CELL-BASED MYOGENIC AND
ANGIOGENIC THERAPY FOR
MYOCARDIAL REGENERATION
J.C. CHACHQUES, MD, PhD
POMPIDOU HOSPITAL
PARIS - FRANCE
René
Magritte
Surrealist
painter
1898-1967
EUROPEAN HOSPITAL GEORGES POMPIDOU
CARDIAC BIOASSIST PROCEDURES
MOLECULAR BASIS OF CONGESTIVE
HEART FAILURE
• LACK OF MYOCARDIAL STEM CELLS
• UNABILITY OF THE DAMAGED CELLS TO
UNDERGO SUFFICIENT REPAIR
HEART FAILURE THERAPY
•
PHARMACOLOGICAL THERAPY
• CARDIAC PACING FOR RESYNCHRONIZATION
• VENTRICULAR RESTORATION, LV REDUCTION
• CARDIOMYOPLASTY, AORTOMYOPLASTY
• VENTRICULAR CONTAINMENT, MITRAL VALVULOPLASTY
• HEART TRANSPLANTATION
• MECHANICAL ASSIST DEVICES - ARTIFICIAL HEART
• CELLULAR CARDIOMYOPLASTY
CELL TRANSPLANTATION THERAPY
• VASCULAR SURGERY: CRITICAL LEG ISCHEMIA
• HEMATOLOGY : LEUKEMIA
• DERMATOLOGY : EPIDERMAL CELLS
• OPHTHALMOLOGY: CORNEAL REGENERATION
• ORTHOPEDICS : CHONDROCYTES
• MYOLOGY : DUCHENNE DYSTROPHY
• NEUROLOGY : PARKINSON DISEASE
• HEPATOLOGY : HEPATOCYTES
• DIABETES : LANGERHANS ISLETS
NOVEL EMERGING TREATMENTS
IN ISCHEMIC HEART FAILURE
• CELL-BASED MYOGENIC THERAPY
• CELL-BASED ANGIOGENIC THERAPY
LIMITATIONS OF ANGIOGENIC GROWTH
FACTORS AND GENE THERAPY
• GROWTH FACTOR PROTEINS
Systemic effects: angiogenesis in the retina.
Intimal arterial hyperplasia and development of
atheromatous plaque.
Potentiation of growth and metastasis of occult tumors
• GENE THERAPY
Instability and adverse response to transfection vectors
ANGIOGENIC CELLS
• BONE MARROW DERIVED ANGIOBLASTS
Hematopoietic stem cells (HSC): CD34+
Progenitor endothelial cells (PEC): CD 133+
• VASCULAR ENDOTHELIAL CELLS
collected from the intima of arteries or veins
CliniMACS
instrument
for immunomagnetic cell
selection
(microbeads)
Miltenyi Biotec
Germany
MYOGENIC CELLS
• SKELETAL MYOBLASTS ( satellite cells )
• SMOOTH MUSCLE CELLS
• BONE MARROW CELLS :
Multipotent adult progenitor cells (MAPC)
• CARDIOMYOCYTES : fetal, neonatal
SKELETAL MUSCLE BIOPSY
MYOBLAST CULTURE TECHNIQUE
Manipulations in a laminar flow hood
• REMOVAL OF FIBROUS AND ADIPOSE TISSUE FROM
MUSCLE BIOPSY
• MUSCLE MINCING WITH SCISSORS
• ENZYMATIC (collagenase + trypsin) & MECHANICAL
DIGESTION
• MULTIPLE CENTRIFUGATIONS
MYOBLAST CULTURE TECHNIQUE
•
ISOLATION OF MYOBLASTS, EXCLUSION OF FIBROBLASTS
•
IN VITRO MYOBLASTS CULTURE : 3 WEEKS, 37°C, 5% CO2
• ASSESSMENT OF % CELL VIABILITY : TRYPAN BLUE
• ASSESMENT OF % MYOBLASTS / FIBROBLASTS :
CD 56 and DESMIN ANTIBODIES
•
STERILITY TESTS : BACTERIAL, VIRAL, FUNGIAL
•
CELL SUSPENSION IN 0.5 % HUMAN ALBUMIN FOR
MYOCARDIAL INJECTION
CELL CULTURES with AUTOLOGOUS-HUMAN-SERUM
- obtained from plasmapheresis or blood sample • AVOIDS THE FIXATION OF ANIMAL PROTEINS (FBS) ON THE
CELL SURFACE
• AVOIDS IMMUNOLOGICAL AND INFLAMMATORY ADVERSE
EVENTS LEADING TO FIBROSIS AND MICRO-REENTRY
CIRCUITS
• REDUCE THE RISK OF ARRHYTHMIA AND THE NEED OF A
DEFIBRILLATOR
• AVOIDS THE RISK OF PRION, VIRAL, OR ZOONOSES
CONTAMINATION
TRANSPLANTED SKELETAL MYOBLASTS
INTRAMYOCARDIAL ORGANIZATION
• ISOLATED MYOBLASTS
• MULTINUCLEATED MYOTUBES
• MYOFIBRES
IN VIVO MYOBLAST ORGANIZATION
CELL IMPLANTATION TECHNOLOGIES
EPICARDIAL APPROACH
• SURGICAL : CLASSIC OR MINIMALLY INVASIVE
• THORASCOCOPIC
ENDOVENTRICULAR : CATHETER BASED
• ASSISTED BY 3D ELECTROMECHANICAL MAPPING
• ASSISTED BY ECHO & FLUOROSCOPY, MRI
INTRAVASCULAR
• CATHETER BASED INTRACORONARY
• CORONARY VENOUS ROUTE or INTRAVENOUS SYSTEMIC
ELECTROMAGNETIC 3D CARDIAC MAPPING
PRE / POSTOPERATIVE EVALUATION
MYOCARDIAL VIABILITY
• POSITRON EMISSION TOMOGRAPHY : 2 fluoro
deoxyglucose.
