Investigation of molecular markers in the diagnosis of refractory coeliac disease in a large patient cohort.

AIMS: Some patients with coeliac disease, despite strict adherence to a gluten-free diet, continue to have significant symptoms and/or a severe small intestinal histological lesion. The term "refractory coeliac disease" (rCD) is used to describe this condition. The purpose of this study was to investigate the value of tissue molecular markers reported to help in the diagnosis of rCD. METHODS: Details on 61 patients with suspected rCD were collected. The clinical and laboratory findings in these patients were carefully evaluated, in part to determine whether patients were adhering to a strict gluten-free diet. The co-expression of CD3 and CD8 on intraepithelial lymphocytes was investigated by monoclonal antibody staining of small intestinal biopsy tissue; a finding of less than 50% CD3+ cells co-expressing CD8 was defined as an aberrant phenotype. T cell receptor gene rearrangement was assessed when a sufficient tissue sample was available. RESULTS: A diagnosis of rCD was made in 38 patients based on clinical, laboratory and histological data. An aberrant intraepithelial lymphocyte population was found in 20 of these patients and in this group a clonal T cell population was found in five of seven patients tested. In the remaining 18 patients, the CD3/CD8 ratio was normal and two of seven tested had a clonal T cell population. After detailed monitoring, a diagnosis of rCD was excluded in the remaining 23 patients. CONCLUSIONS: This study supports the use of phenotypic and T cell clonality investigations in identifying patients with true rCD.

Epidemiological findings and medical, legal, and public health challenges of an investigation of severe soft tissue infections and deaths among injecting drug users -- Ireland, 2000.

In May 2000, public health authorities in Dublin, Ireland, identified a cluster of unexplained severe illness among injecting drug users (IDUs). Similar clusters were also reported in Scotland and England. Concurrent investigations were undertaken to identify the aetiology and source of the illnesses. In Dublin, 22 IDUs were identified with injection-site inflammation resulting in hospitalization or death; eight (36%) died. Common clinical findings among patients with severe systemic symptoms included leukaemoid reaction and cardiogenic shock. Seventeen (77%) patients reported injecting heroin intramuscularly in the 2 weeks before illness. Of 11 patients with adequate specimens available for testing, two (18%) were positive by 16S rDNA PCR for Clostridium novyi. Clinical and laboratory findings suggested that histotoxic Clostridia caused a subset of infections in these related clusters. Empiric treatment for infections among IDUs was optimized for anaerobic organisms, and outreach led to increased enrolment in methadone treatment in Dublin. Many unique legal, medical, and public health challenges were encountered during the investigation of this outbreak.

Dermatomyositis associated with angiotropic lymphoma.

We describe a 64-year-old man with dermatomyositis sine myositis who presented with unusual cutaneous plaques and nodules. Initial investigations failed to reveal evidence of malignancy. Eighteen months later he became systemically unwell and repeat biopsies from the nodules confirmed angiotropic T-cell lymphoma. Cutaneous lymphoma is rarely reported with dermatomyositis and in the previously reported cases, mycosis fungoides was the variant seen. This is the first report of angiotropic T-cell lymphoma associated with dermatomyositis.

Whole-mount NADPH-diaphorase histochemistry is a reliable technique for the intraoperative evaluation of extent of aganglionosis.

Multiple seromuscular biopsies at three levels (narrow segment, transitional zone, and dilated segment) were taken and investigated intraoperatively to determine the extent of aganglionosis. Using the whole-mount preparation technique, circular muscle fibers were separated from the specimens. After a short prefixation, the muscle fibers were stained by the NADPH-diaphorase technique and were examined within 20-25 min. A fine and dense neuronal meshwork was observed between circular muscle fibers in the normal and ganglionic part of the bowel. In contrast, there was a complete lack of NADPH-diaphorase-positive fibers in the circular muscle of aganglionic colon. In the transitional zone, NADPH-diaphorase-positive fibers were markedly reduced compared to the ganglionic region. The density of these fibers increased and attained normal levels in the proximal bowel above the transition zone. These results suggest that whole-mount NADPH-diaphorase histochemistry is a three-dimensional technique suitable for the intraoperative evaluation of extend of aganglionosis. The technique is sufficiently rapid to be used in conjunction with routine frozen sections to assist in the diagnosis and in selecting the optimal level of resection at the time of pull-through operation.

