Influence of neoadjuvant chemoradiotherapy on the anal sphincter: ultrastructural damage may be critical.

BACKGROUND: Neoadjuvant chemoradiotherapy (nCRT) in addition to the curative surgery has been the first of treatment for local advanced rectal cancer because of its benefits in local recurrence and sphincter-saving. However, its side effects on anorectal function have been recognized. The histopathological changes on internal anal sphincter (IAS) have been reported, but ultrastructure changes of external anal sphincter (EAS) are unknown. The aim of this study is to detect the alterations on the gross morphology of IAS and ultrastructure of EAS after nCRT. METHODS: We collected 34 anal canal specimens of patients undergoing abdominoperineal resection (APR) prospectively. The length and thickness of IAS were measured with vernier caliper. The EAS was dissected for observation with transmission electron microscope (TEM). RESULTS: Ten patients received nCRT (nCRT group) before surgery and 24 underwent APR directly (control group). The length and thickness of IAS in nCRT group were 22.68 ± 3.56 and 5.39 ± 0.74 mm, respectively. These parameters were 21.28 ± 3.62 and 5.35 ± 1.12 mm in control group, respectively. There were no significant differences in the length and thickness of IAS between the two groups (P>0.05). In nCRT group, the sarcomere and myofibril were arranged disorderly and parts of them that were filled with collagenous fiber, triads, and mitochondria were destroyed severely and the glycogenosome also distributed disorderly. Such alterations of EAS did not occur in control group. CONCLUSIONS: The nCRT cannot change the gross morphology of IAS, while it induces serious damages to the ultrastructures of EAS which may adversely affect the anorectal function.

Nerves supplying the internal anal sphincter: an immunohistochemical study using donated elderly cadavers.

PURPOSE: Nerves serving the internal anal sphincter (NIAS) have been described as the lower rectal branches of the pelvic autonomic nerve plexus. However, their topographical anatomy and fiber components have remained unclear. METHODS: Using histological sections from ten elderly donated cadavers, we investigated the topographical anatomy and composite fibers of the NIAS using immunohistochemistry for S100 protein, neuronal nitric oxide synthase (nNOS), vasoactive intestinal polypeptide (VIP) and tyrosine hydroxylase (TH). RESULTS: At the 2-3 o'clock position in the lower rectum, the NIAS originated from nerves at the posterolateral corner of the prostate in males or in the lower paracolpium in females. The nerves ran inferiorly along the internal aspect of the levator ani muscle, and joined branches of the myenteric plexus at a level slightly above the epithelial junction. The NIAS contained both nNOS-positive parasympathetic nerve fibers and TH-positive sympathetic fibers, but VIP-positive fibers were few in number. CONCLUSIONS: The origin of the NIAS at the posterolateral corner of the prostate as well as in the lower paracolpium might be sacrificed or damaged during radical prostatectomy or tension-free vaginal tape insertion. Low anterior resection of rectal cancer will most likely render damage to the NIAS because of its intersphincteric course. Although the nerve composition of the NIAS is characterized by a higher proportion of sympathetic nerve fibers than the myenteric plexus in the large intestine, their role is unclear. However, evaluation of sphincteric function after surgery would appear to be difficult because of the complex control mechanism independent of nerve supply.

Morphology of the epithelium of the lower rectum and the anal canal in the adult human.

The anal canal is an important body part clinically. However, there is no agreement about the epithelium of the anal canal, the anal transitional zone (ATZ) epithelium in particular. The aim of this study is to clarify the structure of the epithelium of the human lower rectum and anal canal. Intact rectum and anus obtained from patients who underwent surgery for rectal carcinoma were examined by light and scanning electron microscopy (LM and SEM). By LM, three types of epithelium were observed in the anal canal: simple columnar epithelium, stratified squamous epithelium, and stratified columnar epithelium. The lower rectum was composed of simple columnar epithelium. SEM findings showed stratified squamous epithelium that consisted of squamous cells with microridges, changing to simple columnar epithelium consisting of columnar cells with short microvilli at the anorectal line. LM and SEM observations in a one-to-one ratio revealed that the area of stratified columnar epithelium based on LM corresponded to the anal crypt and sinus. In conclusion, the epithelium of the human anal canal was fundamentally composed of simple columnar epithelium and stratified squamous epithelium. We found no evidence of the ATZ.

