Cell kinetics, DNA integrity, differentiation, and lipid fingerprinting analysis of rabbit adipose-derived stem cells

Abstract

Human adipose tissue has been described as a potential alternative reservoir for stem cells. Although studies have been performed in rabbits using autologous adipose-derived stem cells (ADSC), these cells have not been well characterized. The primary objectives of this study were to demonstrate the presence of adipose-derived stem cells isolated from rabbit inguinal fat pads and to characterize them through osteogenic and adipogenic in vitro differentiation and lipid fingerprinting analysis. The secondary objective was to evaluate cell behavior through growth kinetics, cell viability, and DNA integrity. Rabbit ADSCs were isolated to determine the in vitro growth kinetics and cell viability. DNA integrity was assessed by an alkaline Comet assay in passages 0 and 5. The osteogenic differentiation was evaluated by Von Kossa, and Alizarin Red S staining and adipogenic differentiation were assessed by Oil Red O staining. Lipid fingerprinting analyses of control, adipogenic, and osteogenic differentiated cells were performed by MALDI-TOF/MS. We demonstrate that rabbit ADSC have a constant growth rate at the early passages, with increased DNA fragmentation at or after passage 5. Rabbit ADSC viability was similar in passages 2 and 5 (90.7% and 86.6%, respectively), but there was a tendency to decreased cellular growth rate after passage 3. The ADSC were characterized by the expression of surface markers such as CD29 (67.4%) and CD44 (89.4%), using CD 45 (0.77%) as a negative control. ADSC from rabbits were successfully isolated form the inguinal region. These cells were capable to differentiate into osteogenic and adipogenic tissue when they were placed in inductive media. After each passage, there was a trend towards decreased cell growth. On the other hand, DNA fragmentation increased at each passage. ADSC had a different lipid profile when placed in control, adipogenic, or osteogenic media.

Periodic injections of adipose-derived stem cell sheets attenuate osteoarthritis progression in an experimental rabbit model

“…More than three passage ADSCs were prepared for the subsequent experiment. Previous reports have confirmed the presence of stemness in ADSCs extracted by this method [6,20].…”

“…However, the entire grafting process remains relatively complicated, time-consuming, and requires special materials. In our study, a simple and inexpensive Vc-mediated procedure was used to obtain ADSC sheets [20].…”

Periodic injections of adipose-derived stem cell sheets attenuate osteoarthritis progression in an experimental rabbit model

“…More than three passage ADSCs were prepared for the subsequent experiment. Previous reports have con rmed the presence of stemness in ADSCs extracted by this method [6,20].…”

“…Additionally, they revealed that the sheets contained collagen I and III, bronectin, and vitronectin, and exhibited stable adhesion to the surface of cartilage matrix. We investigated the optimal condition of Vc-induced ADSC sheets formation and indicated osteogenic potential of the sheets 20 . However, the properties of Vc-induced cell sheets have many unclear parts such as strength, species, incubation period, and quantity of Vc.…”

Expression patterns of mesenchymal stem cell-specific proteins in adipose tissue-derived cells: possible immunosuppressing agent in partial allograft for restoring the urinary bladder in rabbits

“…Rabbits have a large amount of adipose tissue and a used in basic science for different purposes for many years. Regarding tissue engineering approaches, rabbit animal models have been used for urethral, cartilage, and vocal fold reconstruction (Barretto et al 2014). This paper focuses on the characterization of expression profiles of clusters of differentiation (CDs) in rabbit ADSCs used as immunosuppressive agent in rabbits undergoing urinary bladder allografts.…”