Evaluation of Levator Veli Palatini Innervation in Congenitally Clefted and Non-Clefted Goat Palates

Congenital cleft palates are one of the most commonly occurring birth defects in the world, with recent studies reporting rates as high as 7 per 10,000 live births. Despite surgical correction of cleft palates, velopharyngeal incompetence (VPI) persists in 15% of patients, such that additional surgical procedures are required. The authors hypothesized that VPI may be due to inherent levator veli palatini (LVP) muscle dysfunction and, specifically, denervation of this muscle.

A total of six palates from 14-month-old female Spanish goats were used for the study: three control non-clefted goat palates and three congenital chemically-induced clefted goat palates. LVP muscles were analyzed morphometrically by light microscopy and evaluated for myosin ATPase activity to determine slow and fast muscle-fiber composition. Myosin heavy chain content was analyzed by Western blot analysis. Neural cell adhesion molecule (NCAM) and myogenin immunohistochemistry were performed to evaluate velar myogenesis and determine innervation status.

Based on myosin ATPase activity, non-clefted palate (NP) muscles contained only Type 1, slow-oxidative muscle fibers, whereas cleft palate (CP) muscles demonstrated both Type 1 (29.3 ± 10.6%) and Type 2 fast muscle fibers (70.7 ± 10.6%). Cross-sectional areas of muscle fibers from CP goats (1987 ± 739 μm²) were significantly larger than those from the NP group (1656 ± 518 μm²)(p < 0.0003). This difference was explained by the larger percentage of Type 2 muscle fibers in the CP compared with the NP goats. Western blot analysis of myosin heavy chain content confirmed the presence of Type 1 and Type 2 muscle fibers in the CP and the absence of Type 2 fibers in the NP goats. One potential explanation for this difference is that skeletal muscle denervation results in muscle conversion to a physiologically fast phenotype (Type 2). Myogenin immunohistochemistry demonstrated a greater than nine-fold increase in muscle fibers expressing myogenin in the CP (47.8%) compared with the NP goats (5.2%). Myogenin is upregulated in denervated muscle fibers. Initial results also showed a three-fold increase in fibers expressing NCAM on the sarcolemmal surfaces of the CP group(6.9%) compared with the NP group (2.3%). Similar to myogenin, NCAM is upregulated on the sarcolemmal surface of denervated muscle fibers.

Clear differences exist between non-clefted and clefted goat palates. An increase in Type 2 fast glycolytic fibers and myogenin and NCAM expression in CP muscle are all consistent with the increase in denervated muscle fibers in CP muscle, which can impact palatal dysfunction and VPI after cleft-palate repair.