PRGF-Modified Collagen Membranes for Guided Bone Regeneration: Spectroscopic, Microscopic and Nano-Mechanical Investigations

By C. Ratiu, Simona Cavalu et al.

The aim of our study was to evaluate the properties of different commercially available resorbable collagen membranes for guided bone regeneration, upon addition of plasma rich in growth factors (PRGF). The structural and morphological details, mechanical properties, and enzymatic degradation were investigated in a new approach, providing clinicians with new data in order to help them in a successful comparison and better selection of membranes with respect to their placement and working condition. Copyright Simona Cavalu et al.

Whole blood separation upon centrifugation at 580 G for 8 minutes at room temperature (a) and subsequent platelets rich in growth factor (PRGF) separation by pipetting (b); membrane immersion in PRGF (c). Copyright C. Ratiu, Simona Cavalu et al.

Hematology parameters of whole blood and PRGF fraction:

ComponentWhole bloodPRGF
Leukocytes (x 103/μL)5.9 ± 1.20.3 ± 0.2
Erythrocytes (x 106/μL)4.5 ± 0.40.01 ± 0.0
Platelets (x 103/μL)210 ± 20655 ± 85
Hematology parameters of whole blood and PRGF fraction. Copyright Simona Cavalu et al.
Growth factor contentValue
Transforming growth factor TGFβ1: enhances the proliferative activity of fibroblasts and stimulates the biosynthesis of collagen and fibronectin43 ng/mL
Vascular endothelial growth factor VEGF: induces angiogenesis via migrating endothelial cells220 pg/mL
Insulin –like growth factor IGF-1:  is a primary mediator of the effects of growth hormone ; can also regulate cellular DNA synthesis105 ng/mL
Platelet-derived growth factor PDGR: enhances collagen synthesis and bone cells proliferation14 ng/mL
Quantitative assessment of the main growth factors, cytokines, and chemokines in PRGF fraction. Copyright Simona Cavalu et al.
Cross-sectional scanning electron microscopy (SEM) images of different commercial collagen membranes before (a,d,g) and after (b,e,h) PRGF treatment; AFM 3D topography of the membrane surface after PRGF treatment (c,f,i) showing the details of collagen fibers. The images correspond to Biocollagen® (ac), CovaTM Max (df), and Jason® (gi). Copyright Simona Cavalu et al.
Nanoindentation measurements: load–displacement curves recorded for each membrane before (a,c,e) and after (b,d,f) PRGF treatment. The images correspond to Jason® (a,b), Biocollagen® (c,d), and CovaTM Max (e,f). Legend: MC1/MC2 = Jason® membrane before/after PRGF treatment; MCP1/MCP2 = Biocollagen® membrane before/after PRGF treatment; MPP1/MPP2 = CovaTM Max membrane before/after PRGF treatment. Copyright Simona Cavalu et al.
Young modulus calculations with respect to the three collagen membranes before (a) and after PRGF treatment (b). Legend: MPP1/MPP2 = CovaTMMax membrane before/after PRGF treatment; MC1/MC2 = Jason® membrane before/after PRGF treatment; MCP1/MCP2 = Biocollagen® membrane before/after PRGF treatment. Copyright Simona Cavalu et al.
Results of enzymatic degradation test of native (unmodified) and PRGF-modified collagen membranes. Copyright Simona Cavalu et al.

PRGF-modified collagen membranes investigated in our study present new evidence of several advantages, with respect to a rapid and predictable soft tissue healing. The structural and morphological features of three different commercial collagen membranes for GBG/GTR were investigated upon PRGF treatment, revealing that particular characteristics such as porosity, fiber density, and surface topography may influence the mechanical behavior and performance of the membranes. By FTIR spectroscopy, it was demonstrated that the collagen matrix may act as a natural reservoir for growth factor delivery. Nanoindentation measurements revealed that, upon PRGF treatment, the changes of Young modulus values are correlated with the ultrastructural properties of each membrane type, especially the porosity. The mechanical properties of the membranes were analyzed in a comparative manner, before and after PRGF modification. The enzymatic (trypsin) degradation test also emphasized a different behavior—PRGF-modified membranes exhibited a slower degradation compared with the native ones. Within the limitations of the present study, the results are important with respect to the regulation and kinetic release of multiple growth factors that can be adapted to specific therapeutic conditions. Copyright Simona Cavalu et al.

