ACI new technique for condral or osteochondral defect

1. AUTOLOGOUS CONDROCYTE
IMPLANTATION
MODERATOR : PRESENTED BY:
Dr. MAHESHWARAPPA D Dr. VIJENDRA YADAV
(PROFESSOR & UNIT CHIEF) PG IN ORTHOPAEDIC
DR. SHIVASHANKARAPPA DATE-1/3/17
(ASSO. PROFESSOR)

2. INTRODUCTION
• Autologous chondrocyte implantation
(ACI) for the treatment of articular
cartilage lesions of the knee joint
provides successful and durable long-
term outcomes.
• The treatment of symptomatic cartilage
defect is a challenging problem as
injuries of the articular cartilage do not
heal spontaneously.

3. • To address this, ACI was first performed in
Gothenburg, Sweden, in 1987 by Lars
Peterson, and his first cohort of cases with
second-look biopsies was reported in 1994.
• The first-generation ACI were performed with
an autologous periosteal flap from the tibia or
femur to cover the defect and seal in the
cultured autologous chondrocytes.

4. • Second-generation ACI involved the use of a
bioabsorbable scaffold carrier for the cells.
These scaffolds include a hyaluronan weave
(Hyalograft C; Fidia Advanced Biopolymers),
collagen scaffolds (MACI; Genzyme
Biosurgery) and gels derived from collagen
(CaReS; Arthro Kinetics).

5. • ACI is a two-stage procedure, beginning with
arthroscopic assessment of the chondral
injury and biopsy to harvest approximately
200 to 300 mg of cartilage, followed by a
commercial enzymatic digestion and cell
expansion in monolayer culture with
cryopreservation of the cells. The
cryopreserved cells are thawed and expanded
to the cell population needed for the second-
stage open ACI technique.

6. GENERAL FEATURES OF CARTILAGE
• Cartilage is derived (embryologically) from
mesenchyme.
• Some mesenchymal cells differentiate into cartilage
forming cells or chondroblasts.
• Chondroblasts produce the intercellular matrix as well
as the collagen fibres that form the intercellular
substance of cartilage.
• Chondroblasts that become imprisoned within this
matrix become chondrocytes.
• Some mesenchymal cells that surround the developing
cartilage form the perichondrium.
• Perichondrium contains cells that are capable of
transforming themselves into cartilage cells when
required.

7. • Cartilage has very limited ability for
regeneration (after destruction by injury or
disease). Defects in cartilage are usually filled
in by fibrous tissue.

8. • Cartilage is usually described as an avascular
tissue. However, the presence of cartilage
canals, through which blood vessels may enter
cartilage, is well documented. Each canal
contains a small artery surrounded by
numerous venules and capillaries.
• Cartilage cells receive their nutrition by
diffusion from vessels in the perichondrium or
in cartilage canals.

9. • Growth of cartilage: Newly formed cartilage
grows by multiplication of cells throughout its
substance.
• This kind of growth is called interstitial
growth. Interstitial growth, is possible only as
long as the matrix is sufficiently pliable to
allow movement of cells through it.
• As cartilage matures the matrix hardens and
the cartilage cells can no longer move widely
apart; in other words interstitial growth is no
longer possible.

10. • At this stage, when a cartilage cell divides the
daughter cells remain close together forming
cell nests.
• Further growth of cartilage now takes place
only by addition of new cartilage over the
surface of existing cartilage; this kind of
growth is called oppositional growth.
• It is possible because of the presence of
cartilage forming cells in the deeper layers of
the perichondrium

11. • The articular surface of most synovial joints
are lined by hyaline cartilage.
• The articular cartilages provide the bone ends
with smooth surfaces between which there is
very little friction.
• They also act as shock absorbers.
• Articular cartilages are not covered by
perichondrium.
• Their surface is kept moist by synovial fluid
that also provides nutrition.

