation occurs in different steps resulting in the
formation of pre pro-α chains.
1. Activation of AA
2. Initiation
3. Elongation
4. Termination

37. ACTIVATION OF AA
 Polypeptide synthesis takes place in ribosomes located either free in the
cytoplasm or bound to membranes in rough endoplasmic reticulum(RER).
 The ribosomes are compact ribonuclear protein particles consisting large and
small subunits of 60s and 40s in eukaryotes.
 Each ribosomal sub unit is made up of many proteins associated with rRNA.
 A ribosome attaches to the mRNA molecule at the 5” end and moves along in
the direction of 3” direction.
 In this route it translates triplet codons into corresponding amino acids.
 Translation begins at an AUG intiation site and peptide synthesis proceeds
from the amino acid to carboxyl end.
 GTP is an important co factor necessary for translation reactions

38.  Several protein factors play a role in the translation and they are classified
based on their functions they are classified as initiation, elongation,
termination or release factors
 The amino acids for polypeptide synthesis are presented as aminoacyl-tRNAs.
 Each tRNAs contain a three nucleotide anti codon sequence complementary
to the corresponding codon for the particular amino acid it specifies.
 There are 64 possible codons and one amino acid may be specified by one or
more codons

41. ELONGATION:
 Factors needed for the peptide chain elongation are EF1A, EF1B and EF2.
 These protien factors form a complex with GTP and aminoacyl-tRNA.
 A peptide bond forms between the first methionine, which is present at the P
site and the aminoacyl –tRNA at the A site.
 Formation of the peptide bond is catalyzed by the enzyme peptidyl transferase.
 Once a peptide bond is formed the ribosome moves to the next codon,
transfering the pepitdyl tRNA to the P site and releasing the deacylated tRNA.
 This sequence is repeated as the ribosomes move along the mRNA ín the 5’-to-
3’direction

42. TERMINATION:
Termination of the peptide elongation occurs when the ribosomes recognize the
termination codons UAG,UAA or UGA .
Termination involves releasing the completed polypeptide chain from the tRNA
and the free tRNA from the ribosomes.
Many ribosomes work on same mRNA molecule simultaneously and these
aggregates are called POLY RIBOSOMES or POLYSOMES.
This process requires proteins known as release factors.
In bacteria about 15 aminoacids are added per second at 37⁰c .
In eukaryotic cells it is at the rate of 2 aminoacids per second
Translation has a very low error rate of 1 in 10⁴ amino acids.
Initiation factor eIF2 is one of the key regulators of overall eukaryotic protein
synthesis

43. POSTTRANSLATIONAL PROCESSING AND
PROTEIN TARGETING:
TARGETING OR SORTING:
Placement of newly synthesized proteins in target cell organelles is achieved
either by synthesis at the required location as is seen in mitochondria and
chloroplasts or by translocation if proteins are synthesized elsewhere.
The movement of proteins from the site of synthesis to a different destination is
referred to as target or sorting.
Proteins targeted to nucleus are instructed to move this site by a 7 to 9 basic
amino acid sequence.
Ser-Lys-Leu triplet sequence at the carboxyl terminal of proteins such as catalase
places the protein in the peroxisomes.
A sequence of aprox 25 amino acids provides the signal for the polypeptide to
traverse through membranes in the chloroplast and mitochondria.

44.  Once the protein passes through the membrane this sequence is cleaved off.
 Proteins destined to be retained within the membrane contains this sequence
followed by another stretch of uncharged amino acids.

45. TRANSLOCATION:
 Movement of polypepetides across membranes of organelles and RER is
known as translocation.
 In eukaryotes the secretory protein are synthesized at the RER.
 The amino terminus of these proteins contains a signal sequence consisting of
15 to 30 hydrophobic and one or more positively charged amino acids.
 When ribosomes initiate protein synthesis this signal sequence binds to a
signal recognition particle (SRP).
 SRP bound ribosomes attaches a receptor on the ER membrane.
 Signal sequence attached to the polypeptide enters the ER membrane
 SRP then disassociates and the nascent polypeptide enters into the ER lumen