Identification of epithelial label-retaining cells at the transition between the anal canal and the rectum in mice.

In certain regions of the body, transition zones exist where stratified squamous epithelia directly abut against other types of epithelia. Certain transition zones are especially prone to tumorigenesis an example being the anorectal junction, although the reason for this is not known. One possibility is that the abrupt transition of the simple columnar epithelium of the colon to the stratified squamous epithelium of the proximal portion of the anal canal may contain a unique stem cell niche. We investigated whether the anorectal region contained cells with stem cell properties relative to the adjacent epithelium. We utilized a tetracycline-regulatable histone H2B-GFP transgenic mice model, previously used to identify hair follicle stem cells, to fluorescently label slow-cycling anal epithelial cells (e.g., prospective stem cells) in combination with a panel of putative stem cell markers. We identified a population of long-term GFP label-retaining cells concentrated at the junction between the anal canal and the rectum. These cells are BrdU-retaining cells and expressed the stem cell marker CD34. Moreover, tracking the fate of the anal label-retaining cells in vivo revealed that the slow-cycling cells only gave rise to progeny of the anal epithelium. In conclusion, we identified a unique population of cells at the anorectal junction which can be separated from the other basal anal epithelial cells based upon the expression of the stem cell marker CD34 and integrin alpha6, and thus represent a putative anal stem cell population.

Effects of ovarian failure on submucosal collagen and blood vessels of the anal canal in postmenopausal women.

BACKGROUND: Estrogen and progesterone receptors are expressed in the anal canal. Fecal control deteriorates after menopause. This phenomenon is related to decreased circulating levels of estrogen and progesterone due to ovarian failure at menopause. AIM OF WORK: To study the effects of estrogen and progesterone on inflammatory cells, submucosal collagen fibers, and vascular plexus of the anal canal of postmenopausal women. SUBJECTS AND METHODS: Experiments were performed on samples of anorectal tissue obtained from 40 women, 19 menstruating (group I), and 21 postmenopausal women (group II). Investigations included immunohistochemistry of estrogen and progesterone receptors and CD34. RESULTS: In negative estrogen receptors (ER) and progesterone receptors (PR), inflammatory cells, submucosal blood vessels, collagen type I were nonsignificantly changed in postmenopausal women relative to menstruating women (P > 0.05) whereas, in positive ER and PR, inflammatory cells and collagen I were significantly increased and submucosal blood vessels were significantly decreased in postmenopausal women relative to menstruating women (P < 0.05). CONCLUSION: Estrogen and progesterone, in menstruating women, produce beneficial effects by decreasing incidence of inflammation and increasing anal canal submucosal blood vessels number and collagen types I, thus both hormones have a positive effect on anal compliance and pressure.

Fine structure of the anal tonsilliar epithelium in the laboratory shrew (Suncus murinus).

The epithelium of the anal tonsil of the laboratory shrew was studied by scanning and transmission electron microscopy, with particular attention focused on the structure of the epithelium lining the anal tonsillar crypt. The tonsillar crypt surface is lined by two kinds of epithelia: squamous epithelium, which is located mainly at the neck of the crypt and includes keratohyalin granules in the superficial layer, and reticular epithelium, which is invaded by many immigrating cells and has several micropores immigrating cells to pass through. In addition, basal granulated cells are present in the basal layer. These results suggest that the reticular epithelium of the anal tonsil belongs to the well-developed gut-associated lymphoid tissue (GALT) in the alimentary canal. It represents a specialized and important compartment in immunological function, similarly to the palatine tonsils of other mammals, and has as yet unknown roles in digestion.

Light and electron microscopic study of perineal gland in the grasscutter (Thryonomys swinderianus, Temminck 1827).

