at the University of Pécs and at the University of Debrecen
Identification number: TÁMOP-4.1.2-08/1/A-2009-0011
CONTROLLED RELEASE
Dr. Judit Pongrácz
Three dimensional tissue cultures and tissue engineering – Lecture 13
at the University of Pécs and at the University of Debrecen
Identification number: TÁMOP-4.1.2-08/1/A-2009-0011
scaffolds
• Drug release upon matrix degradation
• Drug release upon diffusion
• Long-term maintenance of effective local concentration
• Localized effects ensured
• Limited systemic effects
Ideal scaffold
• 3-dimensional and well defined microstructure
• Interconnected pore network
• Mechanical properties similar to those of natural tissues
• Biocompatible and bio-resorbable
• Controllable degradation and resorption
• Local sequestration and controlled delivery of specific bioactive factors
• Thus enhancing and guideing the regeneration process
design
• ECM is the natural medium where cells proliferate, differentiate and migrate
• ECM is a highly organized dynamic biomolecular environment where motifs governing cell
behaviours are continuously generated and sequestered
• Motifs are locally released according to cellular stimuli
• Relase occurs on-demand upon degradation of the adhesion sites binding them to the ECM
Growth factors and the ECM
• Growth factors (GFs) are locally stored by ECM
• Storage in insoluble/latent forms
• Specific binding with glycosaminoglycans (e.g.
heparins)
• Elicit biological activity once released
• ECM binding provides concentration gradient important in morphogenesis
Mimic the function of ECM
• Future generations of TE scaffolds need to have extended functionality and bioactivity
• Synthetic bio-inspired ECM should broadcast specific cellular events
• The ability of controlled release of multiple bioactive molecules will allow the control of cellular behaviour and successful regeneration
Interspersed signals
• Hydrogels (either natural or synthetic) have been succesfully used for controlled release of bioactive protein compounds
• Molecules were simply mixed with the polymer and were entrapped upon gelation
• Natural (collagen, fibrin, hyaluronan) and synthetic
(PEG-based, peptide-based) hydrogels have been used
• Release characteristic may modulated with crosslinking agents
• Solid-state scaffolds: fabrication method must be mild (to avoid protein denaturation)
Immobilized signals
• Modification of polymer scaffolds to interact with signaling molecules: immobilization
• Prolonged diffusion out of the scaffold platform
• Reversible or irreversible binding to the polymer.
• Released upon degradation of a linking tether or the matrix itself
• Determinants of the amount of bound signal and release profile:
– The number of binding sites – Affinity of the signal for sites
– Degradation rate of the scaffold
Signal delivery from cells
• Inclusion of nucleic acids (NA) encoding the desired protein
• NA are introduced into target cells, which then produce the desired proteins
• Antisense oligos can be used to return abnormal gene expression to a certain state
• Synthetic polymers containing adhesion sites
(RGD) proved to be more effective in delivering the plasmid
Protein delivery systems (DS) in TE
• DS must prevent the protein from inactivation or degradation
• Fine-tuning of the release rate can be achieved by modulating the composition, shape, and
architecture of the platform
• Continous and pulsatile delivery
• Biodegradable and non-degradable platforms
Non-biodegradable systems
Ethylene-vinyl acetate copolymers (EVAc) and silicones:
• Mass transport through polymer chains or pores is the only rate-limiting step
• Possible application in cell encapsulation preventing them to interact with the immune system
Time
• PLGA is a very versatile and widely used system
• Poly-ortho esters are newly in the centre of interest (no heating or solvents, injectable polymers)
• Polyanhydrides usually undergo surface erosion which has a favorable kinetics
Biodegradable systems
Time
systems
Surface erosion
Bulk erosion
Corresponding rate
Typical release profile
t t
dc(t)/dt
Release rate
Amount of drug released
t t t
dc(t)/dt
Toxic dose
ceff(t) Protein or small
molecule drug
Protein or small molecule drug
Amount of drug released
Release rate
Corresponding rate
Typical release profile
On-off drug delivery systems
• Pulsatile mode of protein and peptide release
• Rapid and transient release of a certain amount of drug
molecules within a short time-period immediately after a pre- determined off-release interval
• Classified into “programmed” and “triggered” delivery systems (DS):
– Programmed-DS: the release is governed by the inner mechanism of the device
