Case 4 -

Acute Ischemic Injury with Fibrin Thrombi in Microvasculature Charles R. Lassman Geffen School of Medicine at UCLA

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Clinical History A 31 year old woman, presented to the emergency room with severe cramps and bloody diarrhea. She had been in good health until two days prior to presentation when she suffered from acute onset of watery diarrhea and abdominal cramping. After two days, the diarrhea turned bloody and she sought medical attention. Examination in emergency room was notable for a tender abdomen, which was not felt to require surgical intervention. The patient was afebrile. Routine laboratory work was unremarkable. The patient was placed on levafloxacin and sent home; stool cultures were not performed. The patient continued to pass moderate amounts of blood in her stool and returned to the ER the following day with worsening cramps. A CT suggested "inflammation and ischemia" of the hepatic flexure and right transverse colon; the patient was admitted for further evaluation and observation. On admission stool was sent for culture and analysis and the patient was continued on levafloxacin with a presumptive diagnosis of acute infectious colitis. Stool cultures were negative for pathogenic organisms. Severe cramping and bloody diarrhea continued unabated for two more days. Five days after the onset of symptoms and 4 days after antibiotic treatment was started, colonoscopy was performed and biopsies were taken. Colonoscopy demonstrated normal appearing mucosa from the cecum to the proximal transverse colon. The proximal transverse colon appeared inflamed and markedly edematous with areas of extensive superficial ulceration. The distal transverse colon appeared less edematous, with less severe ulceration. The rectum was minimally involved. Biopsies were obtained from the rectum as well as the transverse colon. Case 4 - Slide 1 Click to view with ImageScope Click to view with a Web-Based Viewer Case 4 - Figure 1 - Colonic tissue demonstrating hemorrhage with superficial mucosal necrosis, mild acute inflammation, preservation of deep crypts, reepithelialization of the surface and occasional deep crypt apoptosis. The submucosa demonstrates edema hemorrhage and a mild mixed inflammatory infiltrate (20x, H &E). Case 4 - Figure 2 - Colonic tissue demonstrating hemorrhage with superficial mucosal necrosis, mild acute inflammation, preservation of deep crypts, reepithelialization of the surface and occasional deep crypt apoptosis. The submucosa demonstrates edema hemorrhage and a mild mixed inflammatory infiltrate (20x, H &E). Case 4 - Figure 3 - Colonic tissue demonstrating hemorrhage with superficial mucosal necrosis, mild acute inflammation, preservation of deep crypts, reepithelialization of the surface and occasional deep crypt apoptosis. The submucosa demonstrates edema hemorrhage and a mild mixed inflammatory infiltrate (20x, H &E). Case 4 - Figure 4 - A small fragment of fibrinopurulent exudate with foreign (fecal) material (40x, H&E) Case 4 - Figure 5 - Deep mucosal capillaries with fibrin thrombi (40x, H &E). Case 4 - Figure 6 - Colonic tissue demonstrating hemorrhage with superficial mucosal necrosis, mild acute inflammation, preservation of deep crypts and reepithelialization of the surface. (40x, H &E). Endoscopic Impression: Acute inflammatory process of the transverse colon, probable severe acute infectious colitis, ischemic colitis can not be excluded, less likely, though still possible is inflammatory bowel disease, specifically Crohn's. Diagnosis: Acute Ischemic injury with fibrin thrombi in microvasculature, rule out E. coli O157:H7. Subsequent Clinical History: Following histologic exam of biopsy specimens, further clinical history was obtained. Two days prior to the onset of symptoms, the patient went to her parent's home to house-sit while they were away. She arrived late in the evening, was quite hungry and found some form of cooked ground meat in the refrigerator. She stated that it tasted bad but thought it was lamb (she did not like lamb) and was so hungry that she continued to eat it. She later found out that it was hamburger. Discussion: Most fecal E. coli. do not cause disease in humans; a subset of organisms however elaborate toxins which are responsible for a wide variety of colonic