Enjekte Edilebilir Hemostatik Fibroin Mikrojellerin Geliştirilmesİ

Abstract

Recently, developing of homeostatic wound dressings and biomaterials capable of stopping the bleeding have been the important research subjects of military and civil trauma researchers of developed countries. At the present time, the increase in the traffic and work accidents, still continuing wars and inadequacies in the field of surgical operation are increasing the frequency of traumatic injuries. Conventionally, in order to stop the uncontrollable bleedings after traumatic injuries, gauze or different homeostatic wound dressings are being used. However, commercially available homeostatic wound dressings are insufficient in terms of getting the shape of the wound area, contributing to the wound healing process, stopping the bleeding safely in every region of the body and are containing side effects such as lack of heat increase for recovery process and clotting disorder. For this reason, a new generation of hemostatic biomaterials is needed to be developed in order to safely stop any uncontrolled bleeding. The aim of this thesis is to develop an injectable fibroin based hemostatic microgel that capable of stopping the bleeding in an effective way in traumatic injuries in which there is an uncontrollable bleeding. In this regard, low molecular weight of silk fibroin protein (200 kDa) was prepared by extraction from B. mori cocoons. Then, by using the polymeric conformational transitions (Silk I and Silk II) of silk fibroin, fibrinogen or Vitamin K containing fibroin microgels less than 5 micron was prepared by phase separation methods without using any surfactant or organic solvent. Thus, fibroin microgels were extensively characterizated in terms of chemical, morphological, thermal, swelling properties. Moreover, rheological measurements were employed to assess the viscoelastic behavior of the microgels. Adaptability to the injectable form was studied and optimized by applying shear stress to the microgells prepared microgels. Then, their biocompatibility was aproved by assessing cell viability. Blood coagulation tests was performed to examine the hemostatic performance of the microgels.

It has been observed that all microgels were in stable Silk II (β-crystallin) conformation and exhibit thermal stability and high biocompatibility. Among them, fibrinogen conjugate microgels (Fibgel-FngC) had the highest thermal stability. The rheological test results demonstrated that yielding storage (G′) for high molecular weight fibroin microgel (HM-Fibgel) and fibrinogen conjugated fibroin microgels (Fibgel-FngC, Fibgel-KFngC) was higher than loss modulus (G″), indicating the typical gel state. In contrast, G′ was lower than G″ in low molecular weight fibroin microgel (Fibgel) and Vitamin K loaded microgel (Fibgel-K), indicating that they still maintained the sol state. For fibrinogen loaded fibroin (Fibgel-Fng) samples, G′ was slightly lower than G″. The current findings suggest that HM-Fibgel and fibrinogen conjugated microgels (Fibgel-FngC, Fibgel-KFngC) exhibit viscoelastic properties and these microgels which gives a fully injectable. Cyctotoxicity test indicated that they are nontoxic on L929 fibroblast cells and also microgels induce cell attachment and proliferation. Further, the blood on the all microgels clotted with variying degree after 60 s, whereas fresh blood did not clot within 60 s. These results indicated that all fibroin based microjel exhibit hemostatic activity.

In conclusion, injectable hemostatic fibroin microgel was successfully developed. Also, in terms of getting the shape of the applied wound, it will improve the patient compliance. Further, due to its protein structure and morphological properties, by initiating the wound healing process, it will have a critical importance for the rapid healing of the damaged tissue.