Carbon Nanotubes for Improved Performances of Endodontic Sealer

By Simona Cavalu et al.

In order to overcome the limitations of current endodontic sealers, especially against resistant bacteria, recent developments in the field of nanotechnology have proved the necessity to reconsider the composition and physico-chemical properties of classical sealers. Nanoparticles with their unique features in terms of small size and high specific surface area, are the best choice for incorporation of antiseptic agents and effective delivery. Multi-walled carbon nanotubes (MWCNTs) encapsulating chlorhexidine (CHX) and colloidal silver nanoparticles (AgNPs) were prepared and incorporated into commercial sealer and investigated in terms of bonding performance to dentin and effectiveness against E. faecalisS. aureus and Candida albicans, which are responsible for the majority of the failures in endodontic treatments. In this context, the challenges related to the long-term biological effects of CHX/AgNPs loaded MWCNTs are discussed.

Development of “smart” endodontic therapeutic agents 

SEM morphological details of CNTs. Copyright Simona Cavalu
SEM morphological details of CNTs as received from the manufacturer, before (a) and after (b) loading with the mixture CHX/AgNPs (High magnification, 100,000×).
Copyright Simona Cavalu et al.
Colloidal CNTs in distilled water (left) compared to colloidal mixture CHX/AgNPs. Copyright Simona Cavalu
SEM morphological details of CNTs as received from the manufacturer, before (a) and after (b) loading with the mixture CHX/AgNPs (High magnification, 100,000×); (c) photographic image of colloidal CNTs in distilled water (left) compared to colloidal mixture CHX/AgNPs (right). Copyright Simona Cavalu et al.
SEM  details of commercial  and modified root canal sealer. Copyright Simona Cavalu
Ultrastructure details of commercial (a) and modified root canal sealer (c) along with the corresponding EDX spectrum (b,d). Copyright Simona Cavalu et al.
Load–displacement curves and Young modulus calculation. Copyright Simona Cavalu.
Load–displacement curves (a) recorded on the surface of commercial (black) and modified sealer (red) and the corresponding Young modulus calculation (b). Copyright Simona Cavalu et al.
TGA and DTG thermograms of the modified and commercial sealers.
Copyright Simona Cavalu et al.
SEM images of interfacial adaptation between sealer and root canal dentine. Copyright Simona Cavalu
SEM images of interfacial adaptation between sealer and root canal dentine. Copyright Simona Cavalu
SEM images (a,c) of interfacial adaptation between sealer and root canal dentine (polished specimens) along with the corresponding EDX spectra (b,d): (a) neat sealer;
(c) CNTs/CHX/AgNPs modified sealer. The transversal section was performed in the middle zone of the root. Copyright Simona Cavalu et al.
Antimicrobial and antifungal effect of different combinations and the mixture CNTs/AgNPs/CHX2%
Antimicrobial and antifungal effect of different combinations and the mixture CNTs/AgNPs/CHX2% against the tested strains. Data are expressed as average value ± standard deviation of triplicate samples (statistical significance * p < 0.05).
Copyright Simona Cavalu

Our original approach, in the context of new generation sealers expecting to have a long-lasting antimicrobial effect, was to demonstrate that the antimicrobial effect of the mixture CNTs/AgNPs/CXH 2% incorporated in commercial sealer, was preserved long enough to efficiently inhibit Gram-positive germs, with excellent results towards E. faecalis in a concentration of 1 mg/mL. The antibacterial and antifungal assay clearly demonstrated a synergic effect of AgNPs, CHX 2% and CNTs with excellent results towards E. faecalis, which is responsible for the primary etiologic factors in pulp and periapical lesions.

The full text of this paper is available at

https://www.mdpi.com/1996-1944/14/15/4248/htm