46. Protein Folding:
Folding of nascent polypeptides into three dimensional conformations proceeds
from the amino to the carboxyl ends.
This process is assisted by accessory proteins and di sulfide linkages.
The formation of disulfide linkages is catalyzed by the enzyme protein di sulfide
isomerase.
Chaperones: Set of proteins that bind with nascent polypeptides translocated into
the ER lumen, facilitate their transport and resist their degradation.
They aid in folding of proteins and also prevent misfolding of proteins under
normal conditions and during heat shock

47. CLEVAGE OF SIGNAL SEQUENCE:
 The mRNAs are translocated to the ribosomes in the cytoplasm that is
attached to RER.
 Signal sequences are cleaved during polypeptide elongation as nascent pro
alpha chain are being transported into rough endoplasmic reticulum.
 As translation procedes , certain proline and lysil residue at Y position are
oxidised to HYP & HYL by enzyme prolyl 4 hydroxylase.
 The cofactors needed for these reactions are vit-C, oxygen, iron and alpha
keto glutarate.

48.  Prolyl hydroxylation is essential for the thermal stability of
collagen.
 The melting temperature of fully hydroxylated collagen is
39°C and that of under hydroxylated collagen is 25°C.
 Lysyl hydroxylation is important because glycosylation takes
place in these residues.

49. Glycosylation
 It takes place in the Hydroxy lysine (Hyl) and Asparagine
(Asn) residues with oligosaccharides like glucose, galactose in
Hyl and mannose in Asn respectively.
 This glycosylation shields the collagen molecules from the
action of other proteolytic enzymes.
 As soon as synthesis of pro alpha chain is completed , 2 pro
alpha 1 ( I ) chain and one pro alpha 2 (I), in type 1 or 3 pro
alpha 1, in type II AND III associate at the C- terminal ends
and the association is stabilised by inter change di sulfide
bond formation catalysed by enzyme ( PROTEIN DISULFIDE
ISOMERASE)

50.  Association of  chains takes place near the C- terminal ends
and proceeds to N- terminal in the form of a zipper.
 Pro collagen molecule thus assembled is then translocated
into the golgi complex where additional glycosylation,
phosphorylation and sulfation takes place depending on the
type of collagen.
 These pro collagen molecules are packaged into vesicles,
which fuse with cell membrane and release their content into
the extracellular space.

51. IN EXTRA CELLULAR
PHASE:
 First the Propeptide sequence at N-C terminal ends are
removed by N-C procollagen peptidases, also known as N-C
proteinases.
 The N-propeptidases cleaves the proline glutamine bond in
pro alpha I and pro alpha III and alanine glutamine bond pro
alpha 2(I).
 Propeptide are removed from type I, II, III procollagen.
 Fibrillar collagen formed in this manner aggregates
spontaneously as ordered fibril due to presence of charged
hydrophobic region on the surface.

52.  Aggregation occurs in parallel overlapping lateral array such
that adjacent molecule are staggered by approx ¼ th of the
molecule.
 This leads to quarter stagger arrangement which permit
interaction with side chain of neighbouring molecule which
gives tensile strength to the fibres .
 Aggregation is nonenzymatic but regulated by presence of
other types of collagen.

53.  Removal of the propeptide extension is necessary for ordered
fibril formation.
 If retained at N-terminus, it leads to poor apposition of
collagen molecule – prevents normal fibril growth and fibril
formed are thinner and disorganised.
 Defects is known as DERMATOSPARAXIS – an inherited
disease seen in cattle and sheep.

54. STABILISATION OF
COLLAGEN FIBRILLS:
 The collagen fibrillar arrays are stabilized by covalent cross links .
 This crosslinking requires enzyme lysyl oxidase.
 The enzyme converts lysine and hydroxy lysine residues to allysine
and hydroxyallysine which then condenses with norleucine and
hydroxy norleucine forming divalent cross links.
 They are subsequently converted into more complex cross links,
merodesmosines and hydroxy pyridiniums.
 Cross linked collagen molecules then combine with other
extracellular matrix to form a three dimensional scaffolding.

