Mucosal Schwann cell hamartoma: a benign and little-known entity.

Dear editor, 50 years-old female with personal history of mutation of the gene BRCA1 and previous prophylactic double anexectomy consulted for rectal bleeding without pain since two weeks. A blood test was performed, with hemoglobin levels of 13.1g/dl and without iron deficiency. In the anal inspection there were neither external hemorrhoids nor anal fistulas, so a colonoscopy was requested. In the colonoscopy, all the colon mucosa was normal but, in the rectal retroflexion, apart from internal engorged hemorrhoids, surrounding the 50% of the anal opening an erythematous and indurated mucosa was found (figure 1). Biopsies were taken. The pathology report informed of proliferation of spindle-shaped cells exclusively in the lamina propria with eosinophilic cytoplasm and unclear cell borders (figure 2). Not nuclear atypia or mitotic activity were observed. On immunohistochemistry, S-100 protein was strongly positive (figure 3) and CD34, SMA, EMA and c-kit were negative. These results are concordant with the diagnosis of Schwann cells in the context of a mucosal Schwann cell hamartoma (MSCH). Given that these lesions seem to not have malignant potential, the patient was discharged without control colonoscopies. The episodes of rectorrhagia were attributed to the presence of internal hemorrhoids. Discussion: MSCH are benign and intramucosal tumors with a mesenchymal origin. They are most commonly located in the distal colon, but they were also found in the gallbladder, the esophagogastric union and in the antrum. They are observed most frequently in middle aged women (around 60 years-old) and they are generally asymptomatic. They are presented as polyps between 1 and 6mm, but in other cases they appeared as small whitish nodules, protruding lesions with normal superficial mucosa or even they were found in random biopsies of the colon. The MSCH are a rare entity with an unknown prevalence. Less than 100 cases are described in the literature. It is essential the differentiation between this entity and the Schwanomas or the gastrointestinal stromal tumors (GIST). Schwanomas are rare in the colon, they are well circumscribed (in contrast with the MSCH) and they are not limited to the lamina propria. GIST are more frequently located in the stomach and they are positive for c-kit. MSCH are not associated with hereditary syndromes such as neurofibromatosis and, in contrast with Schwanomas or GIST, they do not require surveillance because they are benign.

Identification of the microRNA alterations in extracellular vesicles derived from human haemorrhoids.

NEW FINDINGS: What is the central question of this study? What are the morphological features and microRNA (miRNA) expression features of extracellular vesicles (EVs) from haemorrhoids (Hae-EVs) and normal tissues? What are the potential functions of the differentially expressed (DE) miRNAs in Hae-EVs? What is the main finding and its importance? We present, for the first time, the morphological features and miRNA profile of human Hae-EVs. Four hundred and forty-seven significant DE-miRNAs were identified. Gene ontology and pathway analysis of the DE-miRNAs indicated diverse roles of the Hae-EVs through different pathways. Our findings provide EV-based pathological features and the underlying mechanism of haemorrhoids. ABSTRACT: Extracellular vesicles (EVs) play important roles in many pathophysiologies as cell-to-cell communication vehicles. However, the features and potential functions of the EVs in haemorrhoids remain unclear. Therefore, we performed microRNA (miRNA) microarray analysis in EVs derived from haemorrhoid tissue to identify the profile of miRNAs in these EVs and predict their potential functions. We obtained typical EVs from both haemorrhoid and control tissues. Microarray analysis identified 447 miRNAs with significant differential expresssion (DE): 245 upregulated and 202 downregulated. The top three upregulated miRNAs in haemorrhoid EVs (Hae-EVs), namely miR-6741-3p, miR-6834-3p and miR-4254, were detected by RT-qPCR in both Hae-EVs and haemorrhoid tissues. Interestingly, we found a different expression pattern in the haemorrhoid tissues from that in Hae-EVs. The potential target genes of these DE-miRNAs were predicted by the miRWalk and miRDB databases. Gene ontology (GO) analysis of the target genes showed that the DE-miRNAs contributed mainly to protein kinase activity, transcriptional activity and ubiquitin-protein function. KEGG search found that the DE-miRNAs might regulate the MAPK and Ras signalling pathways. These findings revealed, for the first time, the miRNA profiles in Hae-EVs and provided potential targets and pathways involved in the pathological process.

miR-412-5p targets Xpo1 to regulate angiogenesis in hemorrhoid tissue.

