Magister Chirurgiae Dentium (MChD) / Wound healing is a complex process characterised by the repair and reconstitution of lost or damaged tissue. By the mid 1990s, several methods were proposed to enhance wound healing of surgical sites by introducing high concentrations of human platelets to these areas. In the early 21st century, Choukroun et al (2006b) introduced a new type of platelet concentrate that was devoid of any additives, and required no specialised equipment for its production. This concentrate was termed Platelet-rich fibrin (PRF) and although various aspects of this biomaterial had been studied, very little is currently known about its storage properties. Aim: To determine whether storage time had a significant effect on the platelet concentration of Choukroun’s PRF Method: A total of 30 patients were enrolled into the study. Three blood samples of 10ml each were drawn from each patient. Two of the blood samples (Group A and Group B) were centrifuged to form PRF. The third sample was used to measure the baseline blood platelet concentration and was therefore not centrifuged. After PRF had formed in both test groups, it was removed from the test tubes at 2 different times i.e. immediately after centrifuge (Group A) or after 60 min of storage in the blood collecting tube (Group B). The remaining blood was then tested for platelet concentration and compared to each other and the baseline reading. Results: 14 males and 16 females participated in the study (average age 41.7 years). A mean blood platelet concentration of 282.8 ± 58.3 × 109/L was recorded for the baseline reading. Group A had a mean blood platelet concentration 7.9 ± 3.03 × 109/L. Group B had a mean blood platelet concentration of 4.0 ± 1.93 × 109/L. A statistically significant difference was seen between Groups A and B (p < 0.0001). Conclusions: Storage time has a significant effect of the platelet concentration of PRF. If stored over a period of 60 min, the platelet concentration of PRF increases. Further research is required to determine whether this finding is clinically significant.

We detail for the first time the uniquely altered fibrin polymerization of homophenotypic Aα R16H dysfibrinogen. By polymerase chain reaction amplification and DNA sequencing, our new proposita's genotype consisted of a G>A transition encoding for Aα R16H, and an 11 kb Aα gene deletion. High-performance liquid chromatography disclosed fibrinopeptide A release approximately six times slower than its fibrinopeptide B. Turbidimetric analyses revealed unimpaired fibrin repolymerization, and abnormal thrombin-induced polymerization (1-7 μmol/l fibrinogen, > 96% coagulable), consisting of a prolonged lag time, slow rate, and abnormal clot turbidity maxima, all varying with thrombin concentration. For example, at 0.2-3 U/ml, the resulting turbidity maxima ranged from lower to higher than normal control values. By scanning electron microscopy, clots formed by 0.3 and 3 thrombin U/ml displayed mean fibril diameters 42 and 254% of the respective control values (n = 400). Virtually no such differences from control values were demonstrable, however, when clots formed in the presence of high ionic strength (μ = 0.30) or of monoclonal antiβ(15-42)IgG. The latter also prolonged the thrombin clotting time approximately three-fold. Additionally, thrombin-induced clots displayed decreased elastic moduli, with G′ values of clots induced by 0.3, 0.7 and 3 thrombin U/ml corresponding to 11, 34, and 45% of control values. The results are consistent with increased des-BB fibrin monomer generation preceding and during polymerization. This limited the inherent gelation delay, decreased the clot stiffness, and enabled a progressively coarser, rather than finer, network induced by increasing thrombin concentrations. We hypothesize that during normal polymerization these constitutive des-BB fibrin monomer properties attenuate their des-AA fibrin counterparts.

Philosophiae Doctor - PhD / Wound healing is a complex process that involves several overlapping and interacting biological pathways. The consequences of delayed or abnormal wound healing may result in tissue formation that has impaired function or structural abnormalities. As a result, clinicians have sought ways to enhance this process. Recently, the use of autologous platelet concentrates have become popular in the management of wound healing sites. However, controversy exists as to how these biomaterials should be prepared and applied. We therefore sought to investigate whether a biologically viable and clinically effective platelet concentrate could be prepared using standard laboratory equipment. The findings are presented in a series of articles that have been published in peer-reviewed journals. The results suggest that the experimental platelet concentrate produced, has a morphological structure that consists of a dense fibrin network intermingled with platelets, has the ability to accelerate cellular growth in-vitro, has no adverse effects on cells in-vitro, can concentrate and release a systemically ingested antibiotic over a period of 24 hours in-vitro, can be stored for at least 60 minutes without showing signs of deterioration, and has shown clinical evidence of accelerating wound healing in sinus augmentation and alveolar ridge preservation procedures. The reduced cost of producing such a biomaterial allows it to be available to resource poor settings and to wider range of healthcare providers as compared to standard platelet concentration techniques. Further studies are required to investigate the clinical potential of this promising biomaterial.

