Ochratoxin A as possible factor trigging autism and its male prevalence via epigenetic mechanism.

The role of dysbiosis causing leaky gut with xenobiotic production and absorption is increasingly demonstrated in autism spectrum disorder (ASD) pathogenesis. Among xenobiotics, we focused on ochratoxin A (one of the major food contaminating mycotoxin), that in vitro and in vivo exerts a male-specific neurotoxicity probably via microRNA modulation of a specific target gene. Among possible targets, we focused on neuroligin4X. Interestingly, this gene carries some single nucleotide polymorphisms (SNPs) already correlated with the disease and with illegitimate microRNA binding sites and, being located on X-chromosome, could explain the male prevalence. In conclusion, we propose a possible gene-environment interaction triggering ASD explaining the epigenetic neurotoxic mechanism activated by ochratoxin A in genetically predisposed children. This mechanism offers a clue for male prevalence of the disease and may have an important impact on prevention and cure of ASD.

A novel zinc finger gene is fused to EWS in small round cell tumor.

Ewing sarcoma family of tumors share recurrent translocations that fuse EWS from 22q12 to five different members of transcription factors namely FLI-1, ERG, ETV1, E1AF and FEV. Different classes of DNA binding proteins, ATF1, WT1 and CHOP are fused to EWS generating distinct tumor phenotypes: clear cell sarcoma, desmoplastic small round cell tumor, and myxoid liposarcoma, respectively. We have cloned a novel gene located at 22q12 fused to EWS by a submicroscopic inversion of 22q in a small round cell sarcoma showing a translocation (t(1;22)(p36.1;q12). The gene, designated ZSG (Zinc finger Sarcoma Gene), is a putative Cys2-His2 zinc finger protein which contains a POZ transcriptional repressor-like domain at the N-terminus. The rearrangement involves intron 8 of EWS and exon 1 of ZSG creating a chimeric sequence containing the transactivation domain of EWS fused to zinc finger domain of ZSG. This product lacks the transcriptional repressor domain at the N-terminus of ZSG. A rearrangement of the second ZSG allele was also found in tumor cells. This is the first example of an intra-chromosomal rearrangement of chromosome 22, undetectable by cytogenetics, activating EWS in soft tissue sarcoma.

Low grade fibromyxoid sarcoma. a further low-grade soft tissue malignancy characterized by a ring chromosome.

Supernumerary rings in the context of a simple karyotype characterize several low-grade malignant tumors of soft tissue and bone. Low-grade fibromyxoid sarcoma is an uncommon low-grade sarcoma, the cytogenetics of which has not yet been reported. Here we describe the first molecular-cytogenetic characterization of a pulmonary metastasis of low-grade fibromyxoid sarcoma. The histology of the primary and recurrent tumors was consistent with the diagnosis of low-grade fibromyxoid sarcoma of the usual type, whereas the pulmonary metastasis was of the "giant rosettes" variant. Cytogenetic analysis revealed a ring chromosome. Because gain of material of chromosomes 7 and 16 was detected by CGH, the ring chromosome is assumed to be composed of material from these respective chromosomes.

Morphologic-cytogenetic analysis of dedifferentiated liposarcomas with an extensive misleading leiomyosarcomatous component.

This report describes two cases of recurrent retroperitoneal dedifferentiated liposarcoma characterized by an extensive leiomyomatous component that prevented the correct diagnosis before the last recurrence. Strong immunoreactivity with smooth muscle and desmin antibodies and ultrastructural features consistent with leiomyosarcoma were observed in the spindle-cell and/or myxoid-like components in all four recurrences in case 1, and in the spindle-cell component of the primary tumor and the first recurrence in case 2. In case 1, the correct diagnosis was suggested by the cytogenetic evidence of ring markers, a hallmark of well-differentiated/dedifferentiated liposarcoma. In case 2, tumor type was yielded mainly by the morphology of the second recurrence, which consisted entirely of a well-differentiated liposarcoma, a sclerosing inflammatory variant, as confirmed by the karyotype. Reevaluation of the first two surgical specimens of each case revealed small areas consistent with well-differentiated liposarcoma that had been previously overlooked. Despite the smooth-muscle antigen profile, both cases retained an mdm2+/p53+/cdk4+ immunophenotype consistent with the genotype.

Limited role of TP53 and TP53-related genes in myxoid liposarcoma.

