Whole exome sequencing identifies a mutation for a novel form of corneal intraepithelial dyskeratosis.

BACKGROUND: Corneal intraepithelial dyskeratosis is an extremely rare condition. The classical form, affecting Native American Haliwa-Saponi tribe members, is called hereditary benign intraepithelial dyskeratosis (HBID). Herein, we present a new form of corneal intraepithelial dyskeratosis for which we identified the causative gene by using deep sequencing technology. METHODS AND RESULTS: A seven member Caucasian French family with two corneal intraepithelial dyskeratosis affected individuals (6-year-old proband and his mother) was ascertained. The proband presented with bilateral complete corneal opacification and dyskeratosis. Palmoplantar hyperkeratosis and laryngeal dyskeratosis were associated with the phenotype. Histopathology studies of cornea and vocal cord biopsies showed dyskeratotic keratinisation. Quantitative PCR ruled out 4q35 duplication, classically described in HBID cases. Next generation sequencing with mean coverage of 50× using the Illumina Hi Seq and whole exome capture processing was performed. Sequence reads were aligned, and screened for single nucleotide variants and insertion/deletion calls. In-house pipeline filtering analyses and comparisons with available databases were performed. A novel missense mutation M77T was discovered for the gene NLRP1 which maps to chromosome 17p13.2. This was a de novo mutation in the proband's mother, following segregation in the family, and not found in 738 control DNA samples. NLRP1 expression was determined in adult corneal epithelium. The amino acid change was found to destabilise significantly the protein structure. CONCLUSIONS: We describe a new corneal intraepithelial dyskeratosis and how we identified its causative gene. The NLRP1 gene product is implicated in inflammation, autoimmune disorders, and caspase mediated apoptosis. NLRP1 polymorphisms are associated with various diseases.

[A letal case of gastro-intestinal basidiobolomycosis]. / Une forme gastro-intestinale de basidiobolomycose d'évolution fatale.

We report here the case of a 55-year-old man from Mali, who presented with abdominal pain. Radiological exploration revealed an ileo-colonic mass surrounding the appendix. A biopsy was taken and on histology, transmural granulomatous inflammation of numerous eosinophils, lymphocytes, plasmocytes and giant cells was seen. Tuberculosis was suspected clinically and an antibiotic treatment was initiated. Two months later, the patient died of septic complications. Basidiobolus ranarum was identified by PCR. Pathogens were retrospectively highlighted on biopsies. These elements were between 10 and 15 µm in diameter, occasionally pseudo-septated, and were surrounded by a thick eosinophilic cuff. The thick eosinophilic cuff was identified as the Splendore-Hoeppli phenomenon. Basidiobolomycosis is a well-known infection in the tropical areas. Basidiobolus sp., fungus of the order Entomophtorales are a known cause of chronic subcutaneous mycosis. Gastro-intestinal basidiobolomycosis is rare and presents considerable diagnostic difficulty. This infection needs to be diagnosed because surgical resection and prolonged antifungal treatment are curable in most cases.

Genotype-phenotype correlations of TGFBI p.Leu509Pro, p.Leu509Arg, p.Val613Gly, and the allelic association of p.Met502Val-p.Arg555Gln mutations.