55.  Ultimate organisation of collagen fibers depends upon the
specific functional demand of each tissues and presence of
minor collagens and proteoglycans like decorin.

56. REGULATION OF COLLAGEN
BIOSYNTHESIS:
 As collagen is the main structural component of the connective tissue the
quantity and proportion of collagen within each tissue need to be precisely
regulated to maintain tissue integrity.
 The most important step at which regulation of collagen production occurs is
at the gene transcription level. By the action of promoters and enhancers
elements.
 The magnitude of collagen synthesis is dependent upon the levels of mRNA
for its α chains .

57. REGULATION OF COLLAGEN
BIOSYNTHESIS:
Those that upregulate the synthesis,
- PDGF, TGF-, FGF, IGF.
Those that down regulates the synthesis are,
- Cytokines like IL-1, IFN , TNF-
- Hormones like glucocorticoids
- Others like PGE 2

58.  TGF- elevates the level of mRNA by increasing there stability.
 Collagen gene is downregulated by the inflammatory cytokine TNF
α and by the lymphokine interferon gamma IFN-ϒ.
 Growth factors and cytokines affecting collagen synthesis are
secreted by platelets, macrophages, keratinocytes and fibroblast
during inflammation, and their presence at the foci of inflammation
and wound sites generally correlates with changes in collagen
synthesis

59. DEGRADATION AND
REMODELLING:
 Degradation of collagen and other matrix elements is a key component of
normal tissue remodeling.
 It also contributes greatly to pathologic alterations.
 Matrix degradation is a major cause of connective tissue degradation in
chronic periodontitis, rheumatoid arthritis and other inflammatory diseases
whereas hyperplasias and fibrosis are associated with low levels of
degradation.
 Collagen degradation takes place in two different ways,
Intracellular degradation
Extracellular degradation

60. Collagen degradation is primarily mediated by the collagenases.
Collagenases are specialised enzymes that have evolved specifically to hydrolyze
collagen because their triple helical structure is resistant to most common
proteinases.
The collagenases belong to a family of enzymes called matrix metalloprotienases MMP.
They have 13 member with closely related domain structure and discrete functions.
All have 21 –kd catalytic domain that contains a Zn++ binding site.
Based on the substrate specificity they have been classified as
-Collagenases,
-Gelatinases,
-Stomelysins ,
-Matrilysins.

61. Intracellular
degradation:
fibroblast that are in intimate contact with the fibers
identify the alterations in the structure of collagen
cleave the fibrils and phagocytose of fibrils
forming phagosome then the phagolysosome
further degraded by the lysosomal enzymes especially cathepsin.

62. Extracellular
degradation:
 The altered integrity of the extracellular matrix causes
increased water to enter the collagen bundles causing
loosening of the bundles thus allowing the cells that secrete
degrading MMPS to enter the bundles.
 Collagenases :
 Type 1- MMP 1(fibroblast type)
 Type 2 – MMP 8 (neutrophil type)
 Type 3 – MMP 13 (found in breast cancer)
 Type 4 – MMP 18 ( unknown

63.  Gelatinases:
A – MMP 2
B – MMP 9
 Stromelysin:
Type 1 – MMP 3
Type 2 – MMP 10
Type 3 - MMP 11
 Matrilysin: - MMP 7
 Membrane type MMP’s: MMP 14, 15, 16, 17.
 Enamelolysins: MMP 19, 20

64.  The collagenases normally degrades collagen types I, II, III,
VI, VIII & X.
 Collagenase I also called as MMPI or fibroblast type
collagenase is produced by a variety of human epithelial and
mesenchymal cells including keratinocytes, fibroblast and
macrophages .
 This enzyme can hydrolyze type I, II, III, VI, VIII & X.
collagen & gelatin.
 It hyrolyzes type III collagen molecules faster than it does
typeI.

