ABSTRACT
BACKGROUND: In the previous study, we have successfully prepared a silk fibroin/curcumin composite film, which possesses good physicochemical properties, sustained-release properties, antibacterial activity and biocompatibility. OBJECTIVE: To further observe the effect of the prepared silk fibroin/curcumin composite film, as the wound dressing, in the field of wound healing. METHODS: The silk fibroin/curcumin composite film was prepared by referring to the previously established methods. The mechanical properties and light transmittance of the prepared composite film were then investigated. Escherichia coli and Staphylococcus aureus were used as strains. The plate-counting technique was applied to evaluate the antibacterial activity of the light illuminated silk fibroin/curcumin composite film, silk fibroin/curcumin composite film without light exposure, and silk fibroin film without light exposure. The full-thickness excisional lesions were created on the dorsum of 20 Kunming mice. The wounds of 10 mice were covered with silk fibroin/curcumin composite film, and the wounds of other 10 mice were dressed with sterile gauze. The healing status and histological morphology of the wound sites were observed at 3, 7, and 14 days post operation. The animal experimental design and the protocols were approved by the Laboratory Animal Ethics Committee of Southwest University (approval number: IACUC-2019011). RESULTS AND CONCLUSION: (1) Compared to silk fibroin film, the fracture strength of silk fibroin/curcumin composite film increased significantly (P < 0.05), while the Young’s modulus of silk fibroin/curcumin composite film decreased significantly (P < 0.01). (2) Both silk fibroin film and the silk fibroin/curcumin composite film exhibited excellent transparency, but the silk fibroin/curcumin composite film showed a stronger absorption in visible light region at a wavelength of 400-450 nm. (3) Without light illumination, the silk fibroin/curcumin composite film exhibited stronger antibacterial activity than that of silk fibroin film. While, the silk fibroin/curcumin composite film with light illumination demonstrated stronger antibacterial activity than that of silk fibroin/curcumin composite film without light illumination. (4) The wound healing rate of the experimental group was higher than that of the control group at each time point (P <0.05). Analysis on histological morphology reveals that the length of regenerated epithelia of dermal tissue and the thickness of granulation tissue in the experimental group were greater than those in the control group at 3 and 7 days post operation (P < 0.01). While, the thickness of granulation tissue was smaller than that in the control group at 14 days post operation (P < 0.01). (5) The results indicated that the silk fibroin/curcumin composite film can facilitate skin wound healing.
ABSTRACT
In order to provide a basic theory for the materials of repairing central nervous system injury, we have studied the growth and differentiation of neural stem cells (NSCs) on poly (L-lysine) modified silk fibroin film. First, we used poly (L-lysine) to modify silk fibroin film and confirmed by UV-vis and 1H NMR spectra. Then NSCs were isolated and seeded on the silk fibroin film (Silk group), poly (L-lysine) (PLL group) and poly (L-lysine) modified Silk fibroin film (Silk-PIL group). The proliferation of NSCs was evaluated by Cell Counting Kit-8 (CCK-8) assay on days 1, 3, 5 and 7 after seeding. Immunofluorescence was used to analyze the differentiation of NSCs at the 7th day. The levels of apoptosis were detected by Western blotting and TUNEL. The mRNA level of brain derived neurotrophic factor (BDNF) was identified by real-time PCR. UV-vis and 1H NMR spectra confirmed that poly (L-lysine) was successfully grafted onto the silk fibroin film. From the 3rd day after seeding to the 7th day, the CCK-8 test showed that proliferation rate of NSCs in the Silk-PIL was significantly higher than Silk group (P<0.05) but had no significant difference compared with PLL group (P>0.05). Immunofluorescence staining displayed that more NSCs in Silk-PIL group were differentiated into neuron compared with Silk group (P<0.05), however, there was no significant difference compared with PLL group (P>0.05). The number of NSCs differentiated into astrocytes was not significantly different between the three groups. Western blotting and TUNEL test presented that the degree of apoptosis of NSCs in the Silk-PIL group was significantly lower than Silk group (P<0.05). RT-PCR exhibited that mRNA level of brain derived neurotrophic factor (BDNF) of NSCs was higher in Silk-PIL group compared with Silk group (P<0.05) but had no significant difference compared with PLL group (P>0.05). Thus, poly (L-lysine) modified silk fibroin film could promote the proliferation of NSCs and reduce NSCs apoptosis. Furthermore, it also can enhance the differentiation of NSCs into neurons. It is expected to become a new type of tissue engineering scaffold carrying NSCs to repair central nervous system injury.
