Alterations of copy number of methylation pattern in mismatch repair genes by methylation specific-multiplex ligation-dependent probe amplification in cases of colon cancer.

Genetic alterations and changes in genomic DNA cytosine methylation patterns are associated with all types of cancer and are caused by germline mutations in DNA mismatch repair (MMR) genes, predominantly MLH1 (MutL homolog 1, 19 exons) and MSH2 (MutS homolog 2, 16 exons). Genomic DNA was extracted from tissue samples embedded in paraffin from 49 patients with adenocarcinoma and from 21 patients with carcinoma for the study group; genomic DNA was extracted from lymphocytes from 10 healthy donors for the control group. We used methylation specific multiplex ligation-dependent probe amplification (MS-ML-PA), which allows the detection of copy number changes and unusual methylation levels of 10 to 50 different sequences in one reaction by use of the methylation-sensitive restriction enzyme HhaI and sequence-specific capillary electrophoresis for the study of 24 genes. We found the mean methylation rates for MLH1 (97.14%), MSH2 (24.28%), MSH6 (MutS homolog 6) (67.14%), MSH3 (MutS homolog 3) (78.57%), MLH3 (MutL homolog 3) (75.71%), PMS2 (postmeiotic segregation increased 2) (65.71%), MGMT(O-6-methylguanine-DNA methyltransferase ) (82.85%). We conclude that the mismatch repair (MMR) system is critical for the maintenance of genomic stability.

Primary Cardiac Angiosarcoma with Right Coronary-To-Right Atrium Fistula : Case Report

Primary tumours of the heart are rare. About 25of all cardiac tumours are malignant and the most common of these is the angiosarcoma. We present a 61-year-old male with a right atrial angiosarcoma that was detected on coronary angiography. The tumour showed marked vascularity and a right coronary-to-right atrium fistula; and the patient underwent surgical resection. Pathological examination of the tumour was consistent with a cardiac angiosacoma and the diagnosis was also confirmed by immuno-histochemistry. He consequently underwent chemotherapy; however the patient died 60 days after the surgery

Effects of Iloprost and pentoxifylline on renal ischemia-reperfusion in rabbit model.

OBJECTIVE: In ischemia-reperfusion, Iloprost decreases neutrophil activation and aggregation besides inhibition of oxygen-free radical production. Pentoxifylline (Ptx) attenuates reperfusion-associated membrane injury and tissue edema, suppresses leukocyte adhesion and improves hindlimb blood flow during the reperfusion period. The primary hypothesis in this study was that Iloprost could present better protection than pentoxyfillin on renal ischemia-reperfusion in rabbit model. MATERIALS AND METHODS: Forty rabbits were grouped into four. Iloprost was continuously infused starting half an hour before the reperfusion after 2 hours ischemia and during the 4 hours reperfusion period in Group 1 whereas the Group 2 was treated with pentoxyfillin. Group 3 was the control group which didn't receive any medication. Forth group was sham group. Renal tissues were histologically and biochemically evaluated. RESULTS: The histologic scores were obtained according to presence of tubuler necrosis and atrophy, regenerative atypia, hydropic degeneration (Group 1 vs Group 3; p<0.001, Group 2 vs Group 3; p = 0.001, Group 1 vs Group 2; p = 0.331). Malondialdehyde levels of the medicated groups were 109 +/- 11 nmol/gr tissue in Group 1, 119 +/- 15 nmol / gr tissue in Group 2 and 132 +/- 14 nmol / gr tissue in Group 3 (Group 1 vs Group 2; p = 0.130, Group 1 vs Group 3, p = 0.002, Group 2 vs Group 3; p = 0.045). Malondialdehyde levels and histologic scores of all of the groups were significantly different from the sham group. CONCLUSION: Iloprost and pentoxyfillin reduced renal ischemia-reperfusion injury in rabbit model. There was not a significant difference between these two medications.

Toxic hepatitis associated with paroxetine.

Hepatotoxicity is a rare complication of paroxetine, a selective serotonin reuptake inhibitor. Regarded as safe in therapeutic use, there have been reports of cases of severe hepatic dysfunction with gross elevations of transaminase levels that may be related to this drug. We report here severe adverse cholestatic and hepatocellular injury in a patient taking paroxetine probably due to an immune-mediated hypersensitivity reaction.

Desmoplasia measured by computer assisted image analysis: an independent prognostic marker in colorectal carcinoma.

AIMS: The assessment of desmoplasia by traditional semiquantitative methods does not provide reliable prognostic data. The aim of this study was to quantify desmoplasia by computerised image analysis in primary colorectal carcinomas and to investigate its ability to predict overall survival. METHODS: In total, 112 colorectal adenocarcinomas, with a median follow up of 66 months, were studied. The representative tumour sections were stained by the van Gieson method, which stains collagen rich stroma red. For quantitative histochemical measurement, digital images were analysed by a computerised image analysis program to calculate the percentage of red stained tissue area. The percentage of desmoplasia (PD) was related to conventional clinicopathological prognostic factors and overall survival. RESULTS: The mean (SD) PD was 4.85 (3.37). PD was found to be significantly associated with lymph vessel and venous invasion. By Kaplan-Meier analysis, PD was associated with survival-patients with PD > 4 had a shorter survival than those with PD

Glucose transporter-1 (GLUT-1): a potential marker of prognosis in rectal carcinoma?

The aim of the study is to evaluate the pattern and level of expression of glucose transporter-1 (GLUT-1) in rectal carcinoma in relation to outcome as a potential surrogate marker of tumour hypoxia. Formalin-fixed tumour sections from 43 patients with rectal carcinoma, who had undergone radical resection with curative intent, were immunohistochemically stained for GLUT-1. A mean of three sections per tumour (range 1-12) were examined. Each section was semiquantitatively scored; 0, no staining; 1, <10%; 2, 10-50%; 3, >50% and a score given for the whole section, the superficial (luminal) and deep (mural) part of the tumour. Staining was seen in 70% of tumours. Increased staining was noted adjacent to necrosis and ulceration. A diffuse and patchy pattern of staining, with and without colocalisation to necrosis was seen. Patients with high GLUT-1-expressing tumours (score 3 vs 0-2) had a significantly poorer overall survival (P=0.041), which was associated with poorer metastasis-free survival with no difference in local control. No significant correlation was seen with other prognostic factors. There was a strong correlation between the score for the superficial and deep parts of the tumour (r=0.81), but a significant relationship with outcome was only found in the deep part (P=0.003 vs P=0.46). In conclusion, increased GLUT-1 expression in rectal tumours was an adverse prognostic factor and is worth further evaluation as a predictive marker of response to therapy.

Relationship of angiogenesis and p53 protein expression in colorectal carcinomas.

OBJECTIVE: Recently, new functions have been attributed to the p53 protein, particularly a prominent role in the regulation of angiogenesis. Tumours expressing mutant forms of p53 protein may be associated with increased angiogenesis. The aim of this study is to investigate the relationship between p53 protein expression and the quantitative expression of tumour angiogenesis in colorectal carcinomas. PATIENTS AND METHODS: Sections from paraffin-embedded blocks from 46 patients with primary colorectal carcinomas that had been completely removed were analysed. p53 protein expression and all vascular structures were evaluated by immunohistochemistry. The vessel parameters of angiogenesis including vascular surface density (VSD), number of vessels per mm2 (NVES) and number of vessels in unit area (n) were assessed by morphometry. Mann-Whitney U-test was used for comparing the extent of neovascularization in p53-positive and -negative cases. RESULTS: Twenty-four (52%) cases were p53+ and 22 (48%) were p53-. Mean VSD, NVES and n values for p53 protein-positive and -negative groups were as follows: VSD 96.7 +/- 65.4/mm vs 79.6 +/- 45.24/mm; NVES 104.8 +/- 97.5/mm2 vs 62.2 +/- 44.3/mm2; n 79.7 +/- 74.2 vs 52 +/- 35.7, respectively. There was no association between the angiogenesis parameters and p53-positive and -negative cases, when VSD (P=0.226) or n (P=0.176) were considered, but a statistically significant difference was obtained for NVES values (P=0.035). CONCLUSION: The authors concluded that tumoural angiogenesis assayed by morphometric investigation in colorectal carcinomas might be related to p53 protein expression when NVES is considered. This finding supports the possible role of p53 protein in increased angiogenesis in colorectal tumours.

Prognostic significance of angiogenesis in rectal cancer: a morphometric investigation.

BACKGROUND: Neo-angiogenesis is crucial for tumor growth and metastasis and has been proposed as an independent prognostic factor for survival in patients with solid tumors. In this study the quantitative expression of angiogenesis was investigated by direct stereologic assessment of the vascular surface density in rectal carcinoma to determine the possible correlation of angiogenesis with clinicopathological factors and prognosis. PATIENTS AND METHODS: Sections from formalin-fixed paraffin-embedded tissue blocks of 29 primary rectal carcinomas were resected and immunostained for endothelial cell factor-VIII-related antigen. The vascular surface density (VSD), number of vessels per square mm (NVES), maximum NVES (NVESmax) according to the three maximum values of NVES and number of vessels in the unit area (N) were assessed by means of morphometry. The results were related to the main prognostic variables and the survival of patients. RESULTS: There were no significant differences between survivors and non-survivors in terms of the angiogenesis parameters that were investigated. The overall survival rate was not significantly different for sex, age, tumor size and differentiation, extrahepatic metastasis, depth of invasion and the mode of adjuvant therapy. However, a significantly lower overall survival rate was observed in patients with liver metastatic disease (p<0.001), lymph node involvement (p=0.04) and incomplete resection (p<0.001). Multivariate analysis indicated that only the number of vessels in the unit area (HR = 1. 028, p = 0. 04), hepatic metastases (HR=14.94, p=0.007) and type of resection (HR=23.81, p=0.004) predicted overall survival. CONCLUSION: These findings suggest that increased tumoral vascularity, consistent with previous studies, adversely affects survival in rectal cancer patients. Liver metastatic status and completeness of the surgical resection were the most powerful criteria to predict the final outcome of these patients. Thus, neo-angiogenesis is indeed an important and key step in tumorigenesis, but it may not be the single overwhelming factor that determines recurrence and metastasis in rectal carcinoma.

Prognostic value of CD44 expression in colorectal carcinomas.

BACKGROUND: CD44 has diverse functions in cell-cell and cell-matrix interactions and its expression appears to be an indicator of invasive and metastatic behaviour in carcinomas. However, contradictory data have been reported about the correlation between CD44 expression and prognosis in colorectal carcinomas. We aimed (i) to establish whether immunohistochemically detectable CD44 expression is related to tumor aggressiveness, (ii) to correlate CD44 expression with the degree of tumor differentiation and (iii) to determine the relationship between CD44 expression and patient survival and other conventional clinicopathological features. PATIENTS AND METHODS: The immunohistochemical expression of CD44 in a series of 111 colorectal carcinomas was examined using the monoclonal mouse anti-human phagocytic glycoprotein-1, CD44 (clone DF 1485) in correlation with clinicopathological variables. To achieve a reliable semi-quantitative evaluation, not only the staining intensity but also the distribution of positive tumor cells were analyzed. RESULTS: CD44 staining was high-grade positive in 42 and low-grade positive/negative in 69 tumor tissues. There was no association between CD44 expression and tumor size, histological differentiation, depth of invasion, lymph node involvement, clinical stage of the disease, or the radicality of surgical resection. CD44 expression was not correlated significantly with recurrence and distant metastases. Multivariate analysis showed that only the modified Astler-Coller (MAC) staging system was an independent prognostic factor of recurrence (HR=15.267; 15.267-6.808, 95% CI; p=0.001) and survival (HR=37.064; 13.309-103.220, 95% CI; p=0.001). Kaplan-Meier curves showed that there was no significant association between CD44 expression and recurrence and overall survival in either MAC B or C colorectal cancer. CONCLUSION: Expression of CD44 was not associated with any conventional clinicopathological features. CD44 cannot be considered as a prognostic predictor of recurrence, metastasis and overall survival.

Origin of giant cells in osteoclast-like giant cell tumors of the pancreas.

To clarify the origin of giant cells in osteoclast-like giant cell tumors (OGCTs) of the pancreas, we performed microscopical, immunohistochemical, and K-ras gene mutation analyses with a microdissection approach in 3 cases, featuring 4 cellular components (osteoclast-like giant cells [OGCs], pleomorphic large cells [PLCs], mononuclear cells, and ductal carcinoma cells). Two cases had abundant OGCs, and 1 case contained large number of both OGCs and PLCs. In each, none of the microdissected OGCs contained any K-ras gene mutation while they were positive for a histiocytic marker (CD-68). In contrast, PLCs, when present, frequently harbored K-ras gene mutations and were negative for CD-68. In all cases, mononuclear cells, a mixture of histiocyte-like and atypical, from microscopic and immunohistochemical viewpoints, also frequently showed K-ras alteration. Histiocyte-like mononuclear cell was equipped with a regular and oval nucleus similar to those in OGCs and was positive for CD-68. Atypical mononuclear cell showed an irregular, pleomorphic, or sometimes bizarre nucleus similar to those in PLCs and was negative for CD-68. All of the K-ras gene mutations found in PLCs and mononuclear cells were the same as in the ductal carcinoma cells within the same tumor. Thus, OGCs differ in origin from ductal cells and are strongly suggested to be nonneoplastic and of mesenchymal origin, whereas PLCs, which harbor K-ras gene mutations, are neoplastic and presumably derived from ductal carcinoma cells. Moreover, mononuclear cells may be classified into 2 types, histiocyte-like and atypical.

Angiogenesis and p53 and H-ras mutations in pancreatic ductal adenocarcinoma.

OBJECTIVE: To evaluate the correlation of angiogenesis and p53 and H-ras mutations with prognostic factors and proliferative activity assessed with Ki-67 protein expression by studying archival tissues from 24 patients with primary pancreatic ductal adenocarcinoma. STUDY DESIGN: Vascular structures were labeled immunohistochemically using factor VIII-related antigen. Vascular surface density (VSD) and microvessel number (NVES) were assessed by stereology. The tissues were also analyzed with the immunohistochemical method for the expression of proteins, including p53, H-ras and Ki-67. RESULTS: Statistical analysis revealed that tumors with greater NVES and VSD values significantly correlated with occurrence of metastases, higher proliferative activity, poorer histologic differentiation and greater tumor size. p53 Mutations were found in 11 cases (45.8%). However, only three cases (12.5%), all negative for p53 mutations, showed H-ras mutations. p53 Mutation-positive tumors exhibited a statistically significant correlation with occurrence of metastases and higher proliferative activity, whereas H-ras mutations did not show such a correlation. CONCLUSION: Angiogenesis might have a role in predicting prognosis in pancreatic carcinomas, and p53 mutations might be acquired in later stages associated with metastatic progression and higher proliferative activity. Although H-ras mutations were rare in the present study, they might play a role in a different carcinogenic pathway excluding p53 mutations.

Perforated solitary ulcer of the colon. Report of a case.

PURPOSE: A patient with a solitary colonic ulcer had sudden onset of crampy abdominal pain, anorexia, fever, and vomiting, with signs of positive peritoneal irritation. METHODS: The diagnosis was proved by histopathologic examination of right hemicolectomy material. RESULTS: An emergency laparotomy, with right hemicolectomy and ileotransversostomy, gave complete relief from symptoms. The patient was still asymptomatic at the two-year follow-up, and control colonoscopic examinations performed at 6 and 18 months after the operation were normal. CONCLUSION: Preoperative diagnosis of perforated solitary colonic ulcers localized at the right hemicolon may mimic acute appendicitis, and intraoperative findings may mimic colonic carcinoma. If the preoperative diagnosis is not certain, right hemicolectomy and ileotransversostomy, with regular colonoscopic controls, is a safe procedure in the treatment and follow-up of these patients.

A case of juvenile ankylosing spondylitis and Crohn's disease.

Juvenile ankylosing spondylitis (JAS) is a chronic inflammatory arthritis of the peripheral and axial skeleton, frequently accompanied by enthesitis. About four percent of patients with JAS have ulcerative colitis or Crohn's disease. Crohn's disease is the more common of the two and is diagnosed in 26 percent of patients with chronic spondyloarthropathy. In this paper, a 14-year-old male patient is presented as a typical case of juvenile ankylosing spondylitis and Crohn's disease with low back pain, morning stiffness, limited motion in anterior and lateral flexion and extension, left sacroiliitis, ankylosis in the apophyseal joints of the lumbar vertebrae, abdominal pain, bloody diarrhea, characteristic histopathologic changes of colonic involvement such as lymphoid follicles, fissures, submucosal polymorphonuclear cell infiltration and definite ganglion cells. The current therapy with mesalazin, having fewer side effects than sulfosalazin, and its applicability in combination with naproxen sodium is also discussed.

IgA nephropathy in mice following repeated administration of conjugated Haemophilus influenzae type B vaccine (PRP-T).

OBJECTIVE: In this study, the risk of IgA nephropathy in Swiss albino mice following the subcutaneous administration of conjugated Haemophilus influenzae type b vaccine (PRP-T), containing capsular polysaccharide of the organism (PRP) conjugated to tetanus protein (T), was evaluated. METHODS: Three treatment and corresponding control groups, each containing mice, were constituted and given 2, 4, 6 injections of 1/4 HD of PRP-T or placebo, respectively, at 2-week intervals. All mice in each treatment group were sacrificed two weeks from the last injection to examine sequential glomerular changes. RESULTS: The niceoscpic examination of renal tissues revealed mesangial proliferation (6/7; 85%) in the first group given 2 doses of vaccine; mesangial proliferation (5/7; 72%) and increase in matrix (7/7; 100%) in the second group given 4 doses; and mesangial proliferation (7/7; 100%), increase in matrix (7/7; 100%), IgA (7/7; 100%) and C3 (3/7; 42%) deposition within mesangium in the third group given 6 doses. No histopathological changes were detected in the renal tissues of any control mouse. When the experimental groups were compared statistically with their respective controls at the light microscopic level, mesangial proliferation in the first group (p: 0.0047), mesangial proliferation (p: 0.021) and increase in matrix (p: 0.001) in the second group, mesengial proliferation (p: 0.001) and increase in matrix (p: 0.001) in the third group were determined to be significantly different. When study and control groups were compared by immunofluorescence microscopy, only the third group revealed a statistically significant difference with respect to IgA deposition (p: 0.001). C3 deposition was also demonstrated in this group, but it was not significantly different (p: 0.192). However, in no instance was a control mouse found to have any form of immune deposition. CONCLUSION: We concluded that conjugated Haemophilus influenzae type b vaccine, given at two-week intervals to a total of six doses, caused secondary IgA nephropathy in mice.

Behçet's disease with pulmonary involvement, superior vena cava syndrome, chyloptysis and chylous ascites.

Behçet's disease is a chronic multisystem vasculitis of unknown aetiology. This case report describes a patient who applied to the hospital because of dyspnoea, ascites, oedema of lower extremities and recurrent episodes of haemoptysis. For the last 12 yr, he had superior vena cava syndrome (SVCS) and cardiac and pulmonary involvement of Behçet's disease, and biochemical examination of ascite fluid yielded a chylous effusion containing triglyceride 421 mg dl-1 and cholesterol 49 mg dl-1. Chyloptysis was also detected by Sudan III stain. The patient died from cardiac tamponade in spite of cardiac fenestration. To the authors' knowledge, this is the first reported case of Behçet's disease with chylous ascites and chyloptysis in the English literature.

The effect of acute bilateral adrenalectomy on serotonin-induced inhibition of gastric acid secretion and acute gastric mucosal injury in rats.

Serotonin (5-hydroxytryptamine, 5-HT) produces many changes in gastric functional parameters, including the inhibition of gastric acid secretion and changes in mucosal blood flow. Exogenous 5-HT has also been shown to induce gastric erosion. The influence of adrenalectomy on experimental lesions in the rat gastric mucosa remains controversial. The aim of this study was to see the effects of adrenalectomy on pentagastrin stimulated gastric acid secretion in anaesthetized male Wistar rats. Gastric acid was collected via cannulae placed in the stomach. 5-HT (3.5 mumol/kg, i.v.) inhibited pentagastrin stimulated acid output by 54% and produced haemorrhagic gastric lesions with a mean ulcer index of 2 +/- 0.3. Adrenalectomy prevented both 5-HT induced inhibition of gastric acid secretion and mucosal injury. The results suggest that the effects of 5-HT require an intact adrenal gland.

Serotonin causes acute gastric mucosal injury in rats, probably via 5HT1D receptors.

5-HT-induced acute gastric mucosal injury was assessed in rats by using 5HT1, 5HT2, 5HT3, 5HT4 or muscarinic receptor related drugs. Rats were treated with antagonists i.p. and 30 minutes later either vehicle, 5-HT (20 mg/kg) or other agonists were administered s.c. The stomachs were removed 4 hours after the last injection and mucosal integrity was assessed by light microscopy using a histological ulcer index (HUI). The HUI was found to be significantly increased following 5-HT administration (1.57 +/- 0.3) when compared with controls (0.14 +/- 0.1). 5HT1 agonist 5-carboxamidotryptamine (20 mg/kg) produced acute gastric erosion and increased the HUI (P < 0.05). The HUI in the animals receiving 5-HT1D agonist sumatriptan (7 mg/kg) was found to be 1.62 +/- 0.24. 5HT2 antagonist ketanserine (2.5-15 mg/kg), 5HT3 antagonist ondansetron (1-5 mg/kg), 5HT4 antagonist DAU 6285 (1-10 mg/kg) and atropine (1.5-30 mg/kg) exerted no effect whereas 5HT1/2 antagonist metitepine (0.05-0.5 mg/kg) caused a dose dependent inhibition of the effect of 5-HT. The results from this study demonstrate that 5-HT causes acute gastric mucosal injury and this injury is probably due to the activation of the 5-HT1D receptors.

A case of common truncus arteriosus with rare cardiac abnormalities.

Truncus arteriosus communis is a well-known, congenital, cardiac abnormality. In this report, we describe a very rare variant in which the truncus arose from the right ventricle, the atrial and ventricular septa were intact, the left ventricle was hypoplastic and the small right atrium was draining to the truncus arteriosus through a fibrose tunnel-like connection.