12. COMPONENTS OF CARTILAGE
• Like ordinary connective tissue, cartilage is
made up of:
• Cells—chondrocytes
• Ground substance/matrix
• Fibres—collagen fibres

13. CARTILAGE CELLS
• The cells of cartilage are called chondrocytes.
• They lie in spaces (or lacunae) present in the
matrix.
• At first the cells are small and show the features
of metabolically active cells.
• The nucleus is euchromatic.
• Mitochondria, endoplasmic reticulum (ER) and
Golgi complex are prominent.
• As the cartilage cells mature they enlarge, often
reaching a diameter of 40 um
• The nuclei become heterochromatic and
organelles become less prominent.

14. GROUND SUBSTANCE
• The ground substance of cartilage is made up
of complex molecules containing proteins and
carbohydrates (proteoglycans).
• The carbohydrates are chemically
glycosaminoglycans (GAG).
• They include chondroitin sulphate, keratin
sulphate and hyaluronic acid.
• The core protein is aggrecan.

15. COLLAGEN FIBRES OF CARTILAGE
• The collagen fibres present in cartilage are
chemically distinct from those in most other
tissues.
• They are type II collagen.
• However, fibrocartilage and the
perichondrium, contain the normal type I
collagen.

16. HYALINE CARTILAGE
• Hyaline cartilage is so called because it is
transparent (hyalos = glass).
• The fibres are arranged so that they resist
tensional forces. The ground substance resists
compressive forces, while the collagen fibres
resist tensional force.
• In haematoxylin and eosin stained
preparations, the matrix is stained blue and it
is basophilic. However, the matrix just under
the perichondrium is acidophilic.

17. • Towards the centre of a mass of hyaline
cartilage the chondrocytes are large and are
usually present in groups (of two or more).
• The groups are formed by division of a single
parent cell.
• The cells tend to remain together as the dense
matrix prevents their separation.
• Groups of cartilage cells are called cell-nests
(or isogenous cell groups).

18. INDICATIONS AND CONTRAINDICATIONS
Indications
• Symptomatic focal chondral and
osteochondral defects
• defect of > 2 cm2 on the femur or patella
• Age of 13 to 55 years
• Patient's willingness and ability to comply with
postoperative rehabilitation
• Defects for which other treatments have failed

19. Contraindications
• More than 50% loss of cartilage thickness (joint
space narrowing)
• Inflammatory joint disease
• Unresolved septic arthritis or treatment of septic
arthritis within the previous 12 months.
• Metabolic or crystal disorder
• Deficient soft-tissue coverage
• Patient currently smoking
• Obesity (body mass index >35 kg/m2)
• Chronic narcotic use
• Age >55 years (relative contraindication)
• Allergy to collagen.

20. Identifying a Carticel Patient
Patient Factors:
• Younger patients – < age 55 (avg. ~ 35 yr)
• Significant impairment:
– Compromised daily living activities.
– Refractory to treatment
• Obesity
• Demanding Physical activities
• Willing & capable of rehabilitation program

21. Joint Factors:
• Symptomatic cartilage defects
– Moderate to large (> 2cm2 d.= 1.6)
– On the distal femur (mfc / lfc )
– Average defect size > 4 cm2
– Either chondral or osteochondral
• Relatively healthy joint –
– No arthritis
• Co-morbidities(meniscal tear, instability or
mal-alignment) must be corrected prior or
concurrent to implantation.

22. Pre-requisite for surgery :
• Appropriate biomechanical alignment
• Ligamentous stability
• Range of motion
Not recommended for patients who have :
• An unstable knee
• Patients sensitive to materials of bovine origins
• Allergic to the antibiotic gentamicin
• In children
• Yet in any joint other than the knee.

23. INVESTIGATIONS
Wt bearing x-ray
• Evidence of joint space narrowing 50% with
osteophyte formation, subchondral bony
sclerosis or cyst formation eliminates patients
from treatment (ie, if bone on bone changes
are present)

24. MRI
• MRI scanning, while helpful for soft-tissue
evaluation of meniscal or ligamentous injury
as well as assessment of bone bruises and
osteonecrosis.
• Does not have a high sensitivity and
specificity (75% to 93%) for determining the
extent of a chondral injury or subtle
chondromalacia changes.