ABSTRACT
Background Biomaterials for corneal tissue engineering must demonstrate several critical features for potentialutility invivo, includingtransparency, mechanicalintegrity, biocompatibilityand slow biodegradation. Silk film biomaterial had been characterized to meet these functional requirements. ObjectiveThis study was to investigate the feasibility of physico-crosslink regenerated silk fibroin film as tissue engineered corneal scaffold. MethodsHuman corneal epithelial cells(CECs) links were cultured by regular method and CECs in logarithmic phase were than incubated on physico-crosslink regenerated silk fibroin film membrane. The shape of cultured human CECs was observed after 24,48 and 72 hours under the inverted microscope and scanning electron microscope( SEM ) ,and the CECs were cultured on culture plates as controls. The growth state of CECs on regenerated silk fibroin film was observed daily for 7 days by MTT, and cell cycle analysis and the presence of apoptosis of human CECs were examined by flow cytometry after incubation on regenerated silk fibroin film. Regenerated silk fibroin filmCECs (4 mm×3 mm) were implanted into the corneal stroma of the right eyes of New Zealand white rabbits. At the end of 4 and 8 weeks after implantation, the appearance of the ocular surface was examined using slit lamp and corneal neovascular area was measured. Corneal histopathological examination was carried out to assess the degradation of graft materials and immunohistochemistry was performed to detect the expression of CD34 in the corneal tissue after operation. ResultsThe morphology and structure of CECs were identical using the two cultured Methods when observed under the inverted microscope and SEM after 24,48 and 72 hours. No significant difference was found in the A490 value 1,2,3,4,5,6 or 7 days after incubation on regenerated silk membrane and in culture plates ( Fmethod =0. 641 ,P＞0.05 ). The apoptosis rates of CECs on regenerated silk membrane or culture plates were 1.8％ and 2.0％ and the amount of cells in G2/G1 phase was 1. 956 and 1. 945, respectively. Histopathological examination showed that the regenerated silk membrane material degraded and was replaced by regular collagen tissue 2 months after implantation,and the presence of neovascular area and inflammatory cells were less prominent in 2 months than 1 month post-implantation. The expression level of CD34 in corneal tissue was evidently lower 1 and 2 months after operation than the Ad-VEGF165-induced positive control group (P＜0. 05), and no significant differences were seen when compared with normal CECs(P＞0.05). ConclusionsPhysico- crosslink regenerated silk fibroin film is an excellent biomaterial for tissue engineered corneal scaffold with good biocompatibility.
ABSTRACT
PURPOSE: The aim of this study was to examine the effects of silk fibroin film on wound healing of cutaneous burn in hairless mice by using microscopic findings and stem cell markers (nestin, cytokeratin 15) and ki-67 proliferation marker. METHODS: Each mouse received two burns at the dorsal area by applying a metal rod heated with boiling water. Burn wound sites were dressed with Silk Fibroin Film and duoderm (SF group), Aquacel hydrofiber and duoderm (AC group) and duoderm only (Control group). All groups were covered externally with duodermas adhesive bands. Those mice were sacrificed at zero, two, seven, fourteen and twenty one days after burn. Histological findings and immunohistochemical staining for stem cell markers were observed. RESULTS: In SF group, inflammatory cell infiltration, formation of granulation tissue and inflammatory foci are greater than in AC and control group. Those factors appear to enhance mesenchymal stem cell markers such as nestin. Finally mesenchymal tissue regeneration was enhanced. In addition, the length of ki-67 expressed re-generating epithelium, which appeared to be associated with epithelial regeneration, was the longest in SF group. CONCLUSION: The results show that the wound healing effect of SF is the best among other treatment materials including AC in the experimental group and duoderm in the control group through mesenchymal regeneration and epithelial regeneration which are essential factors for wound healing.