PURPOSE: Investigate the genotype-phenotype correlations for five TGFBI (transforming growth factor, beta-induced) mutations including one novel pathogenic variant and one complex allele affecting the fourth FAS1 domain of keratoepithelin, and their potential effects on the protein's structure. METHODS: Three unrelated families were clinically diagnosed with lattice corneal dystrophy (CD) and one with an unclassified CD of Bowman's layer. Mutations in the TGFBI gene were detected by direct sequencing, and the functional impact of each variant was predicted using in silico algorithms. Corneal phenotypes, including histological examinations, were compared with the literature data. Furthermore, molecular modeling studies of these mutations were performed. RESULTS: Two distinct missense mutations affecting the same residue at position 509 of keratoepithelin: p.Leu509Pro (c.1526T>C) and p.Leu509Arg (c.1526T>G) were found to be associated with a lattice-type CD. The novel p.Val613Gly (c.1828T>G) TGFBI mutation was found in a sporadic case of an Algerian individual affected by lattice CD. Finally, the Bowman's layer CD was linked to the association in cis of the p.Met502Val and p.Arg555Gln variants, leading to the reclassification of this CD as atypical Thiel-Behnke CD. Structural modeling of these TGFBI mutations argues in favor of these mutations being responsible for instability and/or incorrect folding of keratoepithelin, predictions that are compatible with the clinical diagnoses. CONCLUSIONS: Description of a novel TGFBI mutation and a complex TGFBI allele further extends the mutational spectrum of TGFBI. Moreover, we show convincing evidence that TGFBI mutations affecting Leu509 are linked to the lattice phenotype in two unrelated French families, contrasting with findings previously reported. The p.Leu509Pro was reported to be associated with both amyloid and non-amyloid aggregates, whereas p.Leu509Arg has been described as being responsible for Epithelial Basement Membrane Dystrophy (EBMD).

Increased HCMV seroprevalence in patients with hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common primary liver cancer, usually arising after years of chronic liver inflammation that could result from viral infections such as hepatitis B virus (HBV) and hepatitic C virus (HCV) infections. Human cytomegalovirus (HCMV) infects primary human hepatocytes and remains an important cause of morbidity in immunocompromised persons where it may manifest as symptomatic end-organ disease including hepatitis. The goal of the present study was to determine a potential correlation between HCMV infection and the appearance of HCC. METHODS: First, we analyzed the seroprevalence of HCMV in a cohort of 11,318 patients hospitalized between 2003 and 2009 in different departments of a French University Hospital. Second, we studied HCMV seroprevalence in a cohort of 190 subjects who were stratified on the basis of age, gender, HCC, cirrhosis (Cir), and the exposition to hepatotropic viruses (HCV, HBV). We further determined whether HCMV DNA was present specifically in tumour area in liver biopsies from HCC-positive patients by using nested PCR. RESULTS: We found that the HCMV seroprevalence was high in the Hepatology department. The HCMV seroprevalence was significantly higher in patients infected with HCV and/or HBV than in patients who were not infected by those later viruses (76.2% versus 56.5%, p < 0.001). The HCMV seroprevalence was significantly higher in patients with HCC (74%) and lower in patients without HCC (54% for HCC-/Cir+ patients, 57% for HCC-/Cir- subjects). We observed a positive correlation between serum IL-6 levels and HCMV seroprevalence in cirrhotic patients, but not in HCC patients. Serum IL-6 levels correlated positively with C-reactive protein (CRP) levels. Preliminary histological studies from liver biopsies from HCC-positive patients highlighted that HCMV DNA can be detected in tumour area of some of the patients studied. CONCLUSIONS: Our results indicate that HCMV seroprevalence in patients with HCC is significantly higher than in patients without HCC, is positively correlated with serum IL-6 levels in cirrhotic patients, and is positively associated with the presence of other hepatotropic viruses such as HCV and HBV.

Detection of alpha- and beta-human papillomavirus (HPV) in cutaneous melanoma: a matched and controlled study using specific multiplex PCR combined with DNA microarray primer extension.

There are few contradictory studies investigating the involvement of HPV in melanoma. We designed a controlled study to evaluate the HPV DNA prevalence in melanoma. One hundred patients with cutaneous malignant melanoma diagnosed between 2002 and 2006 were included. Complementary wide excision (healthy skin) was performed in 85 patients and was used as internal control. After DNA extraction, 68 different HPV types were studied using a multiplex PCR combined with microarray primer extension. We did not observe any statistical significant difference in terms of HPV DNA prevalence in melanoma (38.8%) and in healthy skin from wide excision (42.4%). Twenty-one different HPV types were detected but only one type was present in the majority of our samples (80/85 melanoma vs 59/66 HS). The distribution of HPV genera and types was similar in melanoma and HS, and beta-HPV was predominant (30.6% and 31.8%). Among alpha-HPV (10.6%), high-risk mucosal HPV16 was predominant. Among beta-HPV, melanoma harboured significantly more type 22 than control normal skin from the same patients and significantly less type 21 than paired control normal skin. No correlation between clinical and pathological melanoma characteristics and HPV DNA prevalence was found. Our data do not support a role of HPV infection in melanocarcinogenesis, but confirm the previous data suggesting that HPV DNA is widely distributed among the population and that occult HPV infections are frequent. Furthermore, specific HPV types, such as a-HPV16 and beta-HPV species 2 may be involved in a sub-group of melanoma.

