Macro-microscopic Anatomy Of Anal Conjoined Longitudinal Muscle And Anal Glands

Objectives Anatomy of the anal conjoined longitudinal muscle (CLM) and anal glands (AGs) has significant impact on the development of anal fistula and on the defecation. The aim of this project was to identify histological features of the CLM and AGs of infant and adult cadavers by using histological and sheet plastination techniques and laser confocal microscopy, in order to provide the morphology data for the pathogenesis and surgical approach of anal fistula.Methods (1) A total of 16 adult and 10 infant cadavers were used in this study. The anal regions were harvested from the cadavers and prepared as sets of series of transverse (6 infant and 8 adult cadavers) and coronal (4 infant and 5 adult cadavers) sections that were then stained with H.E. or van-Gieson (VG) or Verhoeff's Van Gieson (EVG) elastic fiber staining. (2) The remaining 3 adult cadavers were prepared as transverse and sagittal plastinated sections with a thickness of 300μm. Some plastinated sections were stained with H.E. or EVG elastin staining and examined under a stereomicroscope microscope or a confocal miscroscope to observe the structures and distributions of CLM and AGs. (3) Some sections were selected to photo. Then the photos were merged into a complete transverse or longitudinal section using Photoshop 7.0 to identify histological features of anal conjoined longitudinal muscle and anal glands.Results (1)On the transverse or longitudinal sections, five layers of structures were identifiable in the wall around the anal canal. From the interior to exterior, they were the mucous, submucous, internal anal sphincter, CLM and external anal sphincter layers. (2) The CLM appeared as a longitudinal layer of the muscular tissue, sandwiched in between the internal and external anal sphincters. It could be further divided into two sublayers: an inner layer consisted of predominantly inner smooth muscle fibers that originated from the longitudinal muscular layer of the rectum and an outer layer containing smaller numbers of skeletal muscle fibers that originated from the levator ani. The inner lays of the CLM gradually became a number of small elastic fibers that penetrated through the internal anal sphincter layer to the submucosal layer, and gathered as elastic fiber bundles, anchoring onto the perianal dermis. The external layer of the CLM continued inferiorly as many relatively larger connective tissue fiber bundles that penetrated the superficial part and deep part of the external anal sphincter, and then faned out and disappeared in the ischiorectal fossa. However, those bundles that penetrated the subcutaneous part of the external anal sphincter as elastic fibers and anchored onto the perianal skin. (3) Most AGs were tubular glands with some irregular branches. Adult AGs were either acinus or tubuloacinar glands but infant AGs only had a tubular appearance. At its opening into the anal canal, the AG was lined with stratified squamous epithelium. As the ducts coursed through submucous layer and internal anal sphincter, the thickness of the lining progressively decreased to 2 to 3 cellular layers, and at the terminal end became a simple epithelium cell layer. The basal cells in anal gland mucosa were similar to those in stratified squamous epithelium. The cells on the surface were cuboidal or columnar. Some mucus-secreting or goblet cells interspersed within the glandular epithelial lining and many glands also showed the presence of intraluminal secretions. Most AGs were distributed in the submucosal layer. In the infant, however, some AGs penetrated the submucosal and internal anal sphincter layers and extended into the CLM, whereas in the adult, occasionally AGs extended into the internal anal sphincter layer. AGs were fairly evenly distributed in the anal circumference at the pectinate line. (4) Compared to the unstained plastinated slices, the tissue layers in the wall of the anal canal in the thin plastinated sections stained with EVG elastin staining, were easily distinguished. (5) Under the confocal microscope, the direct continuation of muscle fibers and collagen fibers was clearly identifiable.Conclusions (1) The detailed macro-/micro-scopic configuration of the CLM and its relationship with the internal and external anal sphincters has been revealed in this study, which may be helpful for surgical planning in the area and better understanding of the pathogenesis of the formation of fistula. (2) The evidence from this study indicates that the AGs and CLM had no direct structural connection in the adult, suggesting that the formation of fistula in ano is more likely resulted from the spread of infection along the fibrous bundles rather than the gland itself. (3) Studying the sectional anatomy of anal canal by using histological and plastination techniques and laser confocal microscopy was a new approach to reveal the histological features of CLM and AGs. (4) The structures and distribution of anal conjoined longitudinal muscle and anal glands in the thin plastinated sections stained with EVG elastin staining were more clearly than the unstained thin plastinated sections. (5) The plastination process results in collagen being endogenously autofluorescent at the 488-nm excitation, but elastic fibers and lamellae was not.