Department of Orthopaedics

Polytechnic University of Marche

Ancona - Italy

A. Gigante

The role of Regenerative Medicine

• Articular environment (patient – pathology – grade – side/size)

• Scaffolds

• Cells

TISSUE ENGINEERING

• Biological environment (patient – pathology – grade – side/size)

• Scaffolds

• Cells

TISSUE ENGINEERING

* Natural polymers

SCAFFOLDS FOR CARTILAGE

• Hydrogels (agarose - alginate) Wang et al., 2007

• Chitosans Muzzarelli et al., 1995

• Synthetic materials (PLA - PGA – PDS)

• Human cartilaginous matrix

• Hyaluronic acid* Pavesio et al., 2003

• Collagen types I and II* Gigante et al., 2003

• Collagen I* + Hydroxyapatite Roveri et al., 1998

• Polysaccharides* (CS photocrosslinkable) Li et al., 2004

• Fibrin gel* Homminga et al., 1993

Biomimicry

BIOMATERIALS PROPERTIES

CELLS SCAFFOLD

BIOMIMIC SCAFFOLDS

• Collagen I membranes for tendon repair: effect of collagen fiber orientation on cell behavior. Gigante A et al. J Orthop Res. 2009 Jun; 27(6):826-32.

• Adult mesenchymal stem cells for bone and cartilage engineering: effect of scaffold materials. Gigante A et al. Eur J Histochem. 2008; 52(3):169-74.

• Engineered articular cartilage: influence of the scaffold on cell phenotype and proliferation. Gigante A et al. J Mater Sci Mater Med. 2003 Aug;14(8):713-6.

CARTILAGE EXTRACELLULAR MATRIX

COLLAGEN

• Chondrocytes in suspension covered by a

membrane (II generation);

• Chondrocytes grown on biological or artificial

scaffolds (MACI – III generation);

• Microfracture covered by a membrane

(AMIC);

• MSCs or buffy coat seeded in a biological or

artificial scaffolds.

SCAFFOLDS

• invasiveness

(mini-open approach or

arthroscopic method)

• operative time

• patient discomfort

Chondrocytes – microfractures – MSCs

grown on biological or artificial scaffolds

COLLAGEN AS SCAFFOLD

CELLS

CELLS FROM MICROFRACTURES

MESENCHIMAL STEM CELLS (buffy coat)

DIFFERENTIATED CHONDROCYTES

TWO STEPS

ONE STEPS

MACI® & NOVOCART® 3D

MACI type I, III collagen (porcine)

Safranin O S-100

protein

Cell density 106

mln/cm2

Gigante et al., 2006

Not controlled, prospective study,

mono and multicentric data

Clinical experience

D’Anchise R., Manta N., Prospero E., Bevilacqua C., Gigante A.

Autologous implantation of chondrocytes on a solid collagen scaffold after two years of follow up.

J Orthopaed Traumatol 6: 36-43, 2005.

Cherubino P., Grassi F.A., Bulgheroni P., Ronga M.

Autologous chondrocyte implantation using a bilayer collagen membrane: a preliminary report.

J Orthop Surg., 11(1): 10-5, 2003

• 59 consecutive cases (10 ♂ - 5 ♀)

• Median age 31

• Follow up 3-5 years

• 51 isolated – 8 multiple lesions

• 33 trauma and microtrauma

• 9 OCD

• 8 degenerative

• 0 connective disease

Patients and methods

Indications

Knee

2 cm2 – max 8 cm2 - mean 4 cm2

Ankle

1 cm2 – max 4 cm2 - mean 2 cm2

33 trauma and microtrauma - 9 OCD

8 degenerative – 0 connective disease

Patients and methods

Associated treatment:

• 9 ACL reconstructions

• 7 valgus osteotomies

• 9 p.f. distal realignments

Patients and methods

Distal realignment and patellar ACI: mid term results in a

selected population.

Gigante A, Enea D, Greco F,

Bait C, Denti M, Schonhuber H, Volpi P

Knee Surg Sports Traum Arthr 2009 ; 17:2-10

Isolated lesions

Multiple lesions

Degenerative lesions: “unshouldered lesions”

• MACI + tissucol + polar sutures

Patients and methods

14 MACI:

Grading III-IV Outerbridge

Size > 2cm2

Causes:

» 7 malallineaments

» 5 trauma

» 2 doc

Patella and throclea

Glued + sutured membrane Patella

Glued + sutured membrane

Patella

Patella: Glued + sutured membrane + TTT

Distal realignment and patellar ACI: mid term results in a selected population.

Gigante A, Enea D, Greco F, Bait C, Denti M, Schonhuber H, Volpi P

Knee Surg Sports Traum Arthr 2009 ; 17:2-10

Knee: maci by arthroscopy

Arthroscopic Delivery of Matrix-Induced Autologous Chondrocyte Implantation:

International Experience and Technique Recommendations

F. Cortese, M. McNicholas, G. Janes, S. Gillogly, S.P. Abelow, A. Gigante, N. Coletti

Cartilage, 2011, in press

6 ankle (talus) Gigante et al., JBJS, 2005

ANKLE Ankle: maci by arthroscopy

• Subjective knee function state

VAS (1,2,3,6,12 and 24 M)

• Objective finding:

1. ICRS-surgeon form part

(1: normal; 2: nearly normal;

3: abnormal; 4: severely abnormal)

2. Lysholm and Tegner scores at

baseline and after 6, 12 and 24 months

Materials and methods

“Modified ICRS score”

TIME

(MONTHS)

0 1 6 12 24 36

N PATIENT/

KNEE 5,42

±1,5

2,71

±1,4

1,03

±1,5

0,45

±0,8

0,21

±0,4

0,23

±0,4

VAS

FUNCTIONAL STATE

TIME

(MONTHS)

0 1 6 12 24 36

2.55±0.

51

* 1.12±0.

31

1.06±0.

27

1.00±0.

25

1.03±0.

25

00:00

01:12

02:24

03:36

04:48

06:00

preop. 6 months 24 months

VAS

Function

Clinical Results at 3 years

IKDC SCORE (from ICRS score)

TIME (MONTHS) 0 6 12 24 36

Functional

test

3.23

±0.45

1.04

±0.39

1.28

±0.37

1.00

±0.25

1.00

±0.02

Ligament

examination

1.17

±0.34

1.05

±0.27

1.07

±0.28

1.06

±0.28

1.06

±0.58

Passive

motion Deficit

1.06

±0.29

1.03

±0.23

1.00

±0.25

1.00

±0.25

1.00

±0.36

Effusion/

compartment

s findings

2.25

±0.49

1.26

±0.36

1.16

±0.33

1.04

±0.27

1.04

±0.17

Jamshidi needle mm. 3 Ø deep > 5 mm

2nd look and biopsy

• 12 - 24 months

• 22 cases fron multicentric national study

2nd look and biopsy at 12 months

MACI at medial condyle and ACL reconstruction

Feature/score 0 1 2 3

I. Surface Discontinuities/

irregularities

Smooth/

continuous

II. Matrix Fibrous tissue Fibrocartilage Misture:

hyaline/fibro

cartilage

Hyaline

III. Cell distribution Individual

cells/disorga

nized

Clusters Mixed/columnar-

clusters

Columnar

IV. Cell population

viability

<10% viable Partially viable Predominantly

viable

V. Subchondral Bone Detached/fractur

e/callus at

base

Bone

necrosis

Increased

remodelling

Normal

VI. Cartilage

mineralization

Abnormal/inappr

opriate

location

Normal

ICRS Visual Histological Assessment Scale

Membrane completely reabsorbed at 12 months

ICRS Visual Histological Assessment Scale

Feature/score 0 1 2 3

V. Subchondral Bone Detached/fractu

re/callus at base

Bone

necrosis

Increased

remodelling

Normal

VI. Cartilage

mineralization

Abnormal/inappr

opriate location

Normal

ICRS Visual Histological Assessment Scale

19/22

The histological data have confirmed that the regenerated cartilage is a typical hyaline cartilage, with a straight tide-mark with subchondral bone, well-organized cell-clusters and strong reactivity for hyaline markers.

MACI: conclusions

The cost efficiency

of this technique

remains to be evaluated

COLLAGEN AS SCAFFOLD

CELLS

CELLS FROM MICROFRACTURES

MESENCHIMAL STEM CELLS (buffy coat)

DIFFERENTIATED CHONDROCYTES

TWO STEPS

ONE STEPS

Cells from buffy coat + collagen membrane

ONE STEP

MERG

membrana per riparazione guidata

• autologous

• lower costs

• one step

• multi-potent?

ONE STEP

Buffy coat (marrow cells) + Collagen scaffold (MERG)

7 biopsies > 1 year

Gigante A. and Calcagno S.

MERG 7 biopsies > 1 year

Gigante A., Calcagno S., Cecconi S., Ramazzotti D., Manzotti S., Enea D.

Use of collagen scaffold and bone marow concentrate as a one step cartilage repair in the knee.

Int. J. Immunopathol. Pharmacol., 24 (suppl.2), 69-72, 2011

• Articular environment (patient – pathology – grade – side/size)

• Scaffolds

• Cells

TISSUE ENGINEERING

Thanks