The expression of Twist has an impact on survival in human bladder cancer and is influenced by the smoking status.

OBJECTIVES: Twist is considered as transcription factor that regulates epithelial mesenchymal transition (EMT) by at least inhibition of E-cadherin expression. EMT is a key event in the tumor invasion process. The purpose of this study is to investigate the expression of Twist but also those of E- and N-cadherin in human primary bladder tumor and to evaluate its prognostic value. As smoking cigarettes is a strong bladder cancer risk factor, we tried to evaluate the impact of the tobacco status on these molecular abnormalities. MATERIALS AND METHODS: To delineate on the oncogenic role for Twist in human bladder cancer, we evaluated the E- and N-cadherin but also Twist expression (n = 70) by immunohistochemistry. We evaluated the prognostic value of these expressions. Moreover, we tried to correlate these protein expressions to the smoking status of the patients. Overall survival (OS) and progression-free survival (PFS) were evaluated using the Kaplan-Meier method, and multivariate analysis was performed using the Cox proportional hazard analysis. RESULTS: Of the 70 bladder tumors, 28 (40%) cases were positive for Twist expression, 16 (23%) cases were negative for E-cadherin expression, and 12 (17%) were positive for N-cadherin expression. When categorized into negative vs. positive expression, Twist was associated with the stage (P = 0.001), the grade (P < 0.001), the progression (P = 0.02), and the E-cadherin expression (P = 0.01). Moreover, positive Twist expression clearly predicted poorer PFS (P = 0.02). In the multivariate analysis, both positive Twist expression and loss of E-cadherin expression were independent prognostic factors for PFS (P = 0.046 and P = 0.001, respectively) and only loss of E-cadherin expression for the OS (P < 0.001). We also demonstrated that almost 60% (16/28) of patients with Twist-positive expression were current smokers at the time of the diagnosis, corroborating the fact that smoking modulates the expression of EMT markers including Twist. CONCLUSION: Positive Twist expression may be a useful prognostic marker for patients with bladder cancer. Its expression seems to be correlated to the tobacco status of the patients.

Human papillomavirus (HPV) genotype distribution in invasive cervical cancers in France: EDITH study.

Invasive cervical cancer (ICC) remains a significant cause of morbidity and mortality in France. Since human papillomavirus (HPV) is the necessary cause of ICC, the aim of this study was to assess the type-specific prevalence of HPV in ICC in France in order to locally evaluate the potential benefit of an HPV 16/18 L1 virus-like particles (VLP) vaccination. A total of 516 histological specimens collected in 15 centers were analyzed. Among them, 86% had a diagnosis of squamous cell carcinoma (SCC) whereas 14% were adenocarcinomas (ADC). HPV genotyping was performed using the INNO-LiPA assay allowing the specific detection of 24 HPV genotypes both high risk (HR) and low risk (LR). The overall HPV prevalence in ICC was 97%. The most prevalent genotypes were HPV 16 (73%) and HPV 18 (19%) followed by HPV 31 (7%), 33, 68, 45, 52 and 58 (4.1-2.3%). HPV 16 and/or 18 were associated with 82% of ICC, 10% being HPV 16 and 18 coinfections. While HPV 16 was the most prevalent type in both SCC (74%) and ADC (64%), HPV 18 was by far more prevalent in ADC (37%) compared to SCC (16%; p < 0.001). Multiple infections with at least two different HR HPV genotypes were observed in 22% of ICC. Given the high HPV 16/18 prevalence and taking into account possible production of crossneutralizing antibodies against other HPV types, HPV 16/18 L1 VLP vaccination would be expected to significantly reduce the burden of ICC in France.