65.  Collagenase -2 also called Polymorphonuclear leukocyte
(PMN) type or MMP -8 is found only in the granules of
polymorphonuclear lymphocytes.
-It hydrolyzes type I & III collagens.
Collagenase -3 is a recently described enzyme found in breast
cancer

66. The action of MMP is controlled by 3 different mechanisms.
1ST MECHANISM
 These enzymes are present in tissue as inactive precursors
and conversion to an active form requires activation by
plasmin , trypsin or other proteinases.
 These are activated when the condition demand (eg During
inflammation ) and they are also regulated by other proteins
such as tissue plasminogen activator.
Inflammation →Tissue Plasminogen
Factor→Plasmin,Trypsin(proteinases)→MMP

67. 2nd mechanism
 Through modulation of synthesis, IL-1,TNF-alpha,PGE2
increase MMP production from both keratinocyte and
fibroblasts.
 TGF-beta also increase MMP production in fibroblasts but
decreases in keratinocytes.in macrophages MMP production
is stimulated by LPS,but decreased by IFN-gamma ,IL-4,IL-
10.

68. 3rd mechanism
 The activities of MMPs are neutralised by inhibitors present in
the serum like alpha 2 macroglobulin the binds to MMP1 and
inhibits MMP1.

69. BACTERIAL COLLAGENASE:
 In contrast to vertebrate enzymes, bacterial collagenases degrade host
collagen molecules to small peptides.
 Theses enzymes are highly potent and have broad specifications and they can
degrade all native collagens fibrils regardless of the collagen type.
 They do not have substrate specificity as host collagenases.

70. Diseases Associated With
Collagen Alterations:
Most tissues contains a mixture of collagen types and damage in their structure,
content, or proportion can lead to functional abnormalities of these tissues.
They can be broadly classified as
1.Inherited diseases.
2.Acquired diseases.
3 Types of alteration can affect collagens and lead to connective tissue changes:
1. Defect in the structure of collagen genes.
2. Molecular defect in the processing enzymes .
3. Mechanisms affecting the expression of collagen genes due pathologies of
acquired disease.

71. INHERITED DISEASES:
 Due to point mutations, deletions or insertions in the structural genes for
collagens or their post translational processing enzymes.
 Severity of diseases varies depending on several factors.
 For type I collagen, mutations that affect the assembly of proα1 chains can
be lethal, whereas those affecting proα2 are not.
 This is because the homotrimers of α1(I) formed in the absence of proα2(I)
are stable at physiologic temperature whereas the α2 (I)3 formed in the
absence of α1(I) is not stable and gets degraded.
 Therefore large deletions, insertions or mutations near the C-terminus of
pro-α1(I) which affects its assembly into collagen are often lethal.

72.  Replacement of glycine with other aminoacids decrease the rate of collagen
folding, its thermal stability and secretion therefore they are retained
intracellularly longer and degraded.
 Mutations associated with genes of collagen processing enzymes are usually
not lethal, but they lead to functional abnormalities of respective tissues.
 Almost 200 mutations have been characterized in COL1A1 and COLIA2
genes.

73. INHERITED DISEASES DUE TO
COLLAGEN ALTERATIONS
 Osteogenesis Imperfecta.
 Ehlers Danlos Syndrome.
 Chondrodysplasia.
 Alport’s Syndrome.
 Epidermolysis Bullosa

74. Osteogenesis Imperfecta: (Brittle
bone disease)
A heterogenous group of disorders associated with
- bone fragility
- dentinogenesis imperfecta
- hearing loss
- blue sclera and soft tissue dysplasia.

75. Classified into 4 types:
Type 1 :
C.F: Autosomal Dominant.
Characterised by blue sclera but normal teeth and near normal
stature.
Pathogenesis:
Due to mutations that decrease Type I collagen production,
Substitution of Glycine by cysteine near the N -terminal end.
Affected Gene:COL1A1

77. Type II:
Lethal in the perinatal period and is associated new dominant mutations.
Pathogenesis:
Due to mutations that affect the procollagen assembly by the substitution of
glycine near the C –terminal end
Rearrangemnet of multiple exons leading to loss of large segment of amino acids
Mutations at the C terminal propeptide of proα(1).
Affected Gene:COL1A1, COLIA2

78. TYPE III:
CF: Autosomal Dominant
Progressively deforming variety.
Pathogenesis:
Mutations in proα1(I) and proα2(I) that prevents its inclusion in
the collagen molecule.
Affected Gene: COL1A1, COLIA2

