Correlation between DNA methyltransferases expression and Epstein-Barr virus, JC polyomavirus and Helicobacter pylori infections in gastric carcinomas.

It' is accepted that aberrant expression of DNA methyltransferases (DNMTs) is responsible for hypermethylation in genes. However, there are limited data related to factors inducing aberrant expression of DNMTs. A total of 43 surgically resected gastrc carcinomas (GC) samples were analysed. Using immunohistochemistry assay we have determined expression level of DNMT1 and 3b. The presence of H.pylori was evaluated by histology, whereas JC polyomavirus (JCV) and Epstein-Barr virus (EBV) detection were carried out by PCR and in situ hybridization techniques, respectively. High expression of DNMT1 and 3b were detected in 46.5% and 53.5% of GC cases, respectively. Co-expression of DNMT1 and 3b were found in 37.2% of cases. Using different techniques, H. pylori, JCV and EBV were detected in 55.8%, 32.6% and 9%, respectively. Moreover, in 37% of cases, we noted the presence of JCV and/or EBV infections. H.pylori co-infection was found in 64.3% (9/14) of JCV positive cases and in 50% of EBV positive GC, without a reliable significant relationship. Correlation analyses have showed a marked increase in DNMT1 expression in EBV associated GC (P= 0.02). Also, co-expression of DNMT1 and 3b was significantly associated with EBV infection in GC (P=0.05). Similarly, JCV associated GC mostly displayed DNMT1 positive status, but the difference did not reach the significant threshold. Nevertheless, infection with JCV and/or EBV was significantly correlated with increased expression of DNMT1 in GC (P= 0.05). Our study suggests that EBV and JCV infections in GC correlated with deregulation of DNA methyltransferases.

Clinicopathologic characteristics of colorectal cancer with microsatellite instability.

Colorectal cancer (CRC) can be classified according to the level of microsatellite instability (MSI) exhibited by the tumor. The aim of this study was to determine MSI status in CRC from Tunisia and to identify clinical and pathological characteristics of MSI-H tumors. Microsatellite status was determined by polymerase chain reaction amplification using standard markers (BAT25, BAT26, D2S123, D5S346 and D17S250, the Bethesda panel) in 44 CRC cases. Molecular results were correlated with pathological and clinical features. Six CRC cases (13.8%) showed high-level instability (MSI-H), 14 cases had low level instability (MSI-L), and the remainders were stable (MSS). Immunohistochemical analysis showed loss of MSH2 protein in 3 cases among the 6 MSI-H tumors, whereas no silencing of MLH1 or MSH6 was found in any case. Significant differences in age and family history of cancers were observed between MSI-H and MSS/MSI-L groups (p=0.01 and p=0.002). However, statistical analysis showed that there were no significant differences between MSI-H and MSS/MSI-L tumors in terms of tumor location, lymph node involvement and stage of disease. Regarding histological features, MSI-H tumors were more likely to be poorly differentiated (p=0.003), to have a medullary pattern (p=0.005), and to harbor increased numbers of peritumoral lymphocytes (p=0.001). These findings indicate that careful observation of the tumor morphology can assist in the identification of unstable colorectal cancers requiring molecular investigations.

Cervical unicentric Castleman's disease in children.

INTRODUCTION: Unicentric Castleman's disease (CD) is a rare benign lymphoproliferative disorder. A cervical location has rarely been reported in the pediatric literature. CASE REPORT: A 13-year-old boy presented with a mass in the right neck region of 3 months' evolution. Clinical examination revealed a smooth mobile 5 cm node deep in the sternocleidomastoid muscle. The diagnosis of hyaline vascular type CD was confirmed by complete surgical excision and histopathologic examination. Postoperative course was simple, without recurrence over 1 year's follow-up. DISCUSSION AND CONCLUSION: Cervically located CD often shows few symptoms, and is rarely suggested by a cervical mass found in children. Definitive diagnosis is histopathological. The hyaline vascular type of unicentric CD has a good prognosis after complete resection of the lesion.

[Virus-induced cancers: interplay between genetics and environment]. / Les cancers viro-induits: interrelations génétique-environnement.