– Triggered-DS: release is governed by changes in the physiologic environment of the device or by external stimuli
• External stimuli involve temperature changes, electric or magnetic fields, ultrasounds or irradiation
systems
• Synthetic polymers can be engineered to be applicable in programmed delivery
• Both surface and bulk-eroding systems may be used
• Biggest interest in triggered delivery is the glucose-sensitive insulin delivery
• The “intelligent” system consists of immobilized glucose oxidase in a pH-responsive polymeric hydrogel
• In the gel, insulin is enclosed
• Upon glucose diffusion into the hydrogel, glucose oxidase converts it into gluconic acid
• Lowering of the pH results in gel swelling and insulin release
scaffolds
Poly-methyl-methacrylate (PMMA) beads with antibiotics (mostly aminoglycosides):
• Orthopedic and trauma surgery
• Treatment of chronic osteomyelitis and/or ulcers
• Bones and joints are „blind spots” of systemic antibiotic therapy because the limited blood supply
• PMMA beads release antibiotics gradually
• High local antibiotic concentration can be achieved
• Limited systemic side effects
scaffolds
VEGF role in tissue vascularization:
• Cells in hypoxic tissues secrete VEGF
• Endothelial cells express VEGFR
• Stimulates endothel proliferation
• Directs endothelial cell migration
• Tissue vascularization is critical in nutrition and oxigenization of implanted TE constructs
• Controlled VEGF delivery is in the focus of TE research
vascularization
Controlled VEGF delivery from alginate microparticles:
• Bivalent cations mediate alginate crosslinking
• VEGF encapsulation efficiency and delivery ratio depends on the cation species (Ca2+ or Zn2+)
• Zn2+-crosslinked particles proved to be more toxic than Zn2+
• Mixture of Ca2+ and Zn2+ beads are the most favorable
bioactive proteins
BMP-2:
• Key role in regulating osteoblast differentiation
• Recombinant hBMP-2 is dissolved in aquaeous solution of polyethylene-oxide (PEO)
• rhBMP-2 solution is then added to scaffold material
• Scaffold materials include silk fibroin, PCLA, PEG, PLGA, collagen, etc.
delivery scaffolds – VEGF
• Half-life of VEGF is 50 min, therefore controlled release is critical
• Controlled release is based on electrostatic
attractions between the carrier (acidic gelatine, IEP=5.0) and VEGF (IEP=8.6)
• Extent of gelatin cross-linking also influences release
• Up to 90% of total VEGF vas released within 30
days from sc. implants, 80% within the first 5 days.
delivery scaffolds – BMP-2
• Use of BMP-2 filled collagen sponges in spinal
degenerative diseases to enhance post-operative bone fusion.
• BMP-2 treated patients regain the ability to self- care and mobility faster, their pain scores are significantly lower.
• Their mood and emotional control is also
significantly better than that of control patients.
BIOSENSORS
Dr. Judit Pongrácz
Three dimensional tissue cultures and tissue engineering – Lecture 14
at the University of Pécs and at the University of Debrecen
Identification number: TÁMOP-4.1.2-08/1/A-2009-0011
Definition
Biosensor is a device that transforms or detects a biological signal and transforms into a more easily detectable one.
sensor
Detector (potentially a mobile phone)
Glucose sensor Implantable potentiostat
Type I
Signal
Type II
Insulin release Glucose sensor
Insulin container
Signal Signal
Dexamethasone-loaded PLGA Microspheres
10m
interactions
Interphase
Microsphere for drug (TRM)
release
Tissue
Angiogenesis Hydrogels + PEO
Endothel cell
Sensor
Biosensor
WBC
Angiogenic factor or other tissue response modifiers Soluble proteins Fibrin Collagen
RBC
The “intelligent” system
• Consists of immobilized glucose oxidase in a pH- responsive polymeric hydrogel, enclosing a
saturated insulin solution.
• As glucose diffuses into the hydrogel, glucose
oxidase catalyzes its conversion to gluconic acid, thereby lowering the pH in the microenvironment of the membrane.
• Low pH causes swelling and insulin release.
biosensors require
1. Novel electrodes are required to decrease invasiveness of the implantable glucose biosensor
2. Bioactive coatings are necessary to enhance the in vivo life of the implantable glucose sensor
3. Biosensor coating using electrospinning nanofibres need to be developed
4. Tissue responses are needed to be studied further to optimize tissue responses to biosensor signals
5. Angiogenesis around the glucose sensor need to be
increased to enhance detection potential of glucose levels and
6. Finally, novel biostable 3D porous collagen scaffolds need to be developed for tissue compatible biosensors