Hemorrhoid is a common and recurrent proctological disease, which is often accompanied by angiogenesis and edema. MicroRNAs in the DLK1-DIO3 imprinted clusters are involved in the development and pathogenesis of mammalian hemorrhoids. Results of the present study indicated multiple, differential expression of DLK1-DIO3 imprinted cluster microRNA between hemorrhoid and normal tissues, where miR-412-5p expression in hemorrhoid tissue was significantly decreased. Fluorescein reporter assays showed that miR-412-5p silenced Xpo1 mRNA expression by targeting its 3'-UTR. Overexpression of miR-412-5p in human umbilical vein endothelial cells (HUVECs) indicated that proliferation, migration and formation of vascular structures in HUVECs were inhibited in vitro. In addition, overexpression of miR-412-5p significantly inhibited Xpo1 expression and promoted upregulation of the p53 protein and its retention in the nucleus. Simultaneously, expression of p66SHC and p16 proteins was activated. In summary, downregulation of endogenous miR-412-5p expression in hemorrhoid vascular endothelial cells leads to high expression of the target gene Xpo1 and translocation of the p53 protein out of the nucleus, rendering it unable to activate p66SHC and p16. This ultimately weakens regulation of the vascular endothelial cell cycle, thereby accelerating the division of hemorrhoid vascular endothelial cells, leading to angiogenesis.

Endothelin-1 and its receptors on haemorrhoidal tissue: a potential site for therapeutic intervention.

BACKGROUND AND PURPOSE: Haemorrhoids is a common anorectal condition affecting millions worldwide. We have studied the effect of endothelin-1 (ET-1) and the role of endothelin ETA and ETB receptors in haemorrhoid tissue. EXPERIMENTAL APPROACH: Protein expression of ET-1, ETA and ETB receptors were compared between haemorrhoids and normal rectal submucosa using Western blot analysis, with the localization of proteins determined by autoradiography and immunohistochemistry. Effects of ET-1 and sarafotoxin 6a on human colonic and rectal arteries and veins was assessed by wire myography and the involvement of receptor subtypes established by selective antagonists. KEY RESULTS: Dense binding of [125 I]-ET-1 to haemorrhoidal sections was reduced by selective receptor antagonists. A higher density of ETB than ETA receptors was found in haemorrhoidal, than in control rectal tissue and confirmed by Western blot analysis. ETA and ETB receptors were localized to smooth muscle of haemorrhoidal arteries and veins, with ETB receptors on the endothelium. Human colonic and rectal arteries and veins were similarly sensitive to ET-1 and affected by the ETA selective antagonist, but sarafotoxin S6a-induced contractions were more pronounced in veins and antagonized by a selective ETB receptor antagonist. CONCLUSIONS AND IMPLICATIONS: ETA and ETB receptors are present in human haemorrhoids with ETB receptors predominating. ETA receptors are activated by ET-1 to mediate a contraction in arteries and veins, but the latter are selectively activated by sarafotoxin S6a - a response that involves ETB receptors at low concentrations. Selective ETB agonists may have therapeutic potential to reduce congestion of the haemorrhoidal venous sinusoids.

Hemorrhoids and matrix metalloproteinases: A multicenter study on the predictive role of biomarkers.

INTRODUCTION: An association between hemorrhoidal disease and matrix metalloproteinases (MMPs) has been described previously. MMPs regulate extracellular structural proteins and tissue remodeling. Neutrophil gelatinase-associated lipocalin (NGAL) is involved in the regulation of MMP activity. The aim of this work was to study the relationship between tissue immunoreactive levels of MMPs and NGAL and different stages of hemorrhoids. METHODS: In a multicenter, open-label, prospective study, the population under investigation consisted of 2 groups: group I (with symptomatic hemorrhoids; Goligher grade I-IV) and group II (healthy volunteers). RESULTS: We enrolled 97 patients with hemorrhoids: 21 with grade I hemorrhoids, 37 with grade II, 14 with grade III, and 25 with grade IV. Finally, 90 healthy volunteers (53 males and 37 females; age range, 19-70 years; median, 56) were enrolled in group II. Enzyme-linked immunosorbent assay and Western blot analysis revealed greater levels of immunoreactive MMPs and NGAL in all patients with hemorrhoids. We recorded significantly greater levels of MMP-1 and MMP-3 in grade I and II patients compared with control, and greater levels of MMP-3, MMP-7, MMP-8, and MMP-9 in grade III compared with grade II. MMP-9 and NGAL were particularly increased in patients with grade IV especially in case of thrombosed hemorrhoids. CONCLUSION: These results provide potentially important insights into the understanding of the natural history of hemorrhoids. MMPs and NGAL play a role in development of disease and may represent molecular markers for the complications such as hemorrhoidal thrombosis.