manifestations. These organisms have been referred to as enterotoxigenic E. coli and those which are associated with bloody diarrhea are referred to as enterohemorrhagic E. coli. (EHEC). Pathogenesis is due to toxins which are closely related to those elaborated by Shigella dysinteriae, so called shiga-like toxins (stx). Numerous shiga-like toxins have been identified and characterized. Enterohemorrhagic E. coli which elaborate shiga-like toxins have been referred to as shiga toxin producing E. coli (STEC). Not all STEC and not all stx are pathogenic; most EHEC associated human disease is caused by organisms elaborating stx 1 and the stx 2. There are numerous subtypes of stx 2 with differing propensity to cause diarrhea, hemorrhagic diarrhea and thrombotic microangiopathy (TTP and HUS). More than 100 serotypes of EHEC have been identified, the most common is O157:H7. The O refers to a cell surface antigen and H refers to a flagellar antigen. Bloody diarrhea occurs more commonly with this serotype than with others. In 1982 an outbreak of bloody diarrhea primarily amongst adults was linked to consumption of hamburgers at different outlets of the same fast food chain. Cultures for usual Campylobacter, Salmonella, Shigella and Yersinia were negative. E. coli O157:H7 was isolated from stool samples of many of the patients (Riley) and was thought to be the cause for the outbreak of hemorrhagic colitis. Around the same time Karmali et al. reported the isolation of a filtratable substance from stool samples of children with HUS in Toronto. The substance was toxic to cultured Vero cells (a cell line derived from epithelial cells of the African green monkey). These toxins were referred to as verotoxins and the E. coli which produced them were referred to as verotoxin producing E. coli (VTEC). Various serotypes of E. coli were isolated from the stool samples of these children with HUS and E. coli O157:H7 was seen in two of eight children. Verotoxin producing E. coli was thought to be a causative agent in some cases of idiopathic HUS. In 1983 O'Brien reported that the verotoxin activity of the E. coli O157:H7 could be neutralized by antibodies to Shiga toxin. Vero toxin producing E. coli are now recognized to be enterohemorrhagic E. coli (EHEC) and the verotoxins are recognized to be the shiga-like toxins. Since then sporadic cases and outbreaks of E. coli O157:H7 have been reported. The most recent being the spinach borne illness in the fall of 2006 which resulted in approximately 200 infections, 3 deaths and 31 cases of kidney failure and the more recent Taco Bell outbreak with another approximately 200 infections (and counting). The main sources of contamination have been undercooked ground beef, unpasteurized milk, unpasteurized apple juice, raw vegetables, water, swimming in contaminated ponds, and human to human contact. Infection is more common in warm weather months, and the symptoms are more severe in children and in the elderly. The bacteria can be isolated from as many as 80% of cattle in the United States. They are not pathogenic to the bovine host as pathogenesis requires attachment of the bacterium to epithelial cells through a highly specific receptor which the cattle lack. It has been suggested that forage fed cattle have a lower rate of infection than grain fed cattle and that forage feeding for 1 week prior to slaughter can markedly reduce the chance of contaminated meat. Proposed Pathogenesis: The bacteria attach to enterocytes, colonize and elaborate a toxin. Receptors for toxin on epithelial and endothelial cells may be upregulated by the immune response to bacterial colonization. The toxin is internalized by epithelial cells, and more importantly endothelial cells. Once internalized, the toxin irreversibly blocks protein synthesis resulting in cell death. Endothelial cell death results in thrombosis of small vessels, primarily capillaries and arterioles. If the toxin becomes widely distributed HUS or TTP may occur. Clinical Presentation: The incubation period is 1-8 days, generally 3-5. The first manifestation in many patients is watery diarrhea with severe cramping. In more severe cases there is progression to blood streaked diarrhea or hemorrhagic colitis. Patients are generally afebrile but suffer from severe cramps and vomiting. Between 5 and 10% of children with