Neonatal Satellite Cells Form Small Myotubes In Vitro

Abstract

Although palatal muscle reconstruction in patients with cleft palate takes place during early childhood, normal speech development is often not achieved. We hypothesized that the intrinsic properties of head satellite cells (SCs) and the young age of these patients contribute to the poor muscle regeneration after surgery. First, we studied the fiber type distribution and the expression of SC markers in ex vivo muscle tissue from head (branchiomeric) and limb (somite-derived) muscles from neonatal (2-wk-old) and young (9-wk-old) rats. Next, we cultured SCs isolated from these muscles for 5, 7, and 9 d, and investigated the in vitro expression of SC markers, as well as changes in proliferation, early differentiation, and fusion index (myotube formation) in these cells. In our ex vivo samples, we found that virtually all myofibers in both the masseter (Mass) and the levator veli palatini (LVP) muscles contained fast myosin heavy chain (MyHC), and a small percentage of digastric (Dig) and extensor digitorum longus myofibers also contained slow MyHC. This was independent of age. More SCs were found in muscles from neonatal rats as compared with young rats [17.6 (3.8%) v. 2.3 (1.6%); P < 0.0001]. In vitro, young branchiomeric head muscle (BrHM) SCs proliferated longer and differentiated later than limb muscle SCs. No differences were found between SC cultures from the different BrHMs. SC cultures from neonatal muscles showed a much higher proliferation index than those from young animals at 5 d (0.8 v. 0.2; P < 0.001). In contrast, the fusion index in neonate SCs was about twice as low as that in SCs from young muscles at 9 d [27.6 (1.4) v. 62.8 (10.2), P < 0.0001]. In conclusion, SCs from BrHM differ from limb muscles especially in their delayed differentiation. SCs from neonatal muscles form myotubes less efficiently than those from young muscles. These age-dependent differences in stem cell properties urge careful consideration for future clinical applications in patients with cleft palate.

Wnt7a induces satellite cell expansion, myofiber hyperplasia and hypertrophy in rat craniofacial muscle

“…At ten-week age, SCs proportion in LVP muscle were much more than 1%, yet remaining well below 1% in MAS muscle and TA muscle. Heterogeneities in SCs proportion among muscles of different embryonic origins or different ages were also reported by other studies 11 , 26 , 27 . It has been proved by Carvajal Monroy that rat LVP muscle and MAS muscle contained more SCs than limb muscle 27 .…”

“…Heterogeneities in SCs proportion among muscles of different embryonic origins or different ages were also reported by other studies 11 , 26 , 27 . It has been proved by Carvajal Monroy that rat LVP muscle and MAS muscle contained more SCs than limb muscle 27 . Keefe reported that the SCs density varied nearly five folds among different muscles 26 .…”

Tissue engineering strategies combining molecular targets against inflammation and fibrosis, and umbilical cord blood stem cells to improve hampered muscle and skin regeneration following cleft repair

“…They express the paired box transcription factor 7 (Pax7) and are located between the sarcolemma and basal lamina surrounding a single muscle fiber. After the injury, SCs become activated and migrate to the site of injury, proliferate, and differentiate into myoD‐positive myoblasts that fuse to form new multinucleated myosin heavy chain (MyHC)‐positive myofibers or repair damaged myofibers (Figure ) . A small portion of these SCs stay quiescent to allow future regeneration cycles.…”

“…SCs from different branchiomeric muscles, however, behave in a similar way. The lower regenerative capacity of branchiomeric muscles compared to limb muscle may impair muscle healing after cleft surgery . The reduced blood supply and aberrant muscle orientation in a cleft lip may further impair the regeneration process .…”

“…During embryonic development, the limb and trunk muscles are derived from the somites whereas lip muscles are derived from the CPM in the branchial arches . Head muscles also contain less SCs than limb muscles and injuries regenerate slower . In addition, Pax7 and myogenic regulatory factor 4 levels in limb muscles are lower than in head muscles.…”