Non-Mulberry and Mulberry Silk Protein Sericins as Potential Media Supplement for Animal Cell Culture

Abstract

Silk protein sericins, in the recent years, find application in cosmetics and pharmaceuticals and as biomaterials. We investigate the potential of sericin, extracted from both mulberry Bombyx mori and different non-mulberry sources, namely, tropical tasar, Antheraea mylitta; muga, Antheraea assama; and eri, Samia ricini, as growth supplement in serum-free culture medium. Sericin supplemented media containing different concentrations of sericins from the different species are examined for attachment, growth, proliferation, and morphology of fibrosarcoma cells. The optimum sericin supplementation seems to vary with the source of sericins. The results indicate that all the sericins promote the growth of L929 cells in serum-free culture media; however, S. ricini sericin seems to promote better growth of cells amongst other non-mulberry sericins.

Figure 1

Degumming of silk cocoons of different species of silkworms using different methods of isolation of sericins.

Figure 2

Scanning electron micrographs of the cocoon pieces of mulberry and non-mulberry silks. The cocoons are observed before (50x) and after degumming (100x) using urea and autoclave degumming methods. Scale bar represents 100 μm.

Figure 3

SDS-PAGE (8%) analysis of 0.1% sericin solutions from cocoons of A. mylitta, B. mori, S. ricini, and A. assama: (a) isolated by autoclave method: Lane 1: sericin hope, Lane 2: B. mori, Lane 3: S. ricini, Lane 4: A. assama, and Lane 5: A. mylitta; (b) isolated by urea method: Lane 1: sericin hope, Lane 2: B. mori, Lane 3: S. ricini, and Lane 4: A. mylitta. (c) Lane 1: A. assama isolated by urea method. The protein molecular weight standards are indicated by the numbers on the left. M: molecular weight marker.

Figure 4

(a) CD spectra of 0.1 % (w/v) sericin solution from cocoons of different species: (A) A. mylitta, (B) A. assama, (C) S. ricini, and (D) B. mori. (b) FTIR spectrum of sericin powders from the various species: (A) A. mylitta, (B) A. assama, (C) S. ricini, and (D) B. mori. (c) TGA curves of lyophilized sericin powders of (A) S. ricini, (B) A. mylitta, (C) A. assama, and (D) B. mori.

Figure 5

Time-dependent attachment of L929 cells growing in DMEM supplemented with 0.05% sericin of B. mori, A. mylitta, A. assama, and S. ricini sericin. Cells grown in DMEM supplemented with 10% serum and without serum were used as controls (error bars denote standard deviation for n = 3).

Figure 6

(A) Phase contrast and (B) fluorescence microscopic observations showing L929 fibroblasts growth and attachment on (a) 10% serum supplemented DMEM and (b) serum-free DMEM as controls and in DMEM supplemented with 0.05% silk protein sericins of B. mori (c), A. mylitta (d), A. assama (e), and S. ricini (f). Scale bar represents 10 μm.

Figure 7

Estimation of cell viability and proliferation by Alamar Blue assay on L929 cells grown on 0.1% (a) and 0.05% (b) concentrations of sericin of B. mori, A. mylitta, A. assama, and S. ricini, 10% FBS supplemented DMEM and FBS free DMEM (error bars denote standard deviation for n = 3).