*Article* **Roles of Silk Fibroin on Characteristics of Hyaluronic Acid**/**Silk Fibroin Hydrogels for Tissue Engineering of Nucleus Pulposus**

#### **Tze-Wen Chung 1,2,\*, Weng-Pin Chen 3,4,\*, Pei-Wen Tai 1,**†**, Hsin-Yu Lo 1 and Ting-Ya Wu 1**


Received: 28 February 2020; Accepted: 11 June 2020; Published: 17 June 2020

**Abstract:** Silk fibroin (SF) and hyaluronic acid (HA) were crosslinked by horseradish peroxidase (HRP)/H2O2, and 1,4-Butanediol di-glycidyl ether (BDDE), respectively, to produce HA/SF-IPN (interpenetration network) (HS-IPN) hydrogels. HS-IPN hydrogels consisted of a SF strain with a high content of tyrosine (e.g., strain A) increased viscoelastic modules compared with those with low contents (e.g., strain B and C). Increasing the quantities of SF in HS-IPN hydrogels (e.g., HS7-IPN hydrogels with weight ratio of HA/SF, 5:7) increased viscoelastic modules of the hydrogels. In addition, the mean pores size of scaffolds of the model hydrogels were around 38.96 ± 5.05 μm which was between those of scaffolds H and S hydrogels. Since the viscoelastic modulus of the HS7-IPN hydrogel were similar to those of human nucleus pulposus (NP), it was chosen as the model hydrogel for examining the differentiation of human bone marrow-derived mesenchymal stem cell (hBMSC) to NP. The differentiation of hBMSC induced by transforming growth factor β3 (TGF-β3) in the model hydrogels to NP cells for 7 d significantly enhanced the expressions of glycosaminoglycan (GAG) and collagen type II, and gene expressions of aggrecan and collagen type II while decreased collagen type I compared with those in cultural wells. In summary, the model hydrogels consisted of SF of strain A, and high concentrations of SF showed the highest viscoelastic modulus than those of others produced in this study, and the model hydrogels promoted the differentiation of hBMSC to NP cells.

**Keywords:** silk fibroin; hyaluronic acid; Tyrosine; viscoelastic modulus of HS-IPN hydrogels; hBMSC differentiations; nucleus pulposus