AIMS: Circumstantial evidence suggests that genetic changes may lead to tumor progression within the myxoid liposarcoma tumors (MLTs) carrying non-random chromosomal translocation t(12;16). METHODS: To address this subject an immunophenotypic analysis, applying antibodies against proteins encoded by TP53, MDM2 and CDK4 genes, complemented by molecular analysis of eight suitable cases, was performed on 104 consecutive cases. Chromosomal translocations were assessed either by cytogenetic analysis or by RT-PCR in 9 suitable cases and chimeric transcripts were found in all cases but two pleomorphic liposarcomas. RESULTS: Based on immunophenotyping and tumor site, the case material consisted of three groups. The first one was made up of 92 non-retroperitoneal cases carrying a null p53, mdm2, cdk4 immunophenotype, which remained unchanged over the time of recurrences and along the gamut of histologic subtypes. The second group was represented by five p53+, mdm2-, cdk4- non-retroperitoneal cases, 4 of which were further analysed by PCR-SSCP for p53 mutation. The immunophenotypic profile of these cases, complemented by the molecular findings, supported a role of TP53 in tumor progression in three high-grade MLTs. The third group, consisting of 7 retroperitoneal cases, showed a heterogeneous immunophenotype, sharing immunophenotypic and molecular features with the well-differentiated/evoluted (dedifferentiated) liposarcoma group. CONCLUSIONS: TP53 mutations seem to play a role in tumor progression in a few cases of MLTs (2.8%) showing more aggressive histologic characteristics. The unexpected finding that a number of retroperitoneal LMTs display the immunophenotypic profile of the well differentiated/evoluted (dedifferentiated) liposarcomas, deserves further investigation.

Genome Sequence of Rhodococcus opacus Strain R7, a Biodegrader of Mono- and Polycyclic Aromatic Hydrocarbons.

Rhodococcus opacus strain R7 (CIP107348) degrades several mono- and polycyclic aromatic hydrocarbons. Here, we present the high-quality draft genome sequence of strain R7, consisting of 10,118,052 bp, with a G+C content of 67.0%, 9,602 protein-coding genes, and 62 RNAs genes.

Myosin-binding protein C DNA variants in domestic cats (A31P, A74T, R820W) and their association with hypertrophic cardiomyopathy.

BACKGROUND: Two mutations in the MYBPC3 gene have been identified in Maine Coon (MCO) and Ragdoll (RD) cats with hypertrophic cardiomyopathy (HCM). OBJECTIVE: This study examined the frequency of these mutations and of the A74T polymorphism to describe their worldwide distribution and correlation with echocardiography. ANIMALS: 1855 cats representing 28 breeds and random-bred cats worldwide, of which 446 underwent echocardiographic examination. METHODS: This is a prospective cross-sectional study. Polymorphisms were genotyped by Illumina VeraCode GoldenGate or by direct sequencing. The disease status was defined by echocardiography according to established guidelines. Odds ratios for the joint probability of having HCM and the alleles were calculated by meta-analysis. Functional analysis was simulated. RESULTS: The MYBPC3 A31P and R820W were restricted to MCO and RD, respectively. Both purebred and random-bred cats had HCM and the incidence increased with age. The A74T polymorphism was not associated with any phenotype. HCM was most prevalent in MCO homozygote for the A31P mutation and the penetrance increased with age. The penetrance of the heterozygote genotype was lower (0.08) compared with the P/P genotype (0.58) in MCO. CONCLUSIONS AND CLINICAL IMPORTANCE: A31P mutation occurs frequently in MCO cats. The high incidence of HCM in homozygotes for the mutation supports the causal nature of the A31P mutation. Penetrance is incomplete for heterozygotes at A31P locus, at least at a young age. The A74T variant does not appear to be correlated with HCM.

Genome Sequence of Rhodococcus sp. Strain BCP1, a Biodegrader of Alkanes and Chlorinated Compounds.

Rhodococcus sp. strain BCP1 cometabolizes chlorinated compounds and mineralizes a broad range of alkanes, as it is highly tolerant to them. The high-quality draft genome sequence of Rhodococcus sp. strain BCP1, consisting of 6,231,823 bp, with a G+C content of 70.4%, 5,902 protein-coding genes, and 58 RNA genes, is presented here.

Multiplex polymerase chain reaction and ligation detection reaction/universal array technology for the traceability of genetically modified organisms in foods.