The thin and ultrathin structure of the perineal gland of the grasscutter was studied using routine histological, histochemical and electron microscopic techniques. The perineal glands were identified in the zona cutanea of the anal canal of males more than 4 months old. These glands are organized on a lobular model made up of sebaceous cells and isolated by connective parietal cells. The paler in colour the sebaceous cells, the more lipid vacuoles they contain. PAS-negative and Sudan III-positive secretions were demonstrated in these cells. Electron microscopy showed that the intracellular organelles were well-developed, which is a mark of high metabolic activity and storage by the glandular cells. Cystic vesicles were found in the lobules. The size and number of these cysts were variable. The position of the cyst cells and the variability of their size suggest that they are formed after lysis of some holocrine secretory cells. These observations suggest a progressive maturation of the gland towards its secretory site. No specific secretory duct for the purpose of draining secretions was observed, however, there were possible discharge pathways, although with no real walls, as is found for the biliary canaliculi.

Fibroepithelial polyps of the anus: a histologic, immunohistochemical, and ultrastructural study, including comparison with the normal anal subepithelial layer.

Fibroepithelial polyps of the anus (FEPA) are relatively common lesions that have attracted little attention. A series of 40 FEPA, together with sections from normal anal mucosa, were studied histologically, histochemically, immunohistochemically, and ultrastructurally. The polyps consisted of myxoid or collagenous stroma covered by squamous epithelium. Stromal cells with two or more nuclei were found in 30 polyps (80%). In five cases (12%) these cells showed atypical nuclear features. All the polyps harboring atypical cells were of large size. Mast cells were frequently present and sometimes intimately related to the stromal cells. Stromal cells stained positive for vimentin and negative for actin. Desmin was expressed in 30% of the cases. Electron microscopic examination confirmed the fibroblastic and myofibroblastic nature of the stromal cells. Examination of the subepithelial connective tissue from the normal anal mucosa showed bizarre multinucleated cells and mast cell infiltration. It is concluded that FEPA are benign lesions characterized by the presence of mononucleated and multinucleated, sometimes atypical stromal cells showing fibroblastic and myofibroblastic differentiation. The morphologic similarity between FEPA and normal anal mucosa supports the hypothesis that FEPA may represent a reactive hyperplasia of the subepithelial connective tissue of the anal mucosa. Mast cells, by means of their fibrogenic, fibrolytic, and angiogenic activities may play an important role in the pathogenesis of FEPA.

[The epithelial junctional zones of the anal canal and cervix uterus: the ultrastructure of tumors of these zones]. / Epitel'ial'nye stykovye zony anal'nogo kanala i sheiki matki: ul'trastruktura nekotorykh opukholei étikh zon.

Relationships between squamous and columnar epithelia in the anal canal and cervix uteri of postnatal period and fetus were studied. The transitional stratified epithelial lining, which is called junctional epithelium, is interposed between the mentioned epithelia. The junctional epithelium has variable numbers of layers of epidermoid cells, which differ from cells of atypical squamous epithelium by some ultrastructural features of the cytoskeleton and cell surface and by a low content of glycogen. The hypothesis on the physiological significance of this epithelium is proposed. Ultrastructural features of the cytoskeleton and cell surface suggest that anal basaloid carcinomas and some cervical squamous carcinomas may develop from so-called junctional epithelium.

Ultrastructural changes in internal anal sphincter in neurogenic faecal incontinence.

The ultrastructural features of the internal anal sphincter (IAS) muscle were studied in biopsies from five patients with neurogenic anorectal incontinence and six control subjects undergoing anorectal excision for cancer, or for inflammatory bowel disease. In the patients with idiopathic neurogenic anorectal incontinence the internal anal sphincter showed loss of smooth muscle cells, disruption of the normal relationships of the remaining cells, stretching of elastic tissue, and increased collagen fibril content. These ultrastructural changes in the morphology of the internal anal sphincter, although probably not the primary cause of faecal incontinence, have functional relevance in the clinical syndrome, as shown by the reduction in resting anal canal pressure found in some patients with this syndrome.

Density of myosin filaments in the rat anococcygeus muscle, at rest and in contraction. II.