The purpose of this study was to develop a fibrin gel system capable of serving as a three dimensional scaffold for the chondrogenesis of rabbit bone marrow mesenchymal stem cells (BM-MSCs) and to examine the effect of two fibrinolytic inhibitors, aprotinin and aminohexanoic acid, on this system. Rabbit BM-MSCs were obtained from the tibias and femurs of New Zealand white rabbits. After chondrogenic potential of BM-MSCs was verified by pellet culture, 2 x 106 cells were pelleted and suspended in fibrinogen (80mg/ml) and then mixed with equal parts of thrombin (5 IU/ml). The specimen were then divided into four groups: aprotinin control (with aprotinin); aprotinin + transforming growth factor (TGF-beta) (with aprotinin and TGF-beta 1); amino control (with aminohexanoic acid); and amino+TGF-beta (with aminohexanoic acid and TGF- beta1). Each of these groups was further divided into three groups depending on the concentration of the inhibitor. Both of the aprotinin groups received 0.0875, 0.175, or 0.35 TIU/ml of aprotinin and both of the aminohexanoic acid groups were supplemented with 2, 4, or 8 mg/ml of aminohexanoic acid. The gels were harvested and analyzed at 7, 14, and 21 days. All of the aprotinin+TGF-beta groups exhibited a significantly higher aggrecan gene expression than control groups whereas only the amino+TGF-â group treated with 8mg/ml was significantly higher than those of the control groups. In addition, the 0.0875 and 0.175 TIU/ml aprotinin+TGF-beta groups exhibited significantly higher levels of expression than the 2 and 4 mg/ml amino+TGF-beta groups. There were no significant differences among the different concentrations of aprotinin or aminohexanoic acid with or without the treatment of TGF-beta. Similar trends were also seen when the glycosaminoglycan (GAG) content was measured and analyzed. These findings suggest that fibrin gels are a suitable environment for the chondrogenesis of BM-MSCs and that aprotinin in combination with TGF-beta1 is the optimal condition for stimulating BM-MSCs to differentiate into chondrocytes.

No description available.

Thesis (M.S.)--Worcester Polytechnic Institute. / Keywords: myocardium; microthreads; fibrin; infarct; human mesenchymal stem cells. Includes bibliographical references (leaves 71-77).

Rheology is a discipline of continuum mechanics that is concerned with the mechanical properties of matter as it flows. Key to the study of rheology is the concept that materials do not behave as Newtonian liquids of as heterogenous, homogenous mateirials, but as a combination of the two. This combination and blurring of the line between liquid and solid peoperties is knows as viscosity. Furthermore, the viscosity of a material or liquid will not necessarily remain constant when it is subjected input forces or stresses at different frequencies. This consideration brings with it the idea of viscoelasticity which can account for the variations in the characteristics of a sample medium.Magnetic tweezers are tools that allow examination of and investigation into the viscoelastic properties of a sample on the mesoscopic scale. Magnetic matter can be inserted into and bound onto a sample. This magnetic matter can then be manipulated using an external magnetic or electromagnetic field. Calibrated magnetic tweezers apparatus can be used to investigate the mechanical and viscoelastic properties of a material with the novel application of time-variant forcesand stresses. The resultant behaviour, or response, of the sample can be observed using a microscope and analysed further.Fibrin is the highly extensible, fibre-like protein that makes up blood clots. Its particularly high levels of extensibility combined with interesting material properties such as viscoelasticity can strain-hardening make it an ideal test sample for magnetic tweezers experiments. The high elastic limit of fibrin ensures that plastic deformation does not usually occur under the range of input forces and stresses exerted by magnetic tweezers. This allows non-destructive and repeatable tests to be performed.Magnetic tweezers have been developed and used in a series of experiments on fibrin to produce a viscoelastic characterisation of the fibrous networks. The key results in this work are the design of high-sensitivity apparatus for the experiments and associated techniques for high-frequency analysis, use of the tweezers with a high-speed CCD attached to a microscope and the analysis of the viscoelastic properties of fibrin over a several decades of frequency.