79. TYPE IV:
CF: Autosomal Dominant
Dentinogenesis Imperfecta.
Pathogenesis:
Mutation in α2(I) and small deletions in α2(I) gene
Affected Gene: COL1A2, COLIA1

80. Ehlers- Danlos syndrome
(Rubber Man Syndrome):
Hetrogenous group of connective tissue disorder charecterized by skin fragility
and hyperextensibility and hypermobility of joints.
EDS types I-III
CF: Autosomal dominant hereditary disorder whose severe form exhibits this
triad of symptoms:
– “Rubber skin” ( A): Large folds of skin can be lifted off the subcutaneous
tissue; minor injuries produce skin tears (dermatorrhexis).
– “Rubber joints”: Joints are hypermobile and are easily dislocated due to lax
ligaments.
– Childhood history of weak connective tissue:Manifestations include inguinal
hernia, intestinal diverticula, and rectal prolapse.

81. Pathogenesis:
Abnormal transcription of the type I procollagen chains causes deranged
fibrillogenesis. This in turn produces collagen fibrils whose cross section
resembles a bear paw (“bear-paw” collagen)

82. EDS TYPE IV: (vascular or
ecchymotic type)
CF:
Autosomal dominant hereditary disorder in which tissue tears like wet toilet
paper,
characterized by quartet of symptoms:
– Vulnerable skin (dermatorrhexis).
– Aneurysm.
– Alveolar tearing (spontaneous pneumothorax).
– Hollow organ ruptures in the genital and intestinal.
Pathogenesis: Mutation or multiple exon deletions within COL3A1 gene

83.  EDS TYPE VI(OCCULAR):
 CF:Blindness
 Pathogenesis: Lysl hydroxylation is severely depressed and
lysine is under hydroxylysed in Type I and Type III collagens.

84. EDS TYPE VII(A &B):
CF: Autosomal dominant hereditary disorder leading to joint
dislocation (usually the hip); the skin is rarely involved.
Pathogenesis: Impairment of pro- collagen to collagen
conversion .Assembly of collagen onto functional fibrils is
affected due to hindrance by the retained pro-peptide domain
.
Affected Gene: COLIAI,COLIA2(Mutations at the N –
propepatidase cleavage site exon 6 of COLIAI or COLIA2

85. EDS TYPE VIIC (Dermatosparaxis): Enzyme N –protienase itself
could be affected in this condition.
EDS TYPE VIII(PERIODONTAL):
CF: Autosomal dominant
Characterised by periodontal involvement, Loss of teeth.
Underlying Molecular cause is not yet identified.
EDS TYPE IX(cutis laxia or occiptal horn syndrome):
Reduced activity of lysyl oxidase due to defective intracellular
distribution of copper a co factor for lysysl oxidase.
Affected Gene: ATP7A

86. CHONDRODYSPLASIAS:
Approximately 50 mutations that leads to this condition has been reported.
CF: Heterogenous group of disease.
Abnormal growth and development of cartilage.
Exhibit abnormalities in type II collagen- containing tissues such as growth
plates ,nucleus pulpous and viterous humor.
3 major forms of this disease :
Achondrogenesis/Hypochondrogenesis
Spondyloepiphyseal dysplasia
Stickler Syndrome.

87. ALPORT SYNDROME:
 Progressive hereditary nephritis associated sensoneuronal deafness and
occular abnormalilties.
 It has an X linked inheritance pattern.
 Characterized by splitting and thinning of the glomerular basement
membrane.
 Pathogenesis:
 Point mutations in triple helical and C-propeptide domains.
 Multiple exon deletions
 Affected Gene: COL4A5 GENE

88. Dystrophic epidermolysis bullosa:
Molecular defects is seen in type VII collagen gene. In this disease the dermal
epidermal integrity is affected due to abnormal or absent anchoring fibrils.
Pathogenesis:
Mutations in COL7AI gene.