Myofibrotic malformation vessels: unique angiodysplasia toward the progression of hemorrhoidal disease.

BACKGROUND: The etiology and pathogenesis of hemorrhoids is unclear, although hemorrhoids are a worldwide disease in men and women, with peak prevalence at 45-65 years of age. Hemorrhoidal cushions as the anal venous plexi are normal anatomical structures from infancy. This study attempts to reveal the angiodysplasia and other pathological changes in association with different degrees of symptomatic hemorrhoids. MATERIALS AND METHODS: A total of 281 patients with internal hemorrhoids from degree I to IV underwent hemorrhoidectomy. The vascular changes were analyzed by microscopic assessment and software analysis, with Masson's trichrome, CD34, and smooth muscle actin. RESULTS: The hemorrhoidal tissues exhibited abnormal vessels in the mucosae and submucosae that we termed them as myofibrotic malformation vessels (MMVs). MMVs are not ascribed to arteries or veins because they exhibit enlarged and tortuous lumens with smooth muscle dysplasia and fibrotic deposition in the walls without overlying mucosal ulceration. The muscularis mucosae also showed smooth muscle dysplasia and fibrosis, even if it were interrupted by the intruding MMVs. The statistical data indicated that the severity of all the changes correlate positively with the progression of hemorrhoids (P<0.001). Hemorrhoidal patients are prone for reoccurrence even with prolapsing hemorrhoid when compared with the conventional hemorrhoidectomy. Multiple logistic regression analysis showed that MMVs in mucosal propria, mean thickness of mucosal muscularis layer, and fibrotic changes in MMV were independent risk factors for MMVs in hemorrhoidal disease. CONCLUSION: MMVs and muscularis mucosae dysplasia reciprocally contribute to hemorrhoidal exacerbation. The novel findings of this study propose that the characteristic features of MMVs and muscularis mucosae dysplasia of the anorectal tube ultimately cause symptomatic hemorrhoids, which could affect the clinical management of hemorrhoidal disease through the use of surgery to target the malformed vessels.

Effect of fermented milk containing Lactobacillus casei strain Shirota on constipation-related symptoms and haemorrhoids in women during puerperium.

Constipation and haemorrhoids are common complaints after childbirth. The objective of this pilot study was to evaluate impact of fermented milk containing Lactobacillus casei strain Shirota (LcS) on stool consistency and frequency, constipation-related symptoms and quality of life, and incidence of haemorrhoids in women during puerperium. Forty women who had natural childbirth were randomised to group consuming either one bottle/day of fermented milk containing at least 6.5×109 cfu of LcS, or placebo, for 6 weeks after childbirth. Subjects filled in a diary on their bowel habits including number of bowel movement, stool consistency and incidence of haemorrhoids, and answered questionnaires on constipation-related symptoms (PAC-SYM) and quality of life (PAC-QOL) during the study period. The probiotic group showed the better scores on overall PAC-SYM (P=0.013), PAC-SYM subscales of abdominal symptoms (P=0.043) and rectal symptoms (P=0.031), and PAC-QOL satisfaction subscale (P=0.037) in comparison with the placebo group. In the probiotic group, two to four subjects experienced haemorrhoids during the first 3 weeks of treatment. The number decreased in week 4 and no one had haemorrhoids on most days in week 5-6. In the placebo group, on average four subjects had haemorrhoids from the beginning, and no obvious change was observed until week 6. No statistically significant effect was observed on stool consistency and frequency. The study products did not cause any adverse event in the subjects. Results of this study indicate that continuous consumption of fermented milk containing LcS might alleviate constipation-related symptoms, provide satisfactory bowel habit and result in earlier recovery from haemorrhoids in women during puerperium. Nonetheless, there are several limitations in interpretation of the results attributed to the study design, including lack of baseline data. Further study is required in order to confirm the efficacy.

