Transcript Tissue bioengineering: a new challenge for wound healing

Physiology of wound healing
Wound healing is:
– Complicated process that involves at least
4 distinct cell types
– Commonly referred to as occurring in
PHASES
– Affected by several factors
Phases of wound healing
process (WHP)
Maturation
Proliferation
Inflammation
Haemostasis
Where does a chronic
wound get stuck?
Platelet Activity
WOUND
Tissue factor
Exposed collagen
Messengers for
Aggregation &
coagulation
Growth factors
(PDGF)
Extrinsic pathway
VII
Intrinsic pathway
XII
Intrinsic pathway
intermediates
IX, VII
Platelet
Coagulation pathway
intermediates
V, X
Other enzymes
(proteases)
Fibrinogen
FIBRIN
Prothrombin & thrombin
Cross-linked fibrin clot
(structural support for wound healing)
Xlila
Role of keratinocytes in wound
healing
Keratinocyte
Migration/
Profileration
• Epibloy
• Integrins
ECM
production
• Matrix formation
• Basement
membrane
formation
Protease
release
Angiogenesis
Growth factor/
Cytokine production
• VEGF
• TGF-α
• PDGF
• PD-ECGF
• Chemoattractants
− VEGF
− KGF (FGF-7)
• Dissolves
− Nonviable
tissue
− Fibrin
barrier
Selected growth factors
important to wound healing
• EGF (epidermal growth factor). Stimulates wound reepithelialization and stimulates blood vessels and fibroblasts.
• FGF (fibroblast growth factor). Stimulates new blood vessel
and collagen formation.
• PDGF (platelet derived growth factor). Attracts/stimulates
smooth muscle cells, fibroblasts, and other cells. Important in
ECM formation.
• TGF-β (transforming growth factor-beta). Slows buildup of
epithelial cells, suppresses immunoglobulin secretion and is
helpful in ECM formation.
• TNF-a (tumor necrosis factor-alpha). Activates neutrophils,
causes fibroblasts to multiply, causes bone/cartilage resorption.
• IL-1 (interleukin-1). Attractant for epithelial cells, neutrophils,
mono and lymphocytes; also stimulates collagen synthesis.
Chronic wounds –
characteristics
•
•
•
•
•
•
Increased inflammatory cytokines
Altered fibroblast phenotype
Abnormalities growth factors
Increased proteases
Altered keratinocyte function
Senescent cells (increased number)
Wound bed preparation
• Debridement
• Bacterial balance
• Dressing therapies
(f.i. silver dressings prevent of infections,
help reduce healing time)
Local wound care
debridement
Surgical
Autolytic
Enzymatic
Biological
moisture balance
Foams
Calcium alginates
Hydrogels
Hydrocolloids
Adhesive films
Negative pressure therapy
Tissue engineering
Biology, medicine and technology are
today closely interleaved with each
other
Tissue engineering – combining cells and
biomaterials into functional tissues
Cells are seeded onto a biomaterial
scaffold to be integrated into a specific
tissue
Tissue engineering
Advances
– Biological wound dressings
– Material scaffolds and cell material
interactions
– The use of stem cells for tissue engineering
– Combination of stem cells and material
scaffolds into tissue engineered
replacements of tissues and organs
Tissue engineering implants
Synthetic polymeric biomaterials
• Nonbiodegradable
Is required to provide and maintain optimal
cellular function -> e.g. alginate, liposome,…
• Biodegradable
To restore the histological structure and
replace the cellular function of recipients -> e.g.
poly L-lactic acid, poly glycolic acid, …
Wound healing promoting
anti-adhesive matrix
The collagen grafting is also applied to
produce a healing / promoting
antiadhesive membrane
• Particularly necessary in peritoneal surgery to
prevent postoperative adhesion
• To produce skin wound dressing membranes
Methods of tissue
bioengineering
Skin replacement
– Cultured epidermal graft
– Cultured human autologous and allogeneic
keratinocytes
– Semi synthetic materials (composed of
human neonatal dermal fibroblasts cultured
onto a bioabsorbable mesh)
Methods of tissue
bioengineering
• Active dressings (f.i.: with maggot’s
excret, with honey)
• Photobiomodulation (modulate cellular
activity in red to near infrared light)
• Hyperbaric oxygen therapy: as
therapeutic benefit in WT
• Growth factors (from blood)
Allogeneic cultivated human skin
keratinocytes
• Make rapid healing of the ulcers particularly those
that are difficult to heal
• No clinical or laboratory evidence of rejection
• No evidence of preexisting cytotoxic antibodies
specific fort the HLA class I antigens expressed on
HSE cells
• A fibrin-based skin substitute produced in the
defined keratinocyte medium could be safely used to
threat a number of skin defect
Preparation of autologous
fibrin-based skin substitutes
Methods of tissue
bioengineering
• Autologous platelet rich plasma product
(platelet gel)
• Allogeneic platelet gel
The effect is attributed to the growth
factors
Fracture
Impaired
healing
Delayed union
Pseudoarthrosis - nonunion
(bone defect)
Impaired
healing
Infection
?