Analysis of human papillomavirus type 16 (HPV16) DNA load and physical state for identification of HPV16-infected women with high-grade lesions or cervical carcinoma.

Integration of human papillomavirus (HPV) DNA into the host cell genome is a frequent event in cervical carcinogenesis, even though this phenomenon does not seem to be mandatory for cervical cancer development. Our objective was to describe the load and physical state of HPV type 16 (HPV16) DNA in a series of cervical samples representative of the natural history of cervical cancer. We used a combination of three real-time PCR assays targeting E6, E2, and albumin genes to calculate HPV16 load (E6 and albumin) and the E2/E6 ratio as a surrogate of integration. This method was applied to 173 HPV16-positive cervical samples. Results show that viral load increases with the lesion grade (from 102 HPV16 DNA copies per 10(3) cells in normal samples up to 56,354 copies per 10(3) cells in cancers), while E2/E6 ratio decreases (from 1 in normal samples down to 0.36 in cancers). We propose that, according to this technique, an HPV16 viral load of higher than 22,000 copies/10(3) cells or an E2/E6 ratio of lower than 0.50 allows the identification of women with prevalent high-grade lesions or worse with a high specificity. In conclusion, both viral load and E2/E6 ratio, used in combination with an appropriate cutoff value, are suitable to screen women with prevalent cervical intraepithelial neoplasia grade 2 or 3 or cancer. Therefore, these assays would be useful in addition to routine HPV testing to more accurately identify women with (pre)cancerous lesions.

N-cadherin as a novel prognostic marker of progression in superficial urothelial tumors.

PURPOSE: Loss of intercellular adhesion and increased cell motility promote tumor cell invasion and spreading. In bladder cancer, loss or reduced E-cadherin expression has been associated with poor survival, and aberrant expression of N-cadherin has been associated with the invasive phenotype of bladder carcinoma cells. The purpose of this study was to investigate whether N-cadherin expression was associated with the bladder tumor progression. EXPERIMENTAL DESIGN: E-cadherin and N-cadherin expression was evaluated by immunohistochemistry in 101 tumors (pT1 and pT2-T3) and by reverse transcription-PCR analysis and immunohistochemistry in 28 other fresh frozen tumors (pT(a), pT1, and pT2-T3). RESULTS: N-cadherin expression was absent in normal urothelium, appeared in stage pT1, and increased in pT2-pT3 tumors. In most cases, increased N-cadherin expression in invasive tumors was associated with loss of E-cadherin expression. Progression-free survival and multivariate analyses revealed that N-cadherin expression is an independent prognostic marker for pT1 tumor progression. Analysis of the 28 frozen tumors by immunohistochemistry and reverse transcription-PCR showed a good correlation between protein and gene expression in pT1 and pT2-T3 tumors. Interestingly, in pT(a) tumors, N-cadherin was not immunodetected, whereas mRNA was present in 50% of cases. CONCLUSION: Regulatory defects in the N-cadherin promoter, abnormalities at the translational, or protein processing levels could explain the discrepancies between protein and mRNA expression. Most importantly, this study identified N-cadherin as a novel prognostic marker of progression in superficial urothelial tumors. Clearly, N-cadherin acts in an invasive mode in bladder cancer, but whether it has a primary role in urothelial neoplastic progression has yet to be investigated.

Cervical intraepithelial neoplasia associated with epidermodysplasia verruciformis HPV in an HIV-infected patient: a manifestation of immune restoration syndrome.

We report a case of a cervical intraepithelial neoplasia associated with epidermodysplasia verruciformis human papillomavirus (HPV) type 5 and HPV type 16 in a human immunodeficiency virus-infected patient. Furthermore, epidermodysplasia verruciformis-like cutaneous eruptions after initiation of highly active antiretroviral therapy has never been described as a manifestation of an immune restoration syndrome.

Expression of E-cadherin and alpha-, beta-, gamma-catenins in patients with bladder cancer: identification of gamma-catenin as a new prognostic marker of neoplastic progression in T1 superficial urothelial tumors.