A multiplex polymerase chain reaction (PCR) system was developed for the simultaneous detection of target sequences in genetically modified soybean (Roundup Ready) and maize (MON810, Bt176, Bt11, and GA21). Primer pairs were designed to amplify the junction regions of the transgenic constructs analyzed and the endogenous genes of soybean (lectin) and maize (zein) were included as internal control targets to assess the efficiency of all reactions. This multiplex PCR has constituted the basis for an efficient platform for genetically modified organism traceability based on microarray technology. In particular, the ligation detection reaction combined to a universal array approach, using the multiplex PCR as target, was applied. High specificity and sensitivity were obtained.

T-banding pattern of bovine chromosomes and karyotype reconstitution with physically mapped cosmids.

Bovine T-banded chromosomes were identified by fluorescence in situ hybridization (FISH) using 27 cosmid probes as chromosome landmarks. Pairwise combinations of probes from chromosomes of markedly different size were cohybridized to metaphase spreads of T-banded chromosomes. This confirmed the association of banding patterns to individual chromosomes. The T-banding pattern appears to be related to R-bands: 80% of T-bands were in corresponding position to R-bands, but not always conserved in size. Band assignments placed 20 probes on similar bands in both patterns. The identification of T-banded chromosomes represents a first step toward the construction of a compositional map of the bovine genome and an additional tool for its study on a comparative basis.

Rapid differential diagnosis of myxoid liposarcoma by fluorescence in situ hybridisation on cytological preparations.

In two cases of suspected myxoid liposarcoma, where chromosomal metaphase preparations were not available, fluorescence in situ hybridisation was performed on interphase nuclei of cytological preparations for the detection of the specific translocation, t(12;16), characteristic of this tumour and of trisomy 8, which is the most frequent secondary chromosome aberration. Probes directed against chromosomes 12 and 16 and against the centromeres of chromosomes 12 and 8 were hybridised on cell brushings and cytocentrifuge preparations. The finding of three painting domains of both chromosomes 12 and 16 and of only two signals with the centromeric probe directed against chromosome 12, suggested the presence of t(12;16) in both cases. In one case trisomy 8 was inferred from the occurrence of three centromere 8 signals. This approach can be used to detect specific chromosomal abnormalities when an urgent differential diagnosis is requested or when chromosome preparations are not available, or both.

Detection of bladder cancer by multitarget multicolour FISH: comparative analysis on archival cytology and paraffin-embedded tissue.

Detection of bladder cancer by multitarget multicolour FISH: comparative analysis on archival cytology and paraffin-embedded tissue We have evaluated the possibility of using the same specimen for both cytological diagnosis and multitarget multicolour FISH (MtMcFISH) analysis in order to determine whether the routinely processed specimens used for diagnosis were also suitable for this ancillary procedure. For this purpose 18 positive samples (11 voided urine and seven bladder washings) were selected, together with a representative section of the corresponding immediately previous or subsequent histological specimens. Two negative cytology slides were added as negative controls. FISH analysis revealed a normal pattern for each probe in the two negative controls and an abnormal pattern in the 18 positive cases. In the latter the same FISH alterations were found in the cytology samples and in the corresponding histological sections, and superimposable cytological/histological features were observed in two cases where two different histology samples were analyzed. The results clearly show that MtMcFISH may be successfully applied to destained routinely processed cytology slides.

Esthesioneuroblastoma is not a member of the primitive peripheral neuroectodermal tumour-Ewing's group.

Esthesioneuroblastoma (ENB) is a rare, site-specific, locally aggressive neuronal malignancy so far thought to belong to primitive peripheral neuroectodermal tumour-Ewing's tumour (pPNETs-ETs). Its anatomical location, in addition to morphologic, immunophenotypic and ultrastructural features, suggests its origin in the neuronal or neuroendocrine cells of the olfactory epithelium. However, the cytogenetic and molecular data currently available appear controversial on the presence of the typical translocation t(11;22)(q24;q12) and of trisomy 8, chromosomal changes that characterize the tumours belonging to the pPNETs-ETs. Herein we have analysed five ENB tumour specimens for trisomy 8 by fluorescence in situ hybridization (FISH), for the presence of EWS gene rearrangements by FISH, reverse transcription polymerase chain reaction and Southern blot analyses, as well as for the expression of the Ewing sarcoma-associated MIC2 antigen by immunohistochemistry. Neither EWS/FLI-I, EWS/ERG and EWS/FEV fusion genes nor MIC2 expression were found in any tumour, whereas trisomy 8 was found in one case only. Moreover, DNA from three cases analysed by Southern blot did not show EWS gene rearrangements. Our results support the evidence that ENB is not a member of the pPNETs-ETs.

c-KIT and c-KIT ligand (SCF) in synovial sarcoma (SS): an mRNA expression analysis in 23 cases.

