The Attachment Of Fibroblast Cells

Published Date: 02 Nov 2017

The growth and the attachment of fibroblast cells (L-929) on the matrices of the silk fibroin from domestic silkworm (DSF) and wild silkworm (WSF) were investigated by a cell culture method. The performance of the two types of silk fibroin was compared to that of collagen. The DSF exhibited as high a cell growth and attachment as collagen did. The cells attached to DSF were broadly spread out and their filopodia were visible in the SEM images. The WSF which contains the tripeptide sequence Arg-Gly-Asp (believed to be a specific interaction site for cell-attachment), displayed much higher cell attachment and growth compared to the DSF. The cells attached on WSF became virtually flat and their filopodia could be seen, the results shown that the cells were very strongly held on the surface. [89]

The aim of this study was to modify the surface of poly(d,l-lactic acid) (PDLLA) with different molecular weight of the silk fibroins, and assess the effects of the modified surfaces on the functions of the rat osteoblasts cultured in vitro. The properties of the modified PDLLA surface and the control one were improved by contact angle and electron spectroscopy for chemical analysis (ESCA). The former was indicated the hydrophobicity variation and the last shown that the modified PDLLA film which it is using silk fibroin is enriched with the nitrogen atoms. The PDLLA film biocompatibility may be was altered and in the turn affects the seeded cell functions also, the proliferation and the attachment of osteoblasts seeded on the modified PDLLA films and the control one were checked. The Cell morphologies on these films were examed by the scanning electron microscopy (SEM) and also the cell viability was evaluated by MTT assay. By measuring the alkaline phosphatase (ALP) activity the differentiated cell function was assessed. These results showed that the silk fibroin-modified PDLLA surface can improve the interaction between the PDLLA films and osteoblasts. [90]

The growth and attachment of L-929 cells on films made of Bombyx mori silk fibroin proteins was studied by the cell culture method. Both of the growth and cell attachment were dependent on the minimum of around 90% sericin in the mixture. The results from electron micrography and also from the DSC measurements supported the notion that the mixture of the both types of proteins which it is fibroin and sericin has a phase-separated structure in the solid state. The observed minimum amount of the sericin in the cell attachment and growth is thought to be as result of this phase-separated structure. Films which it had pure component proteins exhibited as high a cell attachment and growth as the collagen, the mammalian cell culture substrate usually was used. The morphological study of the attached cells revealed that the cells attached to silk fibroin were extended and formed in a spindle shape, just like the cells attached to collagen, while the cells attached to the silk sericin had a different shape. Duo to these results the attachment condition on silk fibroin is ideal for the growth, viability, and function of the cells. [92]

Silk fibroin hydrogels is prepared either by treating a 2% (w/v) silk fibroin aqeuous solution at 4°C (thermo gel) or by adding 30% (v/v) of glycerol (glygel),and they were characterized by using Spectroscopy (FT-IR), Fourier Transform Infrared, Environmental Scanning Electron Microscopy (ESEM), Differential Scanning Calorimetry (DSC), Thermogravimetrical Analysis (TGA) and molecular weight determination. The preparation procedure affected the morphology and the molecular weight of the hydrogels, with no or negligible differences being displayed by DSC and FT-IR analyses. While the thermo gel gave a well uniform porous structure, the morphology of glygel appeared to be heterogeneous and non-porous .the Glygel presented lower degradation temperatures, and lower water content, associated with the presence of glycerol but likely also to less-organized protein structures.the Cytoxicity tests with human osteoblast-like cells showed that both gels were not cytoxic for human , while the cell cultures pointed out a faster cell proliferation on glygel and differentiation on thermgel and a higher cell activation. These results shown that these gels could be used as scaffolds able to promote in situ bone regeneration and wide range applications of tissue engineering. [93] Silk fibrion are being reassessed as biomaterial scaffolds or films due to their unique mechanical properties, opportunities for genetic tailoring of structure and thus function, and the studies clarifying the biocompatibility. The report was presented on the covalent decoration of silk fibrion films with integrin recognition sequences (RGD) and also with the parathyroid hormone (PTH, 1–34 amino acids) and a modified PTH 1–34 (mPTH) involved in the induction of bone formation. Osteoblast-like cell (Saos-2) responses to the decorated silk fibrion films indicate that the proteins serve as suitable bone-inducing matrices. Osteoblast-like cell adhesion was significantly increased on PTH and RGD compared to mPTH, plastic, and the control peptide RAD. After 2 weeks of culture, message levels of the alkaline phosphatase were similar on all substrates, but after 4 weeks, the alkaline phosphatase mRNA was greatest on the RGD. After 2 weeks of culture, α1(I) procollagen mRNA was elevated on the silk fibroin, RAD, RGD, and PTH, and hardly detectable on mPTH and plastic. After 4 weeks RGD demonstrated the highest level compared to the other substrates. Osteocalcin message levels were detected by RT-PCR were greatest on RGD at both time of the points. The Calcification was also significantly elevated on the RGD compared to the other substrates with an increase in size and the number of the mineralized nodules in culture. The RGD covalently decorated silk fibroin appears to the stimulate osteoblast-based mineralization in vitro. [94]

There are Three forms of silk fibroin (SF) matrices, film form, woven (microfiber)form, non-and woven (nanofiber)form, were it are used to perform a conformational analysis and cell culture using normal human oral keratinocytes (NHOK). The silk was degummed To obtain the SF microfiber (SF-M) matrix,while the SF film (SF-F) and nanofiber (SF-N) matrices were prepared by casting and electrospinning the formic acid solutions of the regenerated Silk Fibroin, . For desolubilization, as-prepared SF-F and SF-N matrices were chemically treated by the methanol solution of 50%. The conformational structures of as-prepared and chemically treated SF matrices were investigated by using attenuated total reflectance infrared spectroscopy (ATR-IR) and solid state 13C CP/MAS nuclear magnetic resonance (NMR) spectroscopy. The as-cast SF-F matrix formed a mainly β-sheet structure that was so similar to the SF-M matrix structure, while the as-spun SF-N matrix was had a random coil conformation as the predominant secondary structure. The aqueous methanol treatment leaded to Conformational transitions from random coil to β-sheet of the as-spun SF-N occurred rapidly within less than 10 minute, and were confirmed by the solid state of the 13C NMR analysis. To evaluate the cells behavior and the cytocompatibility on the different textures of silk fibroin, the cell attachment and spreading of NHOK that was seeded onto the SF matrices was examined, as well as the interaction between the SF matrices and the cells. The results indicate that the silk fibroin nanofiber matrix may be preferably more than the SF microfiber matrices and the silk Fibroin film for biomedical applications. [95]