Loss of intercellular adhesion facilitates tumor invasion. To clarify the relation between altered expression of cell adhesion molecules and progression of T1 superficial bladder tumors, 101 cases (71 T1 tumors, 30 T2/T3 tumors) were examined immunohistochemically for E-cadherin and alpha-, beta-, and gamma-catenins. A highly significant correlation was observed between the decreased expression of all molecules and increased TNM stage (P < .001). Univariate analysis, performed in cases of T1 tumors, revealed association of abnormal E-cadherin with beta-catenin diminution. Survival curves were established with the Kaplan-Meier method and analyzed according to clinical and histopathologic parameters using the log-rank test. Cox multivariate analysis revealed only gamma-catenin as an independent predictor of progression-free survival in patients with stage T1 bladder urothelial tumors. The characterization of T1 tumors that will progress could lead to the identification of patients who might benefit from surgery to avoid vesical muscle invasion and, consequently, metastasis.

[Malignant rhabdoid tumor of the liver with spontaneous rupture: a case report]. / Tumeur rhabdoïde hépatique maligne avec rupture spontanée: à propos d'un cas.

We report the case of an 8-year-old child, presenting a rhabdoid tumor of the liver with spontaneous rupture, revealed by an intra-abdominal bleeding with rapidly fatal course. This clinical and pathological report raises the problem of the differential diagnosis of primary malignant hepatic tumors of the child with no alpha-foetoprotein increase.

Mesenchymal progenitor cells differentiate into an endothelial phenotype, enhance vascular density, and improve heart function in a rat cellular cardiomyoplasty model.

BACKGROUND: Cellular cardiomyoplasty is a promising approach to improve postinfarcted cardiac function. The differentiation pathways of engrafted mesenchymal progenitor cells (MPCs) and their effects on the left ventricular function in a rat myocardial infarct heart model were analyzed. METHODS AND RESULTS: A ligation model of left coronary artery of Lewis rats was used. MPCs were isolated by bone marrow cell adherence. Seven days after ligation, MPCs labeled with 4',6-diamidino-2'-phenylindole were injected into the infarcted myocardium (n=8). Culture medium was injected in the infarcted myocardium of control animals (n=8). Thirty days after implantation, immunofluorescence studies revealed some engrafted cells expressing a smooth muscle phenotype (alpha SM actin+), as similarly observed in culture. Other engrafted cells lost their smooth muscle phenotype and acquired an endothelial phenotype (CD31+). Furthermore, vessel density was augmented in the MPC group in comparison with the control group. After 30 days, echocardiography showed an improvement on left ventricular performance in the MPCs compared with the control group. CONCLUSIONS: In vivo administration of syngenic MPCs into a rat model of myocardial infarcted heart was safety demonstrated. Some engrafted cells appeared to differentiate into endothelial cells and loss their smooth muscle phenotype. MPC engraftment might to contribute to the improvement on the cardiac function in such a setting.

Detection of a mesenchymal tumor responsible for hypophosphatemic osteomalacia using FDG-PET.

We report a case of oncogenic osteomalacia (OO) in a 71-year-old man. The tumor, which was localized in the left lower mandible, was not found by CT, MRI, or 111-indium octreotide scintigraphy but was easily detected by FDG-PET. The use of this technique in OO has never been reported.

A transthyretin mutation (Tyr78Phe) associated with peripheral neuropathy, carpal tunnel syndrome and skin amyloidosis.