In a previous immunophenotypic molecular-based analysis it was shown that bcl2 over-expression characterizes the SS gene profile in addition to the non-random translocations. Here we show that the over-expression of an additional potentially antiapoptotic gene, the c-KIT gene, is associated with this tumour. Interestingly, whereas bcl2 over-expression appears to be restricted to the spindle cell tumoral component, c-kit mainly involves the epithelial component of biphasic SS. Twenty-three primary and metastatic samples from 21 patients were analysed by immunophenotyping (23/23), immunoprecipitations and Western blotting (3/23), and RT-PCR (23/23). Ten cases were biphasic and 13 monophasic in sub-type. Twelve, 10 and 1 case carried the SYT-SSX1, SYT-SSX2 and SYT-SSX4 fusion transcript, respectively. Co-presence of both c-Kit and SCF mRNA was observed in almost all cases (20/23), suggesting the occurrence of an autocrine loop. Immunophenotyping, confirmed by biochemical analyses, showed a modulation of c-Kit expression which was faint in the spindle and strong in the epithelial component, respectively. The study was complemented by c-Met/HGF receptor/ligand expression and c-Met protein analysis with results superimposable to those already reported. Since in each tumour, epithelial and spindle cell components harbour the same type of translocation t(X;18) the present findings suggest a shifting of the anti-apoptotic role from BCL2 to c-KIT gene during the transition from the uncommitted spindle to the differentiated epithelial cells.

Identification of a novel spliced variant of the SYT gene expressed in normal tissues and in synovial sarcoma.

Synovial sarcoma (SS) is cytogenetically characterized by the translocation t(X;18)(p11.2-q11.2) generating a fusion between the SYT gene on chromosome 18 and one member of the SSX family gene (SSX1; SSX2; SSX4) on chromosome X. Here, we report for the first time that 2 forms of SYT mRNA are present in both normal tissues and SSs. By amplifying the full-length SYT cDNA of two SSs, we detected 2 bands, here designated N-SYT and I-SYT. The latter, previously undescribed, contains an in-frame insertion of 93 bp. Its sequencing revealed a 100% homology with the mouse SYT gene. These two SYT forms were present, although in different amounts, in all human normal tissues examined, including kidney, stomach, lung, colon, liver and synovia. Coexistence of N-SYT and I-SYT (both fused with SSX) was detected in a series of 59 SSs (35 monophasic and 24 biphasic) and in a SS cell line. A preliminary analysis of the differential expression levels of N-SYT and I-SYT in SSs revealed that the latter was consistently overexpressed, suggesting a role in SS pathogenesis.

The expression of MDM2/CDK4 gene product in the differential diagnosis of well differentiated liposarcoma and large deep-seated lipoma.

Ordinary lipomas are cytogenetically characterized by a variety of balanced rearrangements involving chromosome segment 12q13-15, whereas well differentiated liposarcomas (WDL) show supernumerary ring and giant marker chromosomes, known to contain amplified 12q sequences. The tight correlation between the presence of ring chromosomes and both amplification and overexpression of MDM2 and CDK4 genes suggests the exploration of the possibility that immunocytochemistry (ICC) might assist in the differential diagnosis of lipoma-like well differentiated liposarcomas (LL-WDL) and large deep-seated lipomas (LDSL). For this purpose, 21 cases of the former and 19 cases of the latter tumours were analysed by ICC and, according to the availability of material, by molecular and cytogenetic approaches. All lipomas displayed a null MDM2/CDK4 phenotype, whereas all LL-WDL showed MDM2/CDK4 or CDK4 phenotypes. Southern blot analysis performed on 16 suitable cases, complemented by fluorescence in situ hybridization and classical cytogenetic analysis in 11 cases, was consistent with, and further supported the immunophenotyping data. In conclusion, MDM2/CDK4 product-based immunophenotyping appears to represent a valuable method for the categorization of arguable LDSL.