Three-dimentional Distribution Of α-SMA～+, MHC-Ⅰ～+, MHC-Ⅱ～+, Pax7～+ Or Pax3～+ Cells In The Human Pharynx And Esophagus

Objective: To identify characteristics of muscular structures and to reveal their three-dimensional distribution in the human pharynx and esophagus in order to better understand the mechanism underlying the functional chest pain.Methods:①A total of 10 human fetuses were assigned for this project.The pharynx and esophagus from 4 out of 10 fetuses were sectioned at 5 identical levels which were at upper and lower pharyngeal levels, upper, middle and lower esophageal levels. The sections were stained with H.E staining and immunohistochemical staining with anti-α-Smooth Muscle Actin, anti-slow myosin heavy chain, anti-fast myosin heavy chain,anti-Pax7 and anti-Pax3 antibodies. The specimens from the remaining six fetuses were used for Western blotting analysis with anti-α-Smooth Muscle Actin, anti-slow myosin heavy chain, anti-fast myosin heavy chain,anti-Pax7 and anti-Pax3 antibodies.Results: The general configuration of the pharynx and esophagus muscle fibers were examined in the serial sections at each level.We found that the muscular layer of the pharynx expressed Pax7 and slow and fast MHC, but not a-SMA, suggesting that it only contains skeletal muscle fibres. At the upper and middle levels of the esophagus, the muscular layer of the esophagus contained both smooth and skeletal muscle fibers. Interestingly, Pax7, MHC-I and MHC-II positive profiles were always co-localized and were dominant in the external longitudinal muscular layer and gradually disappeared at the inferior end of the esophagus. In contrast,α-SMA expression was detected in the most inner muscular layer at the upper end of the esophagus and then gradually increased downward. The muscle fibers expressingα-SMA were mainly circularly orientated. Pax3 was not detected in the muscular layer of the pharynx and esophagus. To verify identity of Pax3 expression, we used the gastrocnemius muscle as a positive control specimen. We found that Pax7 and Pax3 were always co-localized in the gastrocnemius, Pax7 expression was stronger than Pax3. Western blotting analysis have shown that Pax7, MHC-I and MHC-II were always co-localized in the muscular layer and gradually disappeared at the inferior end of the esophagus. In contrast,α-SMA expression was detected in the muscular layer at the upper end of the esophagus and then gradually increased downward.Conclusion:①The results suggest that differentiation of pharyngeal and esophageal skeletal muscles proceeds from the superior to the inferior ends of the esophagus in the human fetus.②Pax7, MHC-I and MHC-II were always co-localized in the upper and middle esophagus, supporting the assumption that pax7 positive cells were progenitors of pharyngeal esophageal skeletal muscle fibres.③The study also precisely identified the transition zone between skeletal muscle and smooth muscle of the esophagus. Localization of such zone is much more complicated than we generally believe. It depends on three-dimentional distribution ofα-SMA, MHC-I,MHC-II ,Pax7 and Pax3 in the muscular layer of the pharynx and esophagus. In summary, the above conclusion will provide new information for us to evaluate the development of pharynx and esophagus, particularly in human. Such information will be helpful for clinical diagnose of function chest pain, which may provide theoretical basis for location for balloon distentions.