Fibrin is the body's natural provisional matrix activated in response to vascular injury in which noncovalent knob:hole interactions between fibrin monomers lead to the assembly of fibrin for clot formation. In this study we aimed to exploit fibrin knob:hole affinity interactions with swelling, space filling microgels for the development of a potential bio-synthetic hybrid polymer system with hemostatic properties. Previous work has explored the inherent binding interactions of various fibrin knobs and their complementary polymerization holes, which have led to the development of fibrin knob peptide mimic (GPRPFPAC) with enhanced binding affinity for fibrin(ogen) holes. By coupling this enhanced fibrinogen binding peptide with a pNIPAm microgel system capable of being dynamically tuned and self-assembled, we hypothesized the specific and rapidly triggered formation of a bulk hydrogel in a wound environment (i.e. in the presence of fibrinogen). We found that at the peptide ligand density and concentrations of microgels used, that a rapid formation of a gel did not occur in the presence of fibrinogen alone. However with fibrinogen and thrombin, we found that fibrin network polymerization, structure, and viscoelastic properties were greatly altered in the presence of knob peptide-conjugated microgels.

Specific objective: Guided tissue regeneration (GTR) aims to regenerate the lost attachment apparatus caused by periodontal disease through the use of a barrier membrane. For the GTR procedures to be successful, barrier membranes are required to be present at the surgical site for an extended period of time (weeks to months). Synthetic membranes have the advantage of prolonged presence in a wound site; however, they do not actively contribute to wound healing. Biologic membranes are recognized by the host tissue and participate in wound healing but have the disadvantage of early resorption. Therefore, the goal of this study is to create and characterize a hybrid barrier membrane that contains biologically active fibrin matrix within a synthetic polymeric electrospun scaffold. Method: Fibrin matrices and fibrin-incorporated electrospun scaffold were created from fresh frozen plasma at three different centrifugation conditions 400g for 12 minutes, 1450g for 15 minutes and 3000g for 60 minutes. Following centrifugation, half of the membranes were crosslinked with 1% genipin. Biological stability of these scaffolds was evaluated by resistance to trypsin while their mechanical properties were characterized by MTS Bionix Uniaxial Tensile Testing System. Continuous data was analyzed by ANOVA to detect differences between groups (p=0.05). Results: The addition of an electrospun scaffold to the fibrin matrix led to improvements in the mechanical properties as evidenced by an increase in the modulus (p<0.0001), strain at break (p<0.0001) and energy to break (p<0.0001). The effect of crosslinking was marginal but not statistically significant to the mechanical properties of fibrin matrices or the fibrin incorporated scaffold. However, crosslinking did significantly increase resistance against enzymatic degradation by trypsin (p<0.0001). Lastly, centrifugation speeds at 400g and 1450g showed similar mechanical properties and biologic stability; meanwhile 3000g negatively impacted the properties of the scaffold. Conclusion: Fibrin-incorporated electronspun scaffold exhibits enhanced mechanical and biologic stability compared to fibrin matrices alone. Moreover, crosslinking improves the biologic stability of the novel biomaterial. All these characteristics of the fibrin-incorporated matrix make this membrane a potentially more ideal barrier for GTR procedures to enhance periodontal wound healing.

A research report submitted to the Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, in partial fulfilment of the requirements for the degree of Master of Science Johannesburg, 2012 / This experiment was designed to evaluate whether an allogeneic fibrin-fibronectin protein concentrate (AFFP) can not only prevent ankylosis and root resorption of autotransplanted roots during healing but contribute to regenerate a periodontal attachment as well. In two adult male baboons (Papio ursinus), four horizontal alveoli, 2 to 3 mm deep, were prepared bilaterally in the buccal alveolar and basal bone adjacent to the first and second mandibular molars to receive the roots of the adjacent two molars. Following hemisection, the first and second mandibular molars were extracted, the coronal two-thirds of the roots were planed to remove the remnants of the periodontal ligament and cementum and a notch was placed at the junction between the planed and non-planed surfaces. The planed surfaces were demineralised with citric acid at pH 1 for 3 min. Before transplantation, the crowns were resected and the experimental roots and alveoli were coated with the AFFP prepared from pooled fresh-frozen baboon plasma. The animals were killed 55 days after the transplantations. Histometrical evaluation was performed on serial sections cut in a bucco-lingual direction parallel to the long axis of the transplanted roots. An analysis of variance, in relation to the extent of ankylosis and root resorption, revealed minimal differences between the treatments of experimental and control roots both in the planed and non-planed sections. In this primate autotransplantation model, the treatment with AFFP did not prevent ankylosis and root resorption and did not result in the establishment of a new periodontal attachment.