89. Marfan’s Syndrome:
This is the most common hereditary group of connective tissue disorders.
Manifestations include ocular, skeletal, cardiovascular, and dural lesions.
There is defect in organisation of collagen.
There is more amount of soluble collagen.
Symptoms : Skeletal
dolichostenomelia (long, narrow extremities);
scoliosis (abnormal spinal curvature);
pectus excavatum (deeply depressed sternum);
flat foot (pes planus) due to the medial deviation of the medial malleolus

90. Cardiovascular lesions:
-Aortic aneurysm : The media of the aorta is thinned and atrophic. Its elastic
fibers are thin, fragmented, and pushed apart by focal deposits of
proteoglycan As a result, the aorta appears weakened and is susceptible to
rupture. This means that the aortic rupture often occurs at several sites over
time in the form of a recurrent dissecting aneurysm Fatal hemorrhage may
result.
-Aortic valve insufficiency.
- Mitral valve prolapse may occasionally occur.
Dural ectasia: This may occur in the lumbosacral region.
Dislocation of the lens: deficient zonule fibers may cause ectopia lentis,
resulting in impaired vision.

91. ACQUIRED DISEASES:
Acquired diseases are much more common than inherited.
In these disorders, the gene expression is affected, not the gene structure.
The underlying cause of the disease is unknown; the diseases are caused by
variety of factors and all matrix components including collagens and
proteoglycan are affected.
Examples include chronic inflammatory diseases like rheumatoid arthritis,
periodontal diseases, Crohn’s disease, progressive systemic sclerosis, lupus
erythematosis, sjogrens syndrome etc., and also fibrosed conditions like
atherosclerosis, pulmonary fibrosis etc.,

92. COLLAGEN VASCULAR DISEASE:
-Autoimmune diseases that involve formation of immune complex due to autoreactive
antibodies and are associated with fibrinoid collagen necrosis and collagen deposits.
collagen vascular diseases include:
— Systemic lupus erythematosus
— Progressive systemic sclerosis
— Dermatomyositis
— Nodular panarteritis
— Sjogren’s syndrome
— Wegener’s granulomatosis
— Pseudo-lupus erythematosus;
— Combined connective tissue disease (Sharp’s syndrome) with lupus erythematosus,
scleroderma,and dermatomyositis;
— Rheumatic fever
— Rheumatoid arthritis

93. Rheumatoid Arthritis:
Systemic inflammatory autoimmune disorder primarily attacks the synovial
membrane of the minor joints in successive episodes, leading to joint
stiffening.
Pathogenesis:
Occurs in patients with a genetic predisposition (HLA-DRB1 0401, 0404
and0101).
Initial viral infection somehow stimulates autoreactive Th1 cells.
This generates autoantigens, This causes B cells to form autoreactive
antibodies(rheumatism factors).
-anti-idiotypes (primarily IgM against IgG) ,
-antinuclear antibodies, and
-anti-collagen type II antibodies

94. These antibodies cross-link to form deposits on collagen fibers (fibrinoid
necrosis), leading to formation of rheumatoid granulomas.
This activates the complement system which perpetuates the inflammation.
It affects the TMJ causing loss of vertical dimension. Indirectly have perio
complications like dys functionally excessive occlusal forces in the molar
regions and loss of bone support from extrusion of the anterior teeth.

95. Sjogren’s syndrome:
 Systemic autoimmune disorder with symmetrical involvement of the salivary
and lacrimal glands and desiccation of the conjunctiva and oropharynx.
 Clinical triad of – xerostomia, keratoconjunctivitis sicca, rheumatoid
arthritis.
 Primary form involves an isolated keratoconjunctivitis sicca syndrome.
 Secondary form involves a keratoconjunctivitis sicca syndrome in conjunction
with other autoimmune disorders such as rheumatoid arthritis or systemic
lupus erythematosus.
 Pathogenesis: Patients with a genetic predisposition (primarily HLA-DR3)
form autoreactive antibodies mainly against splicosomal proteins as well as
against the excretory epithelia of the salivary and lacrimal glands.

