Optimizing a novel method for low intensity ultrasound in chondrogenesis induction

Abstract

Background:Hyaline cartilage tissue of joints is susceptible to injuries due to avascularity. Mesenchymal stem cells (MSCs) are used for cartilage tissue engineering. Among MSCs, adipose stem cells (ASCs) are attractive because of accessibility, their large number, and rapid growth. Common in vitro protocols successfully induce chondrogenic differentiation by expression of multiple cartilage-specific molecules. However, transforming growth factor β (TGFβ) promotes chondrogenesis to terminal stages. Despite much attention being given to the influences of biochemical factors on chondrogenesis of MSCs, few studies have examined the chondrogenic effect of mechanical factors such as ultrasound as a feasible tool.Materials and Methods:In this study, we focused on inducing chondrogenesis in the early stages of differentiation by using low-intensity ultrasound (LIUS). Four groups of ASC pellets (control, ultrasound, TGFβ, and ultrasound/TGF) were cultured under chondrogenic (10 ng/ml of TGFβ3) and ultrasound conditions (200 mW/cm2, 10 min/day). After 2 weeks, differentiation was evaluated by histology, quantitative gene expression analysis, and immunohistochemistry.Results:Our data demonstrated that ultrasound differentiated pellets showed increased expression of early chondrogenesis marker, Col2A, than those in TGFβ groups (P < 0.001), and Col2B and Col10 expression were more prominent in TGFβ groups. Immunostaining of sections showed Col2 fibrils around lacuna in LIUS and TGFβ treated groups.Conclusion:Using LIUS resulted in early chondrogenesis in comparison with terminally differentiated chondrocytes by TGFβ. Therefore, LIUS might provide an applicable, safe, efficient, and cheap tool for chondrogenic differentiation of ASCs in cartilage tissue engineering.

An Investigation on the Regenerative Effects of Intra Articular Injection of Co-Cultured Adipose Derived Stem Cells with Chondron for Treatment of Induced Osteoarthritis

“… 25 , 59 The utilization of ASCs in cartilage tissue engineering is a rapidly growing area of research and some evidence of therapeutic achievement using ASCs for OA defect has been reported. 48 , 60 , 61 In general, there have been fewer investigations on co-culture of chondrons with mesenchymal stromal cells (MSCs) in in-vivo systems. 62 These studies are dividable in two categories.…”

“…After reaching 80% confluence, the cells were treated with %0.25 Trypsin and 0.02 mM EDTA (Gibco, 25200-056) solution and the cells reseeded under the same conditions for expansion as passage 1. 48 …”

Effects of Low Intensity Ultrasound on the Chondrogenic Differentiation of Adult Stem Cells From Adipose Tissue

“…study confirms our pervious collagen type II protein expression results. LIUS induces production of early stage extracelluar matrix of cartilage and more collagen type IIA than collagen type IIB [35]. In cartilage development collagen type IIA is the splice variant of type II collagen that has been found in prechondrocytes and immature chondrocytes [43].…”

“…As ultrasound has some advantages such as lowcost, safe in comparison to other mechanical forces and also ultrasound is easy to apply for possible future clinical applications [34], we applied LIUS for inducing chondrogenesis in ASC. We showed that LIUS as a physical inducer work better than TGFβ, in differentiating adipose tissue adult stem cells into chondrocytes by expression of collagen type 2 and aggrecan [35].…”

“…The low intensity ultrasound (LIUS) device (Novin, Iran) with continuous wave at 1 MHz and intensity of 200 mW/cm 2 was applied for 10 minutes per day in ultrasound cultures [35,38]. The distance between transducer and cultures was determined by the Sarvazyan method [39][40][41].…”