Among cancers diagnosed worldwide on a yearly basis, 20% are thought to be associated with a viral infection. The viruses involved are, by order of decreasing incidence, the hepatitis viruses, the papillomaviruses and the Epstein-Barr virus. These virus-induced cancers generate a high level of interest not only for the study of mechanisms involved in the neoplastic transformation, but also for the set-up of specific immunotherapies including prophylactic and therapeutic antitumor vaccination.

Patterns of aberrant DNA hypermethylation in nasopharyngeal carcinoma in Tunisian patients.

BACKGROUND: Aberrant methylation in the promoter of tumor-related genes is associated closely with epigenetically mediated gene silencing. The aim of the present study was to evaluate the methylation profile of Tunisian nasopharyngeal carcinoma (NPC) and to determine the clinicopathological features of tumors showing this epigenetic alteration. METHODS: Thirty-six archival NPC biopsies were investigated in comparison with 19 non-tumor nasopharyngeal tissue specimens. DNA methylation status of ten tumor-suppressor and related genes was analyzed by using methylation-specific PCR. The Epstein-Barr virus (EBV) presence was verified by PCR and in situ hybridization and the LMP1 oncoprotein expression was analyzed by immunohistochemistry. Findings were then correlated with clinicopathological variables (Patients' gender and age, tumor histological subtype and stage). RESULTS: Hypermethylation frequencies of the investigated genes in NPC biopsies were 75% for RASSFIA, 58.3% for SHP1, 47.2% for DAPK, 33.3% for P16, 31% for RARß2, 19.4% for GSTP1 and TIMP3, 11% for APC and CDH1, and 5.5% for MGMT. In non-tumor nasopharyngeal samples, hypermethylation was detected in lower frequencies in 6 genes (SHP 26.3%, P16 21%, RARß2 21%, DAPK 15.8%, TIMP3 10.5%, and GSTP 5.3%). Hypermethylation of RARß2 promoter was more frequent in tumors with lymph node metastasis than those without metastasis (43.5% vs 0%, p=0.03). Methylation of RASSF1A was more frequently detected in non-keratinizing NPC than in undifferentiated subtype (100% vs 66.7%; p=0.05). A trend toward positive association was found between an increased number of methylated genes and LMP1 expression (p=0.07). However, no significant association was found for the remaining variables. CONCLUSIONS: This study indicates that hypermethylation of multiple genes is a common alteration in nasopharyngeal carcinomas in Tunisian patients and that this epigenetic change may play a role in the nasopharyngeal carcinogenesis.

Immunodetection of SV40 T/t-antigens in human osteosrcoma in a series of Tunisian patients.

Osteosarcoma is a primary bone malignancy that typically occurs during adolescence but also has a second incidence peak in the elderly. The etiology of osteosarcoma is not well understood. Recent investigations have identified SV40 DNA sequences in osteosarcomas, suggesting that SV40 may contribute to tumor development. However, these studies also demonstrated geographical differences in SV40-positive osteosarcomas. The purpose of this study was to determine the prevalence and clinicopathological characteristics of SV40 positive osteosarcoma in Tunisian patients. Fifty-six formalin-fixed paraffin-embedded specimens of osteosarcomas were retrospectively investigated. Samples investigated were clinical cases examined between 1990 and 2004 in the Laboratory of Pathology at the University Hospital Farhat-Hached of Sousse (Tunisia). The search for SV40 was performed by immunohistochemistry using the Pab108 antibody for the detection of the viral oncoproteins: large T antigen and small t antigen (T/t-ag). SV40 status was correlated with clinico-pathological data. T/t-ag immunostaning was detected in the tumor cells in 31/56 (55.4%) osteosarcoma cases. SV40 positivity was more frequent (83%) in patients older than 40 years (5/6 cases) than in patients under 40 years (52%, 26/50), but the difference does not reach statistical significance (p = 0.33). Moreover, the time between the onset of clinical symptoms and diagnosis was shorter for SV40 positive than SV40 negative cases (p = 0.08). However, the viral status did not differ significantly according to gender, tumor size, histological subtype, tumor location, or metastases. This study documents the presence SV40 T/t-antigens in a proportion of osteosarcomas in Tunisian patients. The expression of these viral oncoproteins supports the hypothesis that SV40 may have a role in the pathogenesis of this tumor.