NADPH diaphorase-containing nerve fibers and neurons in the myenteric plexus are resistant to postmortem changes: studies in Hirschsprung's disease and normal autopsy material.

OBJECTIVE: Nitric oxide is considered to be the most important messenger of inhibitory nonadrenergic, noncholinergic nerves in the enteric nervous system. Histochemical studies have shown that nitric oxide synthase is identical to reduced nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase. Histochemical staining with NADPH diaphorase has been widely used to study nitrergic innervation of the gastrointestinal tract, but fresh tissue is considered a prerequisite for satisfactory results. The purposes of this study were to evaluate whether whole-mount specimens of human bowel obtained after death are suitable for histochemical staining with NADPH diaphorase and to compare the staining properties with those of specimens of resected bowel from patients with Hirschsprung's disease METHODS: Whole-mount preparations of the myenteric plexus were examined using NADPH diaphorase histochemical staining of bowel specimens obtained at autopsy from 18 pediatric subjects (31 specimens). Fresh tissue was also obtained from the colon of four patients with Hirschsprung's disease. The staining properties of postmortem specimens were assessed in relation to the postmortem time before fixation (<12 hours, 13-24 hours, or 25-48 hours) and were compared with those of specimens of ganglionic bowel from patients with Hirschsprung's disease. Specimens of aganglionic bowel were also stained and examined. RESULTS: Strong NADPH diaphorase staining was achieved in 26 of the 31 postmortem bowel specimens, including all specimens from patients who underwent autopsy 25 to 48 hours after death. Staining properties were similar to those obtained in ganglionic bowel specimens from patients with Hirschsprung's disease. In aganglionic bowel the normal myenteric plexus meshwork was absent and was replaced by weakly staining nerve fibers. CONCLUSION: Histochemical staining with NADPH diaphorase is a robust technique suitable for use with whole-mount preparations to demonstrate nitrergic innervation in motility disorders such as Hirshsprung's disease. The technique may be used with both fresh tissue and specimens obtained up to 48 hours after death.

Morphometric aspects of the submucous plexus in whole-mount preparations of normal human distal colon.

BACKGROUND/PURPOSE: The diagnosis of intestinal neuronal dysplasia (IND) has traditionally been based on the finding of hyperplasia of the submucous plexus and increased acetylcholinesterase activity in parasympathetic nerve fibers in the lamina propria. However, recently it has been suggested that proposed diagnostic criteria relating to nerve cell density may overlap with age-related changes and that the finding of giant ganglia (ganglia containing more than seven ganglion cells) is the most relevant diagnostic parameter of IND. Ganglion cell counting is usually performed on conventional histological sections, whereas the topology of whole ganglia has been poorly studied. The aim of this study was to define the number of ganglion cells per ganglion and the ganglion cell density (the number of ganglion cells per surface area) in submucous whole-mount preparations of normal human colon. METHODS: Specimens from distal colon were obtained during postmortem examination from 14 patients who died of nongastrointestinal disease. The submucous layer was prepared as a whole mount and stained for NADPH diaphorase (a nitrergic neurotransmitter marker) and cuprolinic blue (a general neuronal marker). Ganglion cell density was estimated by counting at least 10 mm2. The number of ganglion cells per ganglion was counted in at least 20 ganglia per case. RESULTS: Ganglion cell density (NADPH diaphorase) fell markedly during the first 5 to 6 years of life (r = -0.60, P < .05). Most ganglion cells formed ganglia of 3 to 64 cuprolinic blue staining cells. The mean number of ganglion cells per ganglion did not vary with age. CONCLUSIONS: The density of NADPH diaphorase-positive ganglion cells in the submucous plexus of human distal colon decreases markedly with age. However, the number of ganglion cells per ganglion remains constant. These findings indicate that the age of the patient has crucial importance for the histolopathologic evaluation of enteric nervous system disorders.

Parvovirus B19 infection in AIDS patients.

