Specificity of pork masseter muscle metabolic and contractile type identified by histology
Authors: Vénien Annie , Danon Jeanne , Loison Olivier , Astruc Thierry 
Published: December 2016

Skeletal muscle is usually composed of four types of muscle cells (I, IIA, IIX, IIB) varying on their speed of contraction (slow: type I; fast: type II) and their metabolism pathway of glycogen (oxidative: type I and IIA; glycolytic: Type IIX and IIB). Depending on the anatomical position and muscle function, the proportion of these different muscle fiber types is variable. The different fibers types are identified longstanding by histoenzymology. ATPase activity in type I fibers is labile at alkali pH and resistant at acidic pH. In contrast, the ATPase activity of type II fibers is alkali resistant and acid labile (For review, see Pette & Staron, 1990, 2000, Schiaffino & Reggiani, 2011). The molecular basis of this typology resides in the polymorphism of myosin heavy chains (MyHC). The use of monoclonal antibodies against MyHC isoforms allows to identify precisely the type I IIA IIX and IIB fibers and hybrid fibers expressing simultaneously different isoforms of myosin (Schiaffino & Reggiani, 2011).

Our goal was to characterise the fiber type of porcine masseter muscle.

Materiel and methodes

Ten 6 months old pigs (105‐115 Kg) were slaughtered in a commercial abattoir and masseter muscles (jaw) were extracted from the head at 30 min postmortem. Muscle sample (1X1X1.5 cm) were frozen in cooled isopentane (−160 °C). Serial cross‐sections (10 µm thick, cryostat Microm, HM 560) were collected on glass. Fiber types were identified by histoenzymology both by revealing ATPase activity after acidic incubation, and SDH activity that reflect oxidative metabolism. Fiber types were also identified by immunohistofluorescence using three monoclonal antibodies specific to MyHC isoform (BAD5 specific for type I, S58 H2 all except type IIa, and BF35 all types except type IIb and IIx) (Schiaffino & Reggiani, 2011). Histological sections were observed on a photonic microscope (Olympus BX 61) coupled to a high resolution digital camera (Olympus DP 71) and the Cell F software. The percentage of each fiber type was calculated according to Meunier et al. (2010) using the image analysis Visilog 6.7 Professional Software.

Results and discussion

ATPase histoenzymology (Fig.1) revealed only type I (30.6%) and IIA (69.4%) oxidative fibers according to Ström & Holm, 1997. Immunohistochemistry (Fig.2) revealed that 17% of the masseter fibers are hybrid fibers containing two MyHC isoforms IIa and IIx (Fig.3). These hybrid fibers are generally a transitional step to move from a pure type to another (Schiaffino & Reggiani, 2011). In this study, no pure type IIX is highlighted, suggesting that the transition is not completed or that the IIA‐IIX hybrids are the final stage of transition in this muscle.

For a given muscle, fiber type evolves mainly with physical activity and age of the animals. In our case, pigs were reared in a building without noticeable change in their physical activity. The pigs were slaughtered at the age of 6 months, which corresponds to a period of entry into sexual maturity. It is possible that the fibers are in transition classes, however, no study has highlighted the presence of IIX fibers in pork masseter, suggesting that the hybrid could be permanent. A large proportion of IIA‐IIX fibers (74 %) were found in rat masseter but several pure glycolytic fibers were detected (Pette & Staron, 1990) and unique HCIIm isoforms were identified in primate and carnivore masseter muscles (Pette & Staron, 1990, 2000). All these data highlight the special feature of masseter compared to other skeletal muscles.

Conclusion

The use of monoclonal antibodies revealed the presence of a high proportion of hybrid fibers IIA‐IIX in pork masseter and did not allow the detection of any pure type IIX. These results suggest a specificity of porcine masseter muscle in which the hybrid type IIA‐IIX would not be a transitory state.