Contribution of epigenetic alteration of BRCA1 and BRCA2 genes in breast carcinomas in Tunisian patients.

OBJECTIVE: The aim of this study was to evaluate the contribution of the BRCA1 and BRCA2 promoter methylation in the pathogenesis of sporadic breast cancer in Tunisian patients. METHODS: Breast carcinoma tissues (n=117) and available paired normal breast tissues (n=65) from Tunisian women who had no family history were investigated for the methylation status of BRCA1 and BRCA2 promoters using methylation-specific PCR. Breast specimens from women without carcinoma (16 fibroadenomas and 5 mastopathies) were used as control. RESULTS: Hypermethylation of BRCA1 and BRCA2 promoters was detected respectively in 60.7% and 69.2% of the carcinoma tissues, and in only 7.7% and 4.6% of the paired normal breast tissues. None of the fibroadenomas and mastopathies showed hypermethylation. Correlations were found between BRCA1 and BRCA2 hypermethylation and decrease in their mRNA expression (p=0.02 and p=0.009, respectively). Moreover, BRCA1 methylation correlates with patients age (p=0.01) and triple negative (ER-, PR-, HER2-) tumors (p=0.01). Patients with methylated BRCA1 and/or BRCA2 had a significant prolonged survivals compared to those with unmethylated tumors (p=0.002). CONCLUSION: Our results suggest an important role of BRCA1 and BRCA2 promoter methylation in breast cancer development in the Tunisian population.

Accumulation of mercury and its effects on testicular functions in rats intoxicated orally by methylmercury.

All forms of mercury are considered poisonous. Methylmercury, one organic form, is highly toxic to many organs. The aim of the present study was to assess the effects of this form on the reproductive system in the rat. For this, 20 male rats were divided into two groups. One, which is considered as reference, received tap water. The second group received tap water containing methylmercury at the rate of 20 mg l⁻¹ for 8 weeks. At the end of the experiment, blood samples were collected for the determination of total mercury and plasma testosterone. The left testes were used for the determination of total mercury and histological examination. Appropriate centrifugation was applied on right testes to extract interstitial and seminiferous tubular fluids. The epididymides were homogenised for the sperm count. Our results showed a dramatic fall in the plasma testosterone in the contaminated animals. The fall in plasmatic testosterone seems to be in relation with the decrease in the secretion of testosterone. In association with this, the concentration of testosterone in seminiferous tubules fluid dropped about 55% in the poisoned animals in comparison with the controls. Despite this, no decrease in the epididymal sperm count in contaminated rats was observed.

[Eighteen cases of craniofacial fibrous dysplasia]. / Dix-huit cas de dysplasie fibreuse craniofaciale.

INTRODUCTION: Fibrous dysplasia or Jaffe Lischtenstein's disease is sporadic and rare bone affection, mainly observed in young adults. It is characterized by a bone maturation disorder that can affect one or several bones. Fibrous dysplasia is relatively rare in the craniofacial region, (only 20% of all locations). The authors analyzed the epidemiological, anatomoclinical and evolutive features of fibrous dysplasia in a Tunisian population. MATERIAL AND METHOD: The authors retrospectively studied 18 cases of craniofacial fibrous dysplasia, diagnosed at the pathology department of the Sousse F. Hached hospital, beween1990 and 2005. RESULTS: Most craniofacial dysplasia cases were monostotic (94% of cases). One case was polyostotic. The maxilla and the mandible were the most frequent locations (83.3%). The patients' mean age was 28.6 years, ranging from 6 to 30 years. The sex ratio was 3.5 in favor of women. Follow-up ranged from one month to 10 years with an average of 26.4 months. One case of sarcomatous transformation was observed. DISCUSSION: The authors noted epidemiological discrepancies compared to what was usually reported, especially concerning the strong female predominance, the frequency of maxillary and mandibular locations and the predominance of monostotic presentations. Larger series are needed to validate these observations.