BACKGROUND: More than 80 transthyretin (TTR) mutations have been described, most associated with amyloidosis. Peripheral neuropathy is the most common clinical presentation in TTR amyloidosis although the carpal tunnel syndrome (CTS) may be the first symptom and skin can be involved, as transthyretin amyloidosis is a systemic disease. CASE REPORT: The 78 year-old proband, belonging to a French family of Italian origin, presented with a 5 year history of peripheral neuropathy in the lower extremities. However, 15 years earlier he had had surgery for bilateral CTS. Amyloidosis was diagnosed on salivary gland and skin biopsies. Immunohistochemistry on skin biopsy was positive using anti-TTR. The proband has 10 siblings, 5 have CTS. METHODS: SSCP and direct sequencing of exons 2, 3, and 4 of the TTR gene were done on DNA from the proband and his brother who had had CTS. To confirm the mutation a PCR-IMRA was done. RESULTS: SSCP analysis of TTR exons 2, 3, and 4 did not suggest a mutation. Sequence analysis of TTR exon 3 revealed heterozygosity in both subjects for a single basepair transversion from A to T in codon 78 (TAC-->TTC) indicating a tyrosine to phenylalanine change. The mutation was confirmed by PCR-IMRA. CONCLUSION: This TTR mutation (Tyr78Phe) is associated with peripheral neuropathy, carpal tunnel syndrome and skin amyloidosis. It is also associated with late onset of the disease.

Transthyretin mutation (TTRGly47Ala) associated with familial amyloid polyneuropathy in a French family.

A French family in which three individuals had familial amyloid polyneuropathy (FAP) was investigated. The proband presented cardiomyopathy with atrial arrhythmia and then developed axonal polyneuropathy, carpal tunnel syndrome, and sclerodactyly. Nucleotide sequencing of exons 2, 3 and 4 of the transthyretin (TTR) gene revealed heterozygosity for a single base change in the second position of codon 47. This G to C transversion predicts replacement of a glycine by an alanine at position 47 in the mature protein. This mutation (G47A) was previously identified in two different families of Italian origin both of which had FAP and cardiomyopathy. Here we report the first identification of this mutation in a non-Italian family.

Danger of systemic cyclosporine for corneal graft.

PURPOSE: To report a case of posttransplant lymphoproliferative disorder (PTLD) in a patient receiving oral cyclosporine (CS) for immunosuppression in a high-risk keratoplasty. METHODS: A systemic CS therapy was given to prevent graft rejection of a keratoplasty. Risk rejection was high in regard to a previous graft rejection and persistence of a corneal vascularization. One year after the keratoplasty, the patient developed a gastric Epstein-Barr virus (EBV)-induced B cell lymphoma. The outcome was favorable after chemotherapy. CONCLUSIONS: This unique case of lymphoma (PTLD) in the course of corneal graft management questions the indications and the follow-up of patients with CS therapy and raises the issue of topical CS treatment.

Avascular necrosis of the hallucal sesamoids update with reference to two case-reports.

We report two cases of nontraumatic metatarsal pain with sclerosis and fragmentation of the lateral sesamoid bone on roentgenographs and computed tomography images. One patient underwent magnetic resonance imaging (MRI), which showed low signal from the sesamoid bone. These imaging findings suggested osteonecrosis. Histology of the sesamoidectomy specimen confirmed this diagnosis in one patient. Avascular necrosis of the metatarsal sesamoid is an uncommon disorder. The suggestive roentgenographic and MRI findings rule out the other painful conditions of the sesamoid bone. The features are reviewed and the treatment options discussed.

[Extra neural perineurioma: an unusal renal tumor]. / Périneurome extra-neural: une tumeur rénale inhabituelle.

Neoplasia composed of perineurial cells include intraneural and extraneural perineurioma. The latter is a rare tumor often observed in cutaneous tissue. We report a case of extraneural perineurioma developed at the upper pole of the left kidney and found during the assessment of a repetitive urinary tract infection in a 26 years old man. The tumor was composed of spindle-shaped cells arranged in a storiform pattern. The immunohistochemical pattern was EMA+, PS100-.

[Neurectodermal differentiation of extraskeletal myxoid chondrosarcoma: a classical feature?]. / Différenciation neurectodermique des chondrosarcomes extrasquelettiques myxoïdes: un classique?

Extraskeletal myxoid chondrosarcoma (EMC) is a phenotypically and genotypically distinct entity with a protracted course. A documented case of an extraskeletal myxoid chondrosarcoma characterized by a t(9; 17) (q22; q11) translocation with a neuroendocrine and neural differentiation is reported.