Method of
treatment?
• Large bone defect
Lack of osteogenic progenitor cells
• Diabetes, glucocorticoid treatment,
chemotherapy, ...
Accepted methods of treatment
• Autologous bone transplants
– Cancellous bone graft (contains all necessary
characteristics of bone substitutes)
– Corticocancellous graft (possibly vascularized
limited amount)
• Homologous (allogeneic) graft
Bone banks, treated (no rejection), contains only
osteoconductive properties
• Ilizarow intercallary bone transport (traction
method)
Properties of bone grafts
• Osteogenesis (bone marrow, cancellous
bone)
• Osteoinduction
– Demineralized bone matrix
– Growth factors (platelet rich plasma, bone
morphogenic proteins – BMPs)
• Osteoconduction
– Ceramics
– Collagen
Alternatives
Bone substitute (biomaterials for scaffold):
– Demineralized bone matrix
– Biocompatible ceramics
– Synthetic Calcium phosphate
– Mineral bone
– Collagen
– Composite grafts
– Osteoinductive collagen
Alternatives
Role of GROWTH FACTORS
Role of STEM CELLS
Collagen based matrices in
tissue engineering
• Skin equivalent
• Cartilage repair
• Bone repair
Matrices are also prepared from
synthetic polymers
Fracture healing promoting
molecules
Growth and different factors
– The transforming growth factor-β (TGF-β)
superfamily
• Bone morphogenetic proteins
(osteoprogenitors, mesenchymal cells, osteoblasts and
chondrocytes within the extracellular matrix produce BMPs.)
BMP-2, BMP-4
BMP-5, BMP-6, BMP-7
GDF-5 (BMP-14), GDF-6 (BMP-13), GDF-7 (BMP-12)
BMP-3 (Osteogenin), GDF-10 (BMP-3b)
– Platelet-derived growth factor (PDGF)
– Fibroblast growth factor (FGFs)
– Insulin-like growth factor (IGFs)
Platelet rich plasma (contains high concentrations of growth
factors) especially TGF-B and PDGF
Autologous
Allogeneic
PLATELETS
MONOCITE
NEUTROPHILS
PDGF
FIBROBLAST
TGF-β
SMOOTH MUSCLE
MACROPHAGE
PDGF
TGF-β
ENDOTHELIUM
OSTEOBLASTS
• Mesenchymal stem cell: the promise for
treating skeletal disorders
• Adult stem cell are being isolated from
various tissues
Adult stem cells
Bone marrow contains
– Hematopoietic stem cells (HSCs)
• All types of blood cells
– Bone marrow mesenchymal stem cells (MSCs)
• Generating bone, cartilage, fat, fibrous connective tissue
Bone tissue formation
Osteogenic progenitor cells
Locations:
• Periost
• Peritrabecular soft tissue
• Cancellous bone and bone marrow (in
BM aspirate up to 40x less stem cells
then in cancellous bone)
Our method of tissue
engineering
Combined graft
Autologous cancellous bone with
stem cells
Allogeneic platelet gel
(source of GFs)
Mixed
manually grounded autologous cancellous bone
with stem cells
corresponding amount of allogeneic platelet
concentrate (app. 1,4x109 platelets per 1 ml)
AND
Added 0,06 ml human thrombin in 40 mM CaCl2
for the activation of platelets
in 1 minute
the resulting gelled graft can be shaped
according to the bone defect and implanted
Our graft
Autologous cancellous bone with stem
cells and allogeneic platelet gel
Conclusions
The use of autologous cancellous bone
with stem cells and allogeneic platelet
gel is safety and effective method
for the treatment of nonunion of long
bones
Future
Gene arrays for gene
discovery
FUTURE
Cell and tissue engineering
Detection of numerous signal pathways
activated during physiological processes
Self (re)restoration and differentiation
off mammalian embryonic, fetal and
stem cells of adult tissues
Thank you for attention !