96. Systemic Lupus erythematosis
 is a immunologically mediated inflammatory condition characterized by
production of auto antibodies and causing multi organ damage.
 Pathogenesis: antinuclear antibodies (= ANA) are primarily directed against
double-stranded DNA (anti-ds-DNA), nuclear ribonucleic protein.
 This leads to complement activation necrotizing immune complex vasculitis
(lupus arteritis), and, in the terminal stage, “onion-skin” arteriopathy.
 CF: Common in women than men,
between the ages of 20 and 50.
 Butterfly erythema: Skin inflammation on the cheek side of the nose and over
the zygoma.
 Libman-Sacks endocarditis: Rough wart-like deposits of fibrin on the margins
of the mitral and tricuspid valves increase the risk of embolism.

98. Progressive systemic sclerosis:
Systemic immunologic disorder that begins with progressive sclerosis of the
dermal connective tissue and spreads to the vascular connective tissue of
internal organs.
CF: There is progressive fibrosis of skin and mucosa and limited jaw opening
Pathogenesis: The disorder is characterized by two mechanisms:
— Formation of immune complex and, presumably induced by it,
— Excessive formation of abnormal collagen and microfibrils

99. Scurvy
Definition: Connective-tissue disorder associated with vitamin C deficiency.
Vitamin C is a cofactor of proline hydroxylase that maintains that substance’s
enzyme iron in the bivalent form.
Pathogenesis: Vitamin C deficiency causes proline hydroxylase insufficiency.
This interferes with glycosylation, which impairs collagen fibrillogenesis.
Increased fragility of the vessels leads to hemorraging

100. Diabetes:
Characterized by hyperglycemia, polyuria and polyphagia.
CF: Numerous clinical changes including chelosis, mucosal drying and cracking
, burning mouth and tongue ,diminished salivary flow.
In periodontium there is enlarged gingiva, sessile or pedunculated gingival
polyps, polypoid gingival proliferations, abscess formation, periodontitis and
loosened teeth.
Altered Collagen Metabolism:
Increased collagenase activity and decreased collagen synthesis is found in
individuals with diabetes.
Chronic hyperglycemia affects the synthesis, maturation and maintenance of
collagen and extracellular matrix.

101.  In hyperglycemic state elevated levels of blood glucose lead to pathological
biochemical processes such as glycation of protein-like collagens or lipids
and non-enzymatic oxidative destruction resulting in accumulated glycation
end products (AGEs).
 Collagen is cross linked by these AGE formation, making it less soluble and
less likely to be normally repaired or replaced.
 Advanced glycation end-products can directly affect normal protein function,
or indirectly act by reacting with receptors on the cell membrane of a variety
of cells.
 Cellular migration through cross linked collagen is impeded and tissue
integrity is impaired as a result of damaged collagen in tissues for a longer
period.
 Increased expression oh MMP 8 &9 is seen in diabetic patients
 As a result the collagen in the tissues of patient with poorly controlled
diabetes is aged and more susceptible to breakdown.

102. Distribution of collagen types in
the periodontium
Periodontium maintains the structural and functional integrity because of the
remodeling capacity of the connective tissue component.
The major connective tissue component is collagen and it’s tissue distribution is
as follows,
Gingiva:
Collagen are the most abundant biochemical component of gingival connective
tissue accounting for more than 60% of the total tissue protein.

103.  They are arranged in two different forms :
type I collagen - arranged as large dense bundles of thick fibers is
the major species.
type III collagen - Those that are arranged in a loose pattern of short
thin fibers mixed with a fine reticular network.
type V collagen - accounts for < 1 % of the total collagen.
type VI collagen - is seen in the micro fibrils throughout the lamina
propria.

104. type IV collagen - is present in the basement membrane in lamina lucida along
with laminin. It is also present in the internal basal lamina that serves as an
interface through which junctional epithelium is attached to the root surface.
The basal lamina of the epithelium is invested into the underlying
connective tissue through anchorage fibrils containing type VII collagen. The
attachment of epithelial cells to the basement membranes and tooth surface is
mediated via hemi desmosomes that contain type XVII collagen.

