Prenatal and postnatal findings in a 10.6 Mb interstitial deletion at 10p11.22-p12.31.

Interstitial deletion of the proximal short arm of chromosome 10 represents a rare genetic alteration. Literature review revealed that only 10 postnatal diagnosed clinical cases with deletions overlapping 10p12p11 were published until present. We report the first prenatal diagnosis and postnatal findings in a male fetus with a 10.6 Mb interstitial deletion of the short arm of chromosome 10 (10p11.22-p12.31).

IL-1RN +2018T>C polymorphism is correlated with colorectal cancer.

Epidemiological and experimental evidence indicates chronic inflammation as a risk factor for colorectal cancer. We investigated whether IL-1B -511C>T (rs16944), IL-1B +3954C>T (rs1143634) and IL1-RN +2018T>C (rs419598) cytokine polymorphisms are correlated with colorectal cancer. Blood samples were obtained from 377 Romanian subjects: 144 patients with sporadic colorectal cancer and 233 healthy controls. Polymorphisms were analyzed by allelic discrimination TaqMan PCR assays with specific probes. The results of our study showed that IL-1RN +2018T>C polymorphism is associated with colorectal cancer. We found that there was a significant difference in the frequency of CC genotype between patients with colorectal cancer and the control group (OR 2.42, 95 % CI: 1.06-5.53, p = 0,034) when TT genotype was used as reference. Furthermore, in a stratified analysis, a positive association was found only for IL-1RN +2018CC genotype, that was limited to early I and II stages (OR 2.72, 95 % CI: 1.05-7.03, p = 0,033). We did not find any association between any of the IL-1B polymorphisms and colorectal cancer. In conclusion this study found that IL-1RN +2018T>C polymorphism is associated with colorectal cancer, mainly for localized disease.

Recurrent partial hydatidiform mole, with a first twin pregnancy, after treatment with clomiphene citrate.

We present a patient, treated for 3 months with clomiphen citrate after 5 years of infertility. This treatment resulted in a twin pregnancy, one degenerated into a partial hydatidiform mole and the other into a very early embryo death. The karyotype was a mosaic one: 63% of metaphases showed triploidy - 69 XXX and 37% diploidy - 46 XX. Despite all medical advice, she returned 8 months later with a new pregnancy, which proved to be a new partial hydatidiform mole, this time a single one. Karyotype was, also, a triploidy - 69 XXX. The genetic map of both genitors was performed, showing no aberrations. Unfortunately, the patient came back, once again, 5 months later, with a new positive pregnancy test. Ultrasonography revealed a new very early embryo death, the histopathological analysis establishing to be a single 'pure' stop in evolution of the pregnancy. As all the three pregnancies obtained after treatment with clomiphene were abnormal, two being partial hydatidiform moles and one being a premature miscarriage, without any genetic aberrations of the genitors, it seems very possible that clomiphene, apart from improving fertility, also increases the risk of abnormal ovum appearance.

The influence of irradiation on autophagy process in normