il dottor Spoliti Ortopedico illustra come curare con le Cellule mesenchimali, difetto condrale Ricostruzione con Acido Ialuronico e midollo osseo autologo Aspirare Concentrate

1. Chondral Defect Reconstruction with Hyaluronic Acid
Scaffold (HyaloFast®) and Autologous Bone Marrow
Aspirate Concentrate
M. Spoliti M.D., F.R. Rossetti M.D.
San Camillo Hospital – Rome,
ITALY
9/6/13
Satellite Symposium, Izmir Turkey
1

2. Articular Hyaline cartilage has a limited
regeneration capacity
In 1743 W.Hunter Stated that :
“from Hippocrates to the present age,
it is universally allowed that ulcerated
cartilage is a troublesome thing and
that when destroyed, it is not
recovered”

3. Autonomous hyaline cartilage defect regeneration is due
to infiltration of Mesenchimal Stem Cells (MSCs) from the
bone marrow , through perforation/microfracturing the
subchondral bone.
Unfortunately the reparative tissue consists of fibrocartilage.
This newly formed and less resistant tissue, could have a
breakdown expecially in young active competitive patients.
Minas & Nehrer 1997, Shapiro 1993
To produce cartilage of BETTER QUALITY,
as yet unidentified
FAVOURABLE CONDITIONS MUST BE CREATED.
REPAIR OF LARGE FULL-THICKNESS ARTICULAR CARTILAGE DEFECTS IN THE RABBIT
THE EFFECTS OF JOINT DISTRACTION AND AUTOLOGOUS BONE-MARROW-DERIVED
MESENCHYMAL CELL TRANSPLANTATION
T Yanai et Al , JBJS Br 2005

4. Articular Cartilage
(Buckwalter et al. 1994)
Composed of:
-Chondrocytes (1-10%)
-Extra-Cellular matrix
• Water (65-80%)
• Collagen (90-95% of type II)
• Proteoglycans
• Other matrix proteins and lipids
P.Motta” La Sapienza” University Rome
It is avascular, alymphatic, and aneural, nutrition through
diffusion from the synovial fluid.
Its lack of vascularity, high extracellular matrix to cell ratio, and lack
of progenitor cells, leads to its limited capacity to self-repair injuries.
3

5. Therapeutic Goals of Articular Cartilage
Repair
•
To resolve or at least alleviate the symptoms
•
Ensure the functional recovery of the movement
•
Restore the integrity of the articular surface
•
Prevent further deterioration of the tissue
(Jackson, Scheer et al., 2001)
The final goal should be to produce a repair tissue that has the same
functional and mechanical properties of the original hyaline
articular cartilage to prevent osteoarthritis.

6. Treatment procedures
(Nehrer, et al. 2000; Felson, et al., 2000).
Symptomatic procedures
•
Lavage, Shaving, Debridmen , Abrasion arthroplasty
Biological restoration: tissue repair/regeneration based on cells or living tissues
Repair procedures
•
•
•
•
Subchondral drilling
Microfracture
Osteochondral transplantation (OCT)
Periosteal or perichondral grafting
Regenerative procedures
• Autologous chondrocyte transplantation (ACT)
• One-step mesenchymal stem cells techniques
(MSCs)

7. Regenerative Techniques
•
ACI and MACI (Auotologous Cultured Chondrocytes)
•
New one-step techniques with scaffolds+MSC
The goal of these techniques is to restore the articular surface with a newlyformed hyaline-like tissue having physical, biomechanical and durability
properties as similar as possible to the native cartilage.
7

8. ACI and MACI (Auotologous Cultured Chondrocytes )
Transplantation of cultured autologous chondrocytes into the cartilage defects to
regenerate hyaline articular cartilage.
Two-step technique:
1st surgery: arthroscopic biopsy collection
2nd surgery: arthroscopic graft implantation
Good clinical results at long
term follow up, but…
Disavantages:
• two surgeries
• high costs
• Graft not immediatly
available
ACI 1st generation
MACI 2nd generation

9. One-step techniques with scaffolds + MSCs
The use of autologous bone marrow-derived MSCs in the treatment of
osteochondral lesions, represents an opportunity allowing, at the same time:
•the restoration of both cartilage and sub-chondral bone tissues
•AVOID A TWO-STEP surgical procedure (less invasive, lower cost)
The source of autologous MSCs can be:
1.bone marrow from subchondral bone at the lesion site (microfracture)
2.bone marrow aspirate, collected from the iliac crest, concentrated or not and
then applied on the defect site.
A limitation is that MSCs are simply released (1) or applicated (2) into the joint,
rather then being contained at the site of the defect.

10. Free MSCs?
Some papers found only a small number of MSCs in the
implant site after 10 days
(5% of the implanted BM-MSCs).
Guest DJ, Smith MR, Allen WR. Equine embryonic stem-like cells and
mesenchymal stromal cells have different survival rates and migration patterns following
their injection into damaged superficial digital flexor tendon. Equine Vet J 2010

11. MSCs NEED A SCAFFOLD FOR SUPPORT
• To stay and grow at the defect site
• To build a 3D structure of hyaline-like tissue
The goal is the appropriate scaffold

12. One-step techniques with scaffolds + MSCs
In order to stabilize the blood clot at the defect site, reasorbable
scaffolds are developed to enhance cartilage regeneration
The Ideal Scaffold
•
•
•
•
•
•
•
•
Resistant (Arthroscopic implant)
Tridimensional
Absorbing
Allowing good cellular adhesion
Enhancing MSCs proliferation
Biodegradable
Biocompatible
Promoting cell differentiation towards chondrogenetic and/or osteogenic
phenotype
• Handful

13. HYAFF 3D-Scaffold
•
•
•
•
13
Non woven felt, 2 mm thick, fiber diameter 10 microns.
Biocompatible
Bioresorbable
Main degradation product: Hyaluronic Acid

15. RESUMING
AUTOLOGOUS CELLS IMPLANT
Knee chondral defects:
• III - IV degree
(According to Outerbridge classification)
• area > 1-5 cm2
MACI
vs
BM - derived MSCs
• 2-steps surgery
• Single step surgery
• High costs
• Less expensive procedure
• No bony defect regeneration
• Bony regeneration
• Good results
• Results (?)
• Hyaline like cartilage

16. Our case series
•
•
•
•
163 chondral defect of the knee (III-IV degree)
Age range 15-51 y
Defect size
mean age 36 y
• 64 < 1 cm2
Last 4 years
• 99 >1 -5 cm2
•
•
•
39 MACI
17 MSCs
43 microfractures + HialoFast
>1 -5 cm2
•
•
51 microfractures
13 OATs
< 1 cm2

17. Study
Comparison MACI vs MSCs Implant
Inclusion criteria:
Exclusion criteria:
• Defect: III° - IV° degree
• Arthritis
• Width: 1- 5 cm 2
• Scheletrical malalignement
• Age range: 15-50 years old
• ACL-PCL tear
• Patellar instability
• Kissing lesions

18. Study
Comparison MACI vs MSCs Implant
Group 1 - 17 cases MACI (Two-step procedure ):
Mean age: 35,8 years (range15-49)
Gender: 10 M + 7 F
Defect location: 2 patella ,1 tib.plateau ,5 lat. condyle, 9 med. condyle
Mean follow-up time: 30,5 months (range 3 months to 4 years)
Defect Size: 6 Pts >1/= 2 9 Pts >2 =3 2 Pts >3
Group 2 - 15 cases (BMAC)+ Hyalofast:
Mean age: 31,9 anni (range19-42)
Gender: 11 M + 4 F
Defect location: 2 patella, 2 lateral condyle, 5 medial condyle, 2 troclea
Mean follow-up time: 10,1 months (range 5 to 26 months)
Defect Size: 3 Pts >1/= 2 7 Pts >2 =3 2 Pts >5

19. Study
MACI Implant Protocol
• 1st surgical step: cartilage biopsy collection
• Chondrocites in vitro expansion, seeding and
culture on 3D HA matrix (Hyalograft C autograft).
•
2nd surgical step: arthroscopic graft implantation

20. Study
MSCs implant protocol
• Harvest the bone marrow from postero-superior iliac crest,
with the patient in the lateral decubitus (60 mL of bone
marow).
• Process the collected bone marrow directly in the operating
room by removing erythrocytes and plasma by a cell
separator-concentrator consisting of a centrifuge and a
disposable double chamber.
• At the end of a 15 min centrifugation cycle, 7 mL of
concentrate containing nucleated cells (stem cells,
monocytes, lymphocytes, and other bone marrow resident
cells) are retrieved in the anterior chamber.
• 3ml/cm2 of BMAC can be loaded onto the scaffold together
with the PRP gel and thrombin; the matrix, due to its
hydrophilic properties, allows the homogeneous distribution
of the concentrate fluid rapidly.
•The pre-loaded scaffold
arthroscopic technique
can
be
implanted
by

21. Study
MSCs Implantation: Arthoscopic Technique

22. Study
MSCs Implantation: Large troclear defect

23. Study
Post-operative Rehabilitation
KEY POINTS
Immobilization: first 24 hours
Control Passive Motion (CPM): after 24 hours, for 4 weeks
Joint Loading: not allowed for about 6 weeks. 6th-10th week:
gradual recovery of the joint loading and of the step
Back to Sports:
Low impact : from 4th month (swimming, cycling)
High impact : from 10th month (running, soccer, tennis,
etc…)

24. Follow up evaluations
Clinical evaluations:
•IKDC subjective: pre-op, 5 months, 10 months
•Tegner score: 10 months follow up
•MRI assesment:
•all patients underwent MRI evaluation at 3- 6 -12 - 24 months
•Cartilage repair evaluations:
3 Pts group 1 and 3 Pts group 2
2° look arthroscopy with biopsy and histology

25. Clinical Results
Summary of MACI case series –
Group 1
Patient . Age
Follow-up
IKDC (pre-op) IKDC (5 m)
IKDC (10 m)
B.B.
S.A.
P.F.
P.M.
D.D.
V.M
L.D.
P.M.
P.L.
C.E.
P.A.
G.R.
B.B.
M.S.
A.G.
M.G.
D.S.
4Y
9M
3Y
2Y
2Y
10 M
1Y
1Y
4Y
2Y
4Y
4Y
4Y
3Y
2Y
11M
1Y
32
53
25
35
30
27
36
35
23
41
33
31
40
25
19
17
51
88
90
75
84
98
80
83
80
49
95
88
74
89
85
87
77
65
35
48
46
23
15
42
34
44
49
21
47
45
37
23
24
49
28
90
95
70
84
89
85
85
76
45
90
74
51
73
80
85
80
75
IKDC pre - op: 32,5
Poor
IKDC post- op 5 m: 78,5
Excel.
IKDC post- op 10 m: 81,6
Excel.
Tegner score (10
m)
7
7
5
9
9
7
9
7
4
8
6
5
5
7
9
6
4
TEGNER post op 10 m: 6.7

26. Clinical Results
Summary of MSCs + scaffold case series Group 2
Patient Age
Follow-up IKDC (pre-op)
IKDC (5 m)
IKDC (10 m) Tegner score (10 m)
B.A.
B.B.
C.M
S.L.
R.V.
P.M.
T.G.
M.D
R.M
S.G.
M.L
N.V.
G.O.
F.O.
M.N.
2Y
18 M
1Y
1Y
2Y
10 M
11 M
18 M
2Y
10 M
3Y
1Y
16 M
17 M
15 M
85
88
90
45
73
85
74
72
90
85
93
91
72
77
84
89
76
95
55
75
88
85
83
90
80
87
65
77
81
76
28
42
22
40
35
25
27
33
29
41
32
19
42
39
25
25
35
30
27
36
35
23
41
33
31
40
25
29
27
45
IKDC pre - op: 32,1
9
7
8
4
6
6
6
7
9
7
8
7
6
7
7
Poor
IKDC post- op 5 m: 80,2
Excel.
IKDC post- op 10 m: 80,1
Excel.
TEGNER post op 10 m: 7

27. MRI Results
Uniform post-operative NMR evolution :
• 3 months: subchondral bone edema important
3M
MSCs implant
• 6 months: substantial reduction in subchondral
edema
• 12 months: disappearance of edema
6 M MSCs implant
• In the following assessment, we have found the coverage of the areas of
chondropathy with integrity and restoration of the articular surface of the joint lining

28. MRI Results
24 months: there is a slight remodeling of 'subchondral bone,
that means a cartilage still in the remodeling process.
(in agreement with the results of Marcacci 2005).
MACI
MACI
MSCs implant

29. MRI Results
A
MSCs implant
B
MACI
C
MSCs implant
3M
E
MSCs implant
FF
24 M
D
MACI
12 M
MACI
G MSCs implant
H
3Y
MACI

30. MACI 2nd look
MSCs 2nd look
SECOND LOOK 24 MONTHS
SECOND LOOK 6 MONTHS
D

31. Conclusions
On the basis of the results of this ongoing study, MSCs
implantation via one-step regeneration procedure at
present is a viable and lower cost alternative compared
to two step techniques.
With confirmed long-term results, one-step technique
could eventually replace autologous chondrocyte
implantation.
Furthermore, the use of MSCs implantation allows us to
treat via a "one-step“ technique associated lesions,
such as meniscal or ligament tears .

32. BEWARE!!!
!!
In order to be effective in joint tissue regeneration, MSCs seem to need a
scaffold for :
•Improving bone marrow handling
•Favouring cells attachment and organization at the lesion site
•Stimulating MSCs differentiation into chondrocytes and proper reorganization
of the osteochondral compartment
Some PRP gel can add a supplement of growth factors to stimulate cell
differentiation and optimize implant stability.
The lack of a guide and containment in situ (osteo-chondral defect) leads the
potential of these cells to a "wild” regeneration pattern.

33. www.marcospoliti.com