105. Periodontal ligament:
Majority of the collagen is type I, followed by type III and type V collagen.
Type VI collagen - micro fibrils
Type IV - basement membrane to which anchoring
fibrils are attached that contains type VII collagen.
Type XII collagen, is involved in the three- dimensional organisation of the
extracellular matrix. This collagen is restricted to the mature tissues.
Type XIV collagen is associated with the major collagen fibrils but not the
micro fibrils.
The rate of collagen turn over in the periodontal ligament is greater than any
tissue of the body, being twice than that of gingiva, five times that of alveolar
bone and fifteen times that of the skin.

106. Cementum:
Predominantly contains type I & type III collagen. The sharpey’s fibers do
not have type V & type VI collagens.
Alveolar bone:
Predominantly has type I & type III collagen. The collagen fibers attached to
bone,has
- larger diameter
- small in number
- less matured
- rapid turn over than the collagen attached to
cementum.

107. ALTERATION OF COLLAGEN IN
GINGIVITIS AND PERIODONTITIS Periodontal disease is characterized by destruction of gingival and
connective tissues.
 In gingivitis tissue destruction begins within the perivascular extracellular
matrix where most of the collagen within the foci of inflammation is
degraded.
 Quantitative and Qualitative changes occur in gingival collagen in patient
with the above disease.
 In gingiva collagen becomes more soluble(indicates new and active
synthesis).
 Type I & II collagen are destroyed at the foci of inflammation.
 The ratio of collagen types are altered.
 Type V collagen increases and its amount may exceed that of Type III.
 A new collagen species, Type I trimer can be detected in inflamed gingiva

108. Gingival Overgrowth:
Charecterised by increased gingival mass associated with fibroepithelial changes
and an accumlation of connective tissue matrix .
Type I/III ratio becomes different with loss of type I and elevated levels of type
III collagen.
Two mechanisms are involved .
1 .Reduction in the level or activities of matrix degrading enzymes in these
lesions.
2. Induction of collagen production may occur by direct action of drugs on
collagen synthesis mechanisms.
-by directly affecting the gene transcription rate
3.Indirectly through cytokines such as TGFβ

109. USES OF EXOGENOUS
COLLAGEN
Periodontal regeneration:
 The goal of periodontal therapy is to regenerate the periodontal tissues
affected by diseases to their original architectural form and function. Thus we
need regeneration of cementum, periodontal ligament and alveolar bone over
the diseased root surface.
 Earlier single layered collagen were used,but now they have been replaced by
bi-layered collagen membranes.
 These collagen are derived mostly from achilles tendon of bovine origin and
also from rat’s tail.
 Uses of these collagen are around infrabony defects, grade II furcation
involvement, peri implant defects, over exposed implant surface, even for root
coverage procedures.

110. Root conditioning:
 Mostly citric acid is used.
 collagen based putative cell binding PEPGEN P-15 peptide is used to
enhance fibroblast attachment and cell migration of periodontal cells to root
surfaces

111. As a haemostatic:
Haemorrhage from donor site in free gingival graft results in operative and
post operative complications. Collagen has been used for the arrest of
bleeding.
Microfibrillar Collagen Haemostat (MHC) is used to control surgical bleeding
both from oozing raw surface and also discrete finite bleeding from
transected or penetrated vessel.
Disadvantages – migration from the application site, adherence to wet
instruments, surgical gloves, mucosal tissues.

112. Sutures:
Surgical gut sutures:
Derived from submucosa of sheep intestine / serosa of beef cattle intestine. It is
99% pure collagen, resorbable being degraded by proteolytic enzymes.
Collagen sutures:
Made by extruding homogenized tendon achilles of beef cattle ,almost 100% pure
collagen, stiffer than surgical gut,used in eye surgery since it’s tissue reaction
is less than surgical gut.

113. References
 BIOLOGY OF PERIODONTAL CONNECTIVE TISSUE- BY P . MARK
BARTOLD & A .SAMPAT NARAYANAN
 ORAL CELLS AN TISSUE BIOLOGY BY GARRANT
 TEXT BOOK OF ORAL MEDICINE – BURKIT
 ORAL AND MAXILLOFACIAL PATHOLOGY- NEVILLE & ALLEN
 TEXT BOOK OF ORAL PATHOLOGY –SHAFERS