Bone marrow of 61 HIV-1-infected patients and 23 control patients was examined to determine the incidence of B19 infection and its clinical impact in HIV-1-infected persons. Of the 61 HIV-infected patients studied, ages ranged from 22-47 years with a mean of 33.2 years. There was a man:woman ratio of 3.8:1. With regard to staging of HIV disease at the time of bone marrow sampling, 52 patients were CDC group 4, 5 patients were CDC group 3, and 4 patients were CDC group 2. Control patients, were not known to be HIV-1-infected, and had one of the following conditions: lymphoma, leukaemia, thrombocytopenia, thrombocytosis, anaemia, multiple myeloma, raised serum IgM. Thirteen of 61 HIV-infected patients and 0 of 23 control patients were positive for B19 DNA in bone marrow (two-tailed P value = 0.016). Within the HIV-infected group, the average haemoglobin among persons testing B19 DNA positive (n = 13) was 11.1 g/dl, compared with 11.5 g/dl among persons testing B19 DNA negative (n = 48). In conclusion, B19 persistence may be common and frequently subclinical in AIDS patients.

Expression of epidermal growth factor receptor and proliferating cell nuclear antigen predicts response of esophageal squamous cell carcinoma to chemoradiotherapy.

BACKGROUND: Multimodality therapy with chemotherapy and radiotherapy followed by surgery may improve survival in patients with esophageal squamous cell carcinoma compared with each of the individual treatment options. Histologic assessment of resected tumors after chemoradiotherapy shows that some patients have a complete response with no residual tumor, whereas other patients derive no benefit. The ability to predict response to chemoradiotherapy would allow treatment to be planned accordingly. METHODS: Expression of the tumor growth and proliferation proteins epidermal growth factor receptor (EGFR) and proliferating cell nuclear antigen (PCNA) was determined using immunohistochemical staining of pretreatment endoscopic biopsies from patients with esophageal squamous cell carcinoma who were randomized to chemoradiotherapy before surgery. Response to chemoradiotherapy was assessed by histologic examination of the resected specimens. Response to chemoradiotherapy and survival were correlated with EGFR and PCNA expression individually and with both markers combined as EGFR/PCNA: RESULTS: Of 14 patients available for study, 6 had a complete histologic response (CR) to chemoradiotherapy with no residual tumor in the resected specimen, 3 had a partial response (PR) to chemoradiotherapy, and the remaining 5 had minimal response (MR). Of the nine patients with a CR or PR, tumors of eight patients were negative for one or both markers. Of the five patients with an MR, four tumors were positive for both EGFR and PCNA (P < 0.05, Fisher's exact test). Comparison of survival from the date of randomization shows that patients with tumors negative for one or both markers had a significant survival advantage (P = 0.0003, log-rank test). CONCLUSIONS: Evaluation of PCNA and EGFR status of pretreatment biopsies may identify a group of patients likely to derive the greatest benefit from chemoradiotherapy before surgery in terms of histologic response and long term survival.

Hyperhomocysteinaemia and multiple aneurysms.

Homozygous homocystinuria, the most common genetic disorder of transulphuration, is associated with elevated plasma concentrations of homocystine, homocysteine, multiple clinical abnormalities and life-threatening thromboembolism. Several instances of vascular aneurysms have also been documented. More recently, an association between premature occlusive vascular disease and the heterozygous state has been proposed. We now report an unusual case in whom multiple aneurysms were associated with heterozygous homocystinuria.

Myocardial hypertrophy, fibrosis and infarction following exposure of the heart to radiation for Hodgkin's disease.

A 35 year old man was treated for stage IIA Hodgkin's disease by radiation to the upper thorax, axillae and neck. Three years later he presented with intractable and ultimately fatal congestive heart failure. Autopsy revealed massive biventricular hypertrophy with widespread subendocardial fibrosis and myocardial infarction, but with little coronary artery disease. Such a complex of features has not previously been described after radiation therapy and cannot be adequately explained by other known causes of heart muscle disease. Ventricular hypertrophy with extensive subendocardial fibrosis may be part of the spectrum of radiation heart disease.

Fatal Clostridium welchii septicaemia following acute cholecystitis.

A case of Clostridium welchii septicaemia following acute cholecystitis is described. The onset was acute and a rapidly fatal outcome ensued. Radiological findings were negative. An approach to the antibiotic treatment and general management is discussed.