hemorrhagic colitis associated with E. coli O157:H7 will then develop HUS or TTP. A smaller percentage of adults develop HUS or TTP. HUS may occasionally develop in the context of mild colonic disease. There is currently no way to predict which patients will develop systemic disease. Thrombocytopenia and the presence of schistocytes on a peripheral smear indicate the development of HUS. In patients for whom bloody diarrhea has resolved for 2-3 days, and for whom evidence of thrombotic microangiopathy has not developed, HUS will most likely not develop. Antibiotics have not been demonstrated to be of benefit for either hemorrhagic colitis or for preventing the onset of HUS. They may in fact be contraindicated as there is some evidence that they increase the risk of development of HUS. Histology and Diagnosis: There are a few studies reporting the histology of hemorrhagic colitis associated with E. coli O157:H7. Colonoscopy is generally not performed unless disease is severe and or there is confounding clinical information to suggest IBD, ischemia or some other form of colitis. Resection specimens due to necrosis or perforation are reflective of severe disease. The histologic features are nonspecific and may be suggestive of ischemic and or infectious colitis. Findings are patchy, however the right colon is more frequently involved. When severe the entire colon may be affected. Mild injury is limited to the mucosa but when severe there may be full thickness necrosis resulting in perforation. Griffin et al. described a combination of infectious and ischemic histologic features in 11 patients (19 biopsies and one resection). All had edema and hemorrhage of the lamina propria. Fourteen specimens (9 patients) had histology indicative of ischemia as characterized by superficial necrosis with preservation of the deeper crypts. In only 10 specimens (7 patients) were fibrin/platelet thrombi identified in mucosal capillaries. Ten specimens (6 patients) demonstrated deep crypt apoptosis. Focal active colitis with neutrophilic infiltration of crypts and the lamina propria and with crypt abscesses, typical of infectious, acute self limited colitis was seen in 10 specimens (5 patients). Poorly formed pseudomembranes were seen in 5 specimens (4 patients). Material form 5 patients showed only ischemic changes, four showed a mixed pattern and one showed an infectious without ischemic changes. In 2003 Murray and Patterson reported the pathologic changes in colonic specimens from eight pediatric patients with hemorrhagic colitis and HUS from a 1993 outbreak in the northwest USA. The study is weighted towards those with quite severe colonic disease. There was one patient with biopsy material only, there were 4 subtotal colectomies, two left hemicolectomies and two autopsies. They confirmed many of the previously described findings and reported more significant left sided disease than has been reported for adults with hemorrhagic colitis secondary to E. coli O157:H7. Routine cultures are insufficient for diagnosis. Culture must be performed on MacConkey sorbitol agar and routine media, the sorbitol negative colonies are then tested with serum for O157 and H7 antigens. Cultures are less likely to be positive late in the course of illness and in patients treated with antibiotics. Su et al. have reported the use of a monoclonal antibody which can be used on paraffin embedded tissue sections. In a retrospective study, 10 sections from 2 patients with culture positive E. coli O157:H7 hemorrhagic colitis were positive with strong staining of the exudates overlying the area of ulceration. Interestingly 3 of 11 cases diagnosed as ischemic colitis were also positive. Staining was negative or equivocal for IBD (8 cases) and pseudomembranous colitis (3). This stain may therefore be helpful in identifying cases of E. coli O157:H7 associated hemorrhagic colitis. Selected References: Calderwood SB, Auclair F, Donohue-Rolfe A, Keusch GT, Mekalanos JJ. Nucleotide sequence of the Shiga-like toxin genes of Escherichia coli. Proc Natl Acad Sci U S A. 1987 Jul;84(13):4364-8. Cleary TG. The role of Shiga-toxin-producing Escherichia coli in hemorrhagic colitis and hemolytic uremic syndrome. Semin Pediatr Infect Dis. 2004 Oct;15(4):260-5. Griffin PM, Olmstead LC, Petras RE. 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