25. • The gold standard for determining whether a
symptomatic patient is a candidate for ACI are
normal radiographs, accompanied by an
arthroscopic assessment showing focal
pathology.

26. ARTHROSCOPY
• Extent of lesion, Menisci, AP length of lesion.
• Quality and thickness of the surrounding
articular cartilage will determine whether
healthy cartilage will be available for
periosteal suturing or a non-contained
chondral injury will require suturing through
synovium or small drill holes through the
bone.

27. SURGICAL TECHNIQUES
• Uses your own cartilage cells (chondrocytes)
to repair the articular cartilage damage in your
knee.
• When implanted into a cartilage injury, your
own cells can form new cartilage; this new
cartilage injury is very similar to your original
cartilage.
• The CARTICEL implantation procedure is
called Autologous Chondrocyte Implantation
or ACI.
• It is a two-step process

28. Stage I: Biopsy
• The first stage involves
an arthroscopic
evaluation of the focal
chodral lesion to
assess containment,
depth, and potential
bone loss.

29. • A biopsy of normal hyaline cartilage is
obtained from either the supero-medial or
lateral femoral condyle or inner edge of lateral
femoral condyle at the intercondylar notch
using a curved bone-graft harvesting gouge.

30. • The total volume of
the biopsy should
be approximately
200 to 300 mg .

31. • It is better to penetrate to the subchondral
bone to ensure that the deep chondrocytes
are included in the biopsy.
• The biopsy is sent to product labs for
processing and cellular expansion.
• Biopsy can be stored upto two year.

32. Stage II: Implantation
• The second stage of the procedure is cell
implantation, which takes place between 1
and 24 months following the biopsy.
• During this stage arthrotomy done to expose
knee and any dead or damaged tissue from
the injury removed with curette, leaving only
healthy tissue .

33. Defect before (A) and after (B) preparation.

34. • The periosteal patch
is harvested
through a 3cm
incision on proximal
medial tibia.

35. • A patch that is at least 2mm
larger than the defect is
harvested to account for slight
shrinkage following
detachment.
• The patch is then sewn onto
the cartilage with the
cambium layer facing the
defect.
• Carticel injects under the
patch.

36. • After cell implantation, the sutured periosteal
patch is sealed with fibrin glue

38. In Vitro Human chondrocyte growth
Cartilage Biopsies:
• The cartilage fragments had 1cm² area in
diameter and were transported immediately
after harvesting in sterile tubes containing
phosphate buffered saline
(penicillin/streptomycin and Amphotericin B).

39. Chondrocyte Isolation from Cartilage:
• A sterile scalpel was used to cut the cartilage into
small pieces and transferred to Petri dish.
• Sliced cartilages were maintained overnight at
37°C, 5% CO₂ as in ham's F12: DMEM containing
fetal bovine serum, ascorbic 100 mg/ml and
penicillin/streptomycin 100 u/100 mg/ml to
ensure sterility of the specimens prior to
enzymes digestion, the time of enzymatic
digestion was adapted to the size of tissue
fragments.
• The prepared cartilage was washed in phosphate
buffered saline (PBS) containing antibiotics.

40. • The tissue was then incubated in the above
growth medium containing trypsin at 37°C, 5%
CO₂ for 30 minutes.
• Supernatants were discarded and the cartilage
was further digested in growth medium
containing 0.8 mg/ml collagenase II for 4 hours at
37°C, 5%CO₂.
• The digested tissue was then allowed to settle
and the supernatant containing the cells was
removed and centrifuged at 1500 rpm for 5
minutes at 4°C.
• The final digest was centrifuged at 1500 rpm for
10 min.

41. • The cell pellet was washed three times in PBS
or serum free medium.
• Cell suspensions were passed through a nitex
nylon filter (mesh width 70 mm) and
re-suspended in serum containing medium.
• The cell viability was determined by trypan
blue dye exclusion test in hemocytometric
chamber.

42. Chondrocytes Monolayer Culture:
• The cells isolated cartilage were seeded in 4 well
culture plates at low density (4000 cell/cm²) in
culture medium in an equal volume mix of ham’s
F12 medium and Dulbecco's modified eagle
medium (F12:DMEM).
• Each culture mediums also contained Ascorbic acid
(50µl/ml), penicillin (100µl/ml), streptomycin
(100µl/ml), L-glutamine (200mM) and incubated in
a CO₂ incubator at 5% CO₂, 37° C and saturated
humidity for periods of up to 40 days.
• The culture medium was changed twice or three
times per week.

43. Morphology and Passage Chondrocytes:
• The cell morphologic feature was examined
daily by phase microscopy in an inverted
microscope.
• The cultures were regularly photographed for
monitoring cell morphology.
• When chondrocytes reached 70 to 90%
confluency, the cells were detached
mechanically, using trypsinization 1 to 2 min
and harvested chondrocytes were washed and
re-suspended in PBS than total cell number
and their viability was assessed using the
trypan blue test.

44. Human chondrocytes primary culture (passage zero) after 40 days showed
different morphology feature in DMEM/F12 supplemented with different
factors
A: primary culture after 13 days B: primary culture after 25 days
C: culture of 40 days D: chondrocytes culture

45. SANDWICH TECHNIQUE
• ACI has traditionally been applied to treat
relatively shallow lesions of articular cartilage
without involvement of the subchondral bone.
• For osteochondral defects of more than 8 to
10 mm in depth, bone grafting is
recommended.
• The bone graft may be performed at the time
of biopsy and the implantation may be
delayed to allow for bone graft consolidation.

46. • Alternatively, the “sandwich technique” has
been utilized to replace bone and resurface
the defect in a single step.
• With this technique, the bone defect is filled
with bone graft, periosteum is sutured on top
of the bone graft at the level of the
subchondral bone plate, a second layer of
periosteum is placed over the cartilage defect,
and the chondrocytes are then placed
between the layers of periosteum.

48. Stages of ACI healing
• Healing process has several stages. They
include the
• proliferative stage (0 to 6 weeks),
• the transition stage (7 to 12 weeks), and
• a remodelling and maturation stage which
occurs over a prolonged period (13 weeks to 3
years)

49. Proliferative stage
• During this stage, a primitive cell response
occurs with tissue fill of the defect and poor
integration to underlying bone or adjacent
cartilage.
• Mostly type I and some type II collagen is
produced.
• The tissue is soft and jelly-like and easily
damaged

50. Transition phase
• This marks the production of type II collagen
framework and the early production of
proteoglycans.
• The proteoglycans, which form the matrix, help
imbibe water to give cartilage its viscoelastic
properties.
• The tissue is not yet firm or well integrated and
has the consistency of a firm gelatin.
• It is milkable when probed with an arthroscopic
nerve hook, indicating incomplete integration to
underlying bone.

51. Stage of remodelling and maturation
• The matrix proteins cross-link and stabilize in large
aggregates.
• The collagen framework reorganizes so as to
integrate into the subchondral bone and form
arcades of Benninghoff.
• Usually by 4 to 6 months, the tissue has firmed up
to a putty-like consistency and is integrated to the
underlying bone.
• At this stage, patients experience good symptom
relief.
• During this stage excessive activity may cause
repair tissue degeneration or continued
improvement in remodelling.

52. ONE YEAR ASSESSMENT
• Radiographs
– Alignment
– Bone quality
• MRI
– Healing cartilage
– Graft failure
• Arthroscopy
– Graft integrity
– Pressure
– Biopsy

53. FOLLOW UP: Biopsy

54. FOLLOW UP: MRI

55. FOLLOW UP: ARTHROSCOPIC
• Final appearance of the
periosteum sutured
over femoral condyle
defect. The cartilage
cells have been injected
under the flap and the
final suture placed to
close the "cover" and
provide a watertight
seal.