Rat anococcygeus muscles were fixed at rest or in contraction by conventional methods and prepared for electron microscopy. Myosin filaments were counted on cross sections and their density expressed per unit cytoplasmic area. In contracted muscles, the mean density increased from 86 to 168 filaments per micron 2 (1.95 times), while the density of intermediate (10 nm) filaments increased by 1.25 times. Cell cross sections from the same muscles were measured. Contraction produced a shrinkage which explains the apparent increased density of the 10 nm filaments; however an excess of 61 myosin filaments per micron 2 cannot be explained in this way. These findings provide the structural basis which quantitatively explains the birefringence changes observed in living contracted muscle (Godfraind-De Becker & Gillis, 1988). Our optical and electron optical results provide evidence for a reversible formation of myosin filaments during contraction of the rat anococcygeus muscle.

Comparative study of the periurethral and perianal parts of the human levator ani muscle.

The present study has used histochemical and electron microscopic techniques to provide a quantitative comparison of the striated muscle fibre populations in the periurethral and perianal regions of the human levator ani muscle. Both regions consist predominantly of type I (slow twitch) fibres together with a small number of type II (fast twitch) fibres. However, marked differences have been demonstrated between the two regions with regard to proportions of the two fibre types, diameters of the constituent fibres and the distribution of muscle spindles. These observations are discussed in relation to the differing functional activity of the periurethral and perianal regions of the levator ani muscle.

Histogenesis of cloacogenic carcinoma. Fine structure of anal transitional epithelium and cloacogenic carcinoma.

The fine structural morphology of the anal transitional zone is described and is compared to the ultrastructure of urothelium and anal squamous epithelium. Electron microscopic observations in eight cases of cloacogenic carcinoma are also presented. Anal transitional epithelium is not highly specialized and incorporates features of both urothelium and squamous epithelium; slight urothelial differentiation is considered vestigial. Cloacogenic carcinoma is a specific morphologic entity, which can be distinguished from urothelial and squamous carcinoma. The "transitional" variant is composed of cells similar to the intermediate zone cells of anal transitional epithelium, an observation confirming the concept that the transitional form of cloacogenic carcinoma is derived from the anal transitional zone. The "basaloid" and pleomorphic variants appear to represent less differentiated forms of cloacogenic carcinoma.

Sphincter denervation in anorectal incontinence and rectal prolapse.

Biopsies of the external anal sphincter, puborectalis, and levator ani muscles have been examined in 24 women and one man with long-standing anorectal incontinence, 18 of whom also had rectal prolapse, and in two men with rectal prolapse alone. In 16 of the women anorectal incontinence was of unknown cause, but in eight there was a history of difficult labour. Similar biopsies were examined in six control subjects. In all the incontinent patients there was histological evidence of denervation, which was most prominent in the external anal sphincter muscle biopsies, and least prominent in the levator ani muscles. Myopathic features, which were thought to be secondary, were present in the more abnormal biopsies. There were severe histological abnormalities in small nerves supplying the external anal sphincter muscle in the three cases in which material was available for study. We suggest that idiopathic anorectal incontinence may be the result of denervation of the muscles of the anorectal sling, and of the anal sphincter mechanism. This could result from entrapment or stretch injury of the pudendal or perineal nerves occurring as a consequence of rectal descent induced during repeated defaecation straining, or from injuries to these nerves associated with childbirth.

Microplicae: characteristic ridge-like folds of the plasmalemma.

Scanning electron microscopy reveals that the free surfaces of stratified squamous epithelial cells lining the alimentary tract, cornea, and conjunctiva exhibit characteristic ridge-like folds of plasmalemma. These microplicae are approximately 0.1-0.2 micronm in width, of variable height (0.2-0.8 micronm) and length, may followstraight or winding paths, often branch, and exhibit a wide variety of patterns over the surfaces of cells. Transmission electron microscopy reveals that microplicae often have a fine (100-150 A) electron-dense zone subjacent to their plasmalemma and an intracellular matrix characterized by a disorderly arrary of fine filaments (40-60 A in diameter). Microplicae appear to arise from plasmalemmal fold which once provided for intercellular interdigitation and desmosome abhesion between adjacent cells. Ruthenium red staining demonstrates that microplicae and interplical grooves are covered with a polyanionic glycocalyx. Although free surface microplicae may merely represent the renmants of intercellular interdigitations or a modified expression of microvillous-like extensions, it is also possible that they serve another specific function. In this regard it is speculated that microplical and interplical grooves may function to hold a layer of lubricating and cushioning mucin designed to protect the underlying plasmalemma from abrasive abuse.