Expression pattern, ethanol-metabolizing activities, and cellular localization of alcohol and aldehyde dehydrogenases in human large bowel: association of the functional polymorphisms of ADH and ALDH genes with hemorrhoids and colorectal cancer.

Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are principal enzymes responsible for metabolism of ethanol. Functional polymorphisms of ADH1B, ADH1C, and ALDH2 genes occur among racial populations. The goal of this study was to systematically determine the functional expressions and cellular localization of ADHs and ALDHs in human rectal mucosa, the lesions of adenocarcinoma and hemorrhoid, and the genetic association of allelic variations of ADH and ALDH with large bowel disorders. Twenty-one surgical specimens of rectal adenocarcinoma and the adjacent normal mucosa, including 16 paired tissues of rectal tumor, normal mucosae of rectum and sigmoid colon from the same individuals, and 18 surgical mixed hemorrhoid specimens and leukocyte DNA samples from 103 colorectal cancer patients, 67 hemorrhoid patients, and 545 control subjects recruited in previous study, were investigated. The isozyme/allozyme expression patterns of ADH and ALDH were identified by isoelectric focusing and the activities were assayed spectrophotometrically. The protein contents of ADH/ALDH isozymes were determined by immunoblotting using the corresponding purified class-specific antibodies; the cellular activity and protein localizations were detected by immunohistochemistry and histochemistry, respectively. Genotypes of ADH1B, ADH1C, and ALDH2 were determined by polymerase chain reaction-restriction fragment length polymorphisms. At 33mM ethanol, pH 7.5, the activity of ADH1C*1/1 phenotypes exhibited 87% higher than that of the ADH1C*1/*2 phenotypes in normal rectal mucosa. The activity of ALDH2-active phenotypes of rectal mucosa was 33% greater than ALDH2-inactive phenotypes at 200µM acetaldehyde. The protein contents in normal rectal mucosa were in the following order: ADH1>ALDH2>ADH3≈ALDH1A1, whereas those of ADH2, ADH4, and ALDH3A1 were fairly low. Both activity and content of ADH1 were significantly decreased in rectal tumors, whereas the ALDH activity remained unchanged. The ADH activity was also significantly reduced in hemorrhoids. ADH4 and ALDH3A1 were uniquely expressed in the squamous epithelium of anus at anorectal junctions. The allele frequencies of ADH1C*1 and ALDH2*2 were significantly higher in colorectal cancer and that of ALDH2*2 also significantly greater in hemorrhoids. In conclusion, ADH and ALDH isozymes are differentially expressed in mucosal cells of rectum and anus. The results suggest that acetaldehyde, an immediate metabolite of ethanol, may play an etiological role in pathogenesis of large bowel diseases.

Expression of estrogen and progesterone receptors in the anal canal of women according to age and menopause.

PURPOSE: Fecal incontinence is highly prevalent, especially in menopausal women. The aim of this study was to analyze the expression of estrogen and progesterone receptors in the anal canal of women in relation to menopausal status and age. METHODS: Samples of hemorrhoidal tissue were obtained from 34 women undergoing hemorrhoidectomy. The patients were divided into 2 groups: group 1 consisted of women with a menstrual cycle (n = 17) and group 2 consisted of postmenopausal women (n = 17). Immunostaining of hormone receptors was performed using specific antibodies (DAKO, Copenhagen, Denmark) in cells from the internal anal sphincter, the vascular epithelium, and the squamous epithelium. The percentage of positivity of receptors and the association between age and receptor positivity were compared between the 2 groups. RESULTS: Estrogen receptors were found in the internal anal sphincter in 23.5% in group 1 vs 11.8% in group 2 (P = .656). Progesterone receptors were found in 41.2% in group 1 vs 11.8% of group 2 (P = .118). Squamous epithelium showed estrogen receptors in 52.9% in group 1 vs 64.7% of group 2 (P = .388) and progesterone receptors in 17.6% and 0% in groups 1 and 2, respectively (P = .227). Vascular endothelium showed no receptors. Receptor positivity was not associated with age. CONCLUSION: No significant differences were found in the detection of estrogen and progesterone receptors in structures of the anal canal in women in relation to menopausal status and age.