Evidence for a role of the Simian Virus 40 in human breast carcinomas.

AIMS OF THE STUDY: The aim of this study was to investigate whether the Simian Virus 40 (SV40) is implicated in human breast carcinomas (BC). EXPERIMENTAL DESIGN: SV40 presence was investigated by PCR assays targeting the Tag, the regulatory, and the VP1 regions in 109 invasive breast ductal carcinomas from Tunisian women. We also examined the relationship between the presence of SV40 and promoter methylation status of 15 tumor-related genes. Immunohistochemistry was used to investigate the expression of Tag, estrogen and progesterone receptors, HER2, and P53. RESULTS: SV40 DNA sequences were detected in 22% of tumors and in only 1.8% of the matched non-tumoral tissues. Using immunohistochemistry, SV40 was detected in the tumor cells. Hypermethylation frequencies were 78% for RASSF1A, 66% for SHP1, 61% for HIN1 and BRCA1, 47% for P16 and ER, 42% for CDH1 and APC, 40% for BLU, 35% for DAPK, 34% for RARbeta2, 27% for GSTP1, 17% for TIMP3, 14% for CCND2, and 8% for hMLH1. Interestingly, the frequencies of RASSF1A, SHP1, BRCA1, and TIMP3 methylation, and the mean of the methylation index (MI) were significantly higher in SV40-positive than in SV40-negative cases (P-values ranging from 0.043 to 0.003). Moreover, SV40 presence correlates with P53 protein accumulation (32.7% vs. 13.3%; P=0.015) and HER2 low expression (3.7% vs. 28%; P=0.008). We also found SV40 more frequently in patients over 50 years than in younger patients (34.8% vs. 12.3%; P=0.006). CONCLUSIONS: This study is the first to demonstrate the presence of SV40 in human BC and provides data supporting a role for this virus in the pathogenesis of these tumors.

Prevalence and characteristics of the MMTV-like associated breast carcinomas in Tunisia.

The involvement of a retrovirus homologous to the mouse mammary tumor virus (MMTV) in the pathogenesis of human breast cancer (BC) has long been assumed, but has never been proven. Previous studies have reported the detection of MMTV-like env sequences in variable proportions that did not exceed 40% of BC cases in several countries. However, these viral sequences have been found in higher proportion (74%) in Tunisian diagnosed with BC during the seventies. This study is an attempt to evaluate the current prevalence of MMTV-like env gene in BC in Tunisian women. We used semi-nested PCR that amplify a 190-bp MMTV-like env sequence, followed by direct sequencing to screen a series of 122 cases of BC randomly selected. The findings were correlated to clinicopathological data and immunohistochemical expression status of progesterone and oestrogen receptors, HER2, and P53. Specific MMTV-like env sequences were found in 17 (13.9%) cases of breast carcinomas, whereas the same sequences were not detected in matched normal breast tissues. The presence of the viral sequences correlates inversely with progesterone receptor expression (6.8% versus 20.3%; P=0.03) and HER2 overexpression (3.1% versus 17.7%; P=0.04). This present study confirms the presence of MMTV-like env sequences in BC in Tunisian women but describes an important decrease in the prevalence of the viral sequences compared with previous studies. This reduction may be due to some changes in the virological characteristics or exposure to the virus.

Prognostic significance of aberrant promoter hypermethylation of CpG islands in patients with diffuse large B-cell lymphomas.

BACKGROUND: Diffuse large B-cell lymphoma (DLBCL) exhibits heterogeneous clinical features and a marked variable response to treatment. PATIENTS AND METHODS: We investigated the prognostic significance of the methylation status of DAPK, GSTP1, P14, P15, P16, P33, RB1, SHP1, CDH1, APC, BLU, VHL, TIMP3, and RASSF1A genes in 46 DLBCL specimens from Tunisian patients. Methylation status of each gene was correlated with clinicopathological parameters including the International Prognostic Index (IPI), the germinal center immunophenotype, and response to treatment and survival. Overall survival (OS) and disease-free survival (DFS) rates were calculated by the Kaplan-Meier method and differences were compared with the log-rank test. RESULTS: Hypermethylation of SHP1 was associated with elevated lactate dehydrogenase level (P = 0.031). P16 and VHL were frequently hypermethylated in patients with high IPI scores (P = 0.006 and 0.004) and a performance status of two or more (P = 0.007 and 0.047). In addition, hypermethylation of P16 was significantly associated with advanced clinical stages and B symptoms (P = 0.041 and 0.012). Interestingly, hypermethylation of DAPK was significantly correlated with resistance to treatment (P = 0.023). With regard to survival rates, promoter hypermethylation of DAPK, P16, and VHL were significantly associated with shortened OS (P = 0.003, 0.001, and 0.017, respectively) and DFS (P = 0.006, 0.003, and 0.046, respectively). In multivariate analysis, hypermethylation of DAPK remains an independent prognostic factor in predicting shortened OS (P = 0.001) and DFS (P = 0.024), as well as the IPI and the germinal center status. CONCLUSIONS: This study demonstrates that DLBCLs with hypermethylated P16, VHL, DAPK, and SHP1 commonly show a biologically aggressive phenotype and worse prognosis. Interestingly, hypermethylation of DAPK was found to be an independent prognostic factor that may be used in conjunction with the conventional prognostic factors such as the IPI and the germinal center status.

Granuloma multiforme: first case report in Tunisia.

Granuloma multiforme is a rare granulomatous skin disease, usually reported in sub-Saharan African countries. The exact aetiology of granuloma multiforme is still unknown. We report the case of a patient who presented clinical and histopathological features of granuloma multiforme that can be considered the first described case in Tunisia.

[Salivary duct carcinoma]. / Carcinome des canaux salivaires.

INTRODUCTION: Salivary duct carcinoma is a very rare tumor of the salivary glands that has been principally reported in the parotid gland. It displays histologically a striking similarity to ductal carcinoma of the breast. OBSERVATION: We report the case of a salivary duct carcinoma arising in the parotid gland of a 62 year-old man. The fact that the tumor was confined to the parotid gland has allowed a radical treatment. After two year follow-up, there was no evidence of recurrence. DISCUSSION: This tumor is characterized clinically by local aggressiveness, frequent metastatic evolution and poor prognosis. This case report illustrates both the clinicopathological features and the prognostic factors of this entity.

[Prevalence of Epstein-Barr virus in Sjögren's syndrome in Tunisia]. / Prévalence du virus d'Epstein-Barr dans le syndrome de Sjögren en Tunisie.

PURPOSE: The cause of Sjögren's syndrome is unclear. Several studies suggested the role of Epstein-Barr virus (EBV) in the pathogenesis of this syndrome, but this always remains a subject of numerous controversies. The purpose of this study was to evaluate the prevalence of EBV in Sjögren's syndrome in Tunisia. METHODS: A series of 31 paraffin-embedded biopsies of salivary glands from patients with Sjögren's syndrome were studied in comparison with 19 control glands. EBV was investigated by PCR, EBERs in situ hybridization and by immunohistochemistry for the detection of LMP1, EBNA2 and ZEBRA. RESULTS: EBV DNA was detected by PCR in 3 of 22 PCR beta-globin positive Sjögren's syndrome cases (13.6%) and in 2 of 17 PCR beta-globin positive control glands (11.7%); in situ hybridization positivity was noted in rare lymphocytes in the 3 EBV positive cases of Sjögren's syndrome, but not in control glands; immunohistochemical study was negative in all cases. CONCLUSION: EBV infection does not appear to play a significant role in the pathogenesis of Sjögren's syndrome in Tunisia.

[Human subconjunctival dirofilariasis: two Tunisian case studies]. / La dirofilariose sous-conjonctivale humaine. A propos de deux cas tunisiens.

Two new cases of human subconjunctival dirofilariasis are reported in Tunisia. Dirofilaria immitis was identified in the first case and Dirofilaria repens in the second. Diagnosis was made by histological examination of excised nodules. We note that the patients are young (8 and 10 years, respectively) and that cases of human dirofilariasis caused by Dirofilaria immitis are present in Tunisia.