• MAGNETIC RESONANCE IMAGING : uptake of
gadolinium.
• SCINTIGRAPHY: stress-redistribution-reinjection
201 thallium scintigraphy.
PRE / POSTOPERATIVE EVALUATION
VENTRICULAR FUNCTION
• BASAL AND DOBUTAMINE STRESS ECHOCARDIOGRAPHY
• RADIONUCLIDE VENTRICULOGRAPHY
• CARDIAC CATHETERIZATION + CORONAROGRAPHY
• COLOR KINESIS AND DOPPLER TISSUE ECHOGRAPHY
NEUROHORMONAL ACTIVATION
(BNP)
CELLULAR CMP - INCLUSION CRITERIA
• MYOCARDIAL INFARCT
• DILATED CARDIOMYOPATHIES
• NYHA FUNCTIONAL CLASS 2 - 3
• LV WALL THICKNESS > 4 mm
• LV EJECTION FRACTION : 20 to 40%
• VIRUS-FREE TESTS
• FREE OF UNCONTROLABLE ARRHYTHMIAS
CELLULAR CMP
MECHANISMS OF BENEFICIAL EFFECTS
VENTRICULAR REMODELING
• REDUCES THE SIZE AND FIBROSIS OF INFARCT SCARS
• MINIMIZES GLOBAL VENTRICULAR DILATATION
• INCREASES MYOCARDIAL WALL THICKNESS
• INDUCES MODULATION OF EXTRACELLULAR MATRIX
CELLULAR CMP
PREOP
-
PET 18 FDG
3 MONTHS
CELLULAR CMP
MECHANISMS OF BENEFICIAL EFFECTS
DIASTOLIC FUNCTION
• IMPROVES MYOCARDIAL WALL TENSION AND ELASTICITY
• IMPROVES STRAIN AND DYNAMIC STIFFNESS
• REVERSES DIASTOLIC CREEP
CELLULAR CMP
MECHANISMS OF BENEFICIAL EFFECTS
SYSTOLIC FUNCTION
• IMPROVES REGIONAL VENTRICULAR WALL MOTION
• INCREASES DEVELOPED PRESSURES
• IMPROVES GLOBAL VENTRICULAR CONTRACTION ?
CLINICAL TRIALS
• > 150 ischemic patients (Europe, America, Asia)
• Skeletal Myoblasts <=> Bone Marrow Cell Implants
• Surgical <=> Interventional Cardiology Procedures
• Defibrillators only implanted in myoblast approach
(when cultivated with bovine serum)
CLINICAL DIFFICULTIES TO BE SOLVED
• CHOICE OF CELL TYPE AND DOSE
• CELL ENGRAFTMENT AFTER IMPLANTATION
need of prevascularization ?
• DIFFERENTIATION OF STEM CELLS IN FIBROBLASTS
• HOST-CELL INTERACTIONS
mechanical and electrical coupling ?
• ELECTRICAL INSTABILITY >>ARRHYTHMIAS
CONCLUSIONS
CURRENT EXPERIMENTAL AND CLINICAL RESULTS
SUGGEST THAT CELLULAR CRDIOMYOPLASTY FOR
MYOCARDIAL REGENERATION MAY BE EFFICIENT TO
AVOID PROGRESSION OF VENTRICULAR REMODELING
AND SUBSEQUENT HEART FAILURE IN PATIENTS WITH
MODERATE CARDIAC INSUFFICIENCY RESULTING
FROM ISCHEMIC HEART DISEASE
NEW DEVELOPMENTS
• ASSOCIATION OF ANGIOGENIC AND MYOGENIC CELLS
• PRE-CONDITIONING OF STEM CELLS
• COMBINATION WITH CARDIAC PACING :
DYNAMIC SUPPORT
• NEW INDICATIONS FOR CELLULAR CMP :
ISCHEMIC MITRAL REGURGITATION
NON ISCHEMIC CARDIOMYOPATHIES
PRE-CONDITIONING OF STEM CELLS
• Treatment with 5-azacytidine : toxic
• Co-culture with cardiomyocytes : limited expansion
• In-vitro electrostimulation
ELECTROSTIMULATION
OF STEM CELL CULTURES
Scientific Basis of Myocardial Differentiation
- ANALOGY Embryon and fetus
Cell cultures
Cardiac conduction tissue
Pacemaker (rate 120 min)
120 impulses min
Mesoderm
Stem cell cultures
Myocardium
Cardiomyocytes ?
Human CD34+ cell cultures at 3 weeks
Without Stimulation
Electrostimulated
Human CD34+ cell cultures at 3 weeks
Desmin antibody
Without Stimulation
Electrostimulated
CONCLUSION
• Positive effect of electrostimulation
over human stem cell cultures have been
detected
• Myogenic differentiation was observed
in electrostimulated CD34+ cells