Haemorrhoids - a collagen disease?

OBJECTIVE: The cause of haemorrhoidal disease is unknown, epidemiological data and histopathological findings support the hypothesis that reduced connective tissue stability is associated with the incidence of haemorrhoids. Therefore the aim of this study was to analyse the quantity and quality of collagen formation in the corpus cavernosum recti in patients with III°/IV° haemorrhoids in comparison with persons without haemorrhoids. METHOD: Haemorrhoidectomy specimens of 31 patients with III°/IV° haemorrhoids were examined. The specimens of 20 persons who died a natural death and who had no haemorrhoidal disease served as the controls. The amount of collagen was estimated photometrically by calculating the collagen/protein ratio. The collagen I/III ratio served as parameter for the quality of collagen formation and was calculated using cross polarization spectroscopy. RESULTS: Patients with haemorrhoids had a significantly reduced collagen/protein ratio (42.2 ± 16.2µg/mg vs 72.5±31.0µg/mg; P= 0.02) and a significantly reduced collagen I/III ratio (2.0±0.1 vs 4.6±0.3; P<0.001) compared with persons without haemorrhoidal disease. There was no correlation with patients' age or gender. CONCLUSIONS: There is a fundamental disorder of collagen metabolism in patients with haemorrhoidal disease. It remains unclear whether this is due to exogenous or endogenous influences.

[Pathologic change of elastic fibers with difference of microvessel density and expression of angiogenesis-related proteins in internal hemorrhoid tissues].

OBJECTIVE: To investigate the pathological variations in internal hemorrhoid and evaluate the expression of nitric- oxide synthase(NOS),vascular endothelial growth factor(VEGF),matrix metalloproteinase- 2(MMP2) and MMP9. METHODS: Normal anal cushion and internal hemorrhoids tissue samples were obtained from 24 patients with iii degree hemorrhoids after procedure for prolapse and hemorrhoids(PPH) procedure. The expression of NOS, VEGF, MMP2, MMP9 and CD34 were detected by immunohistochemical staining; the microvessel density (MVD) was counted by anti- CD34 antibody; the elastic fibers were detected by orcein staining. RESULTS: There were statistically significant differences in the expression of MVD, VEGF, MMP9 between internal hemorrhoid tissue and normal anal cushions(P< 0.05). iNOS was significantly increased in hemorrhoid tissue, but no significant difference between normal anal cushions and hemorrhoid tissue. Morphological abnormalities such as breaking, distortion, mortality, hyaline degeneration were found in elastic fibers of internal hemorrhoid tissue, but not in normal anal cushions. CONCLUSION: Angiogenesis is evident in hemorrhoid tissue, suggesting the possible mechanism in the pathogenesis of hemorrhoids. The direct degeneration effect of MMP9 on supporting structure elastic fibers in anal cushion is another important mechanism. The high expression of iNOS suggests the inflammatory factors involve in the pathogenesis of hemorrhoids, and NO may be involve in pathological effect on hemorrhoids.

Endoglin (CD105) expression in the development of haemorrhoids.

BACKGROUND: Conventional pathogenesis of haemorrhoid emphasized the anchoring connective tissue system, whereas the vascular changes were ignored. The aim of this study was to clarify vascular changes of haemorrhoid disease. MATERIALS AND METHODS: Forty-six samples of grade III and grade IV haemorrhoid tissue were selected for an in vitro study. We assessed the expressions in endoglin (CD105), an accessory protein in transforming growth factor-beta receptor complex, in CD34 and in vascular endothelial growth factor by using an immunohistochemical method. Microvascular density was calculated to correlate the expression of endoglin. RESULTS: Microvascular density was higher in haemorrhoid tissue than in normal anal and lower rectal tissues. CD34 was demonstrated in whole vessels in the haemorrhoids. However, endoglin, a proliferative marker of neovascularization, was present in only 25 of 46 (54%) haemorrhoidal vessels, and its immunoactivity was prominent in venules larger than 100 micro m. Thrombosis formation and stromal vascular endothelial growth factor was significantly associated with the presence of endoglin immunoactivity. CONCLUSION: The results of this study suggest that neovascularization is one important phenomenon of haemorrhoid disease, along with conventional venous dilatation and arteriovenous communication. In addition, thrombosis and stromal vascular endothelial growth factor might be important factors in promoting vascular proliferation.