56. • Arthroscopic
appearance of the same
area 12 months after
Carticel™ implantation.
The defect is no longer
visible and there is now
hyaline cartilage filling
the original defect site.

57. REHABILITATION GUIDELINES
• Immobilization: First 12-24 hours
• (CPM): After 12-24 hours, for about 4 weeks
• Complete joint loading: from about 5th week
patella from about 8th week condyle
• Back to sports:
– Low impact → within 6 months
– Repeated impact → from 8th month
– High impact → from 10-12th month

58. ACI REHABILITATION
Weight bearing
• It is recommended to keep you in non-weight
bearing until the second week after surgery
(ACI). You can increase the weight bearing
gradually and you may be able to sustain your
full weight bearing after 6 to 12 weeks from
surgery.

59. Range of motion
• Recovery on your ROM (Range of Motion) is
gradually increased with a continuous passive
motion (CPM) machine and may be completed
to 140 degrees of ROM at 6 weeks to 12
weeks after surgery

60. Indoor exercise
• You can strenghthen your muscles
surrounding the knee joint with a four point
exercise, as well as isometric, hamstring and
squatting exercises, from 4 weeks to 6 weeks
after surgery.
• You may start performing stationary bike
activity without resistance and increase the
resistance gradually.

61. Outdoor exercise
• At 13 weeks after surgery you can start
walking lightly and at 10 months after surgery
you can perform jogging and then you may
enjoy intensity exercise like playing tennis or
volleyball from 18 months after surgery.

62. Rehabilitation Goals at 12 weeks after
ACI
• Full ROM (Range of Motion)
• Minimal edema level
• Minimal occasional pain
• Pain free tolerance to transitional phase
exercise with adequate stability and motor
control

63. ADVANTAGES
• Can produce hyaline-like cartilage.
• Can fill defects regardless of size with functional
repair tissue.
• Moderate to large defects that have failed
previous intervention.
• Repair tissue which matures, rather than damage
over time.
• Expected outcome
• Return to previous level of functioning

64. DISADVANTAGES
• More invasive
• Expense
• Longer recovery

65. COMPLICATIONS
• The overall failure rate is 10%.
The two most common complications include :
• loosening of the transplant tissue,
• formation of fibrous tissue at the repair site
and adhesions with return of pain.
Other adverse events include :
• post-op haematoma (big blood clot)
• hypertrophic synovitis (angry knee) and
superficial wound infection.

66. COMPARISON WITH OTHER
METHODS OF TREATMENTS :
Autologous Chondrocyte Implantation
• Aims to increase the best condition for
cartilage defect.
Advantages:
• Hyaline cartilage is formed
• Permanent reconstructed cartilage tissue
• Most valuable treatment

67. Abrasion arthroplasty
• Aims to decrease the inflammation of the
joint.
Disadvantages:
• Removes many cartilage fragments from the
joint
• Symptoms reoccur within one year

68. Drilling and Microfracture
• Aims to generate a healing response.
Disadvantages:
• The healing response in inadequate
• No hyaline cartilage is formed, but rather
fibrocartilage
• Has a limited lifespan of approximately one
year
• Rapid damage after such procedures can be
expected

69. Perichondral Resurfacing
Disadvantages:
• Isolated cartilage defects are often too large
to be covered by perichondrum
• Long term follow-up of such procedures
indicate that the implants undergo
endochondral ossification

70. Synthetic Materials (i.e. Carbon Fiber Mesh)
Disadvantages:
• It often results in fibrous tissue formation
• Not adequate biomechanically
• Often the cause for synovitis in the joints

71. Osteocartiloginous Grafts
• Aims to reconstruct joints.
Disadvantages:
• Unless fresh cartilage is transplanted, the
cartilage is dead
• Fresh grafts are not commonly used, as they
inevitably carry a risk of disease transmission
• Cyropreserved grafts can survive for many
years, but ultimately damage