CD34+ stromal cells and hyalinized vascular changes in the anal fibroepithelial polyps.

AIMS: To elucidate the pathogenesis of the anal fibroepithelial polyp, we examined surgically resected lesions histopathologically. METHODS AND RESULTS: Twenty-seven surgically resected anal fibroepithelial polyps were investigated histologically with an additional immunohistochemical examination using anti-CD34. For a control study, the surgical specimens of the anal canal showing non-polypoid lesions, obtained from haemorrhoidectomy (18 specimens) and rectectomy (five specimens) due to rectal cancer without anal canal involvement, were also analysed. We demonstrated characteristic spindle or stellate cells immunohistochemically positive for CD34 in the anal fibroepithelial polyps (24/27, 89%). The number of CD34+ cells was statistically related to the size of anal fibroepithelial polyps, although CD34+ stromal cells were recognized in the non-polypoid anal submucosa and haemorrhoids. We also found hyalinized vascular changes in the base of six anal fibroepithelial polyps examined. These features were not detected in the non-polypoid anal canal. CONCLUSIONS: An increase in CD34+ stromal cells may play a role in the enlargement of anal fibroepithelial polyps. CD34+ stromal cells are suggested to be distinctive mesenchymal cells with a capability for tissue repair and overgrowth. The vascular impairment could be secondary change associated with localized tissue damage by abnormal traction.

Pagetoid dyskeratosis is a frequent incidental finding in hemorrhoidal disease.

BACKGROUND: Pagetoid dyskeratosis is considered a selective keratinocytic response in which a small part of the normal population of keratinocytes is induced to proliferate. Pagetoid dyskeratosis has been found incidentally in the squamous epithelium of the skin in various locations and in the ectocervix in uterine prolapse. In cases in which these pale cells are conspicuous, there is a hazard of overdiagnosis. It has been suggested that friction is the most probable inductor of the lesion. To the best of our knowledge, pagetoid cells have not been reported in surgically resected hemorrhoids. OBJECTIVE AND DESIGN: We here describe the location of pagetoid dyskeratosis in the squamous epithelium of hemorrhoids and the incidence of this lesion in a group of 100 unselected patients surgically treated for hemorrhoidal disease. In addition to the conventional histologic method, special staining procedures and an immunohistochemical study of cytokeratins were performed in selected cases. RESULTS: Pagetoid dyskeratosis was found in 68 cases (68%) and was a prominent finding in 22 cases (22%). The cells of pagetoid dyskeratosis were strongly positive for high-molecular weight cytokeratin. These cells showed an immunohistochemical profile that was different from that of the surrounding squamous cells and indicative of premature keratinization. CONCLUSIONS: In hemorrhoidal disease, the cushions are susceptible to trauma as a result of prolapse. In this setting, friction may be the stimulus for the appearance of pagetoid dyskeratotic cells. These cells must be distinguished from the artifactual clear cells of the squamous epithelium, glycogen-rich cells, and koilocytes. The lesion must also be distinguished from extramammary Paget disease, pagetoid spread of carcinoma cells, pagetoid Bowen disease, and pagetoid melanoma. Pathologists should be familiar with the histologic features of pagetoid dyskeratosis in hemorrhoidectomy specimens to avoid misdiagnosis. Routine histologic study is usually adequate for recognizing this lesion.

Colonic substance P levels are increased in ulcerative colitis and decreased in chronic severe constipation.

Substance P content was determined by radioimmunoassay in colonic mucosa from 24 patients with chronic severe constipation, 16 with active ulcerative colitis, and 28 normal controls. In patients with chronic severe constipation, the mean concentration of substance P (19.9 +/- 8.2 pg/mg) was significantly lower than in normal subjects (71 +/- 18 pg/mg). In patients with ulcerative colitis, colonic substance P concentration in inflamed mucosa (170 +/- 46 pg/mg) was significantly higher than its levels in normal subjects. Substance P may therefore have a role in the pathogenesis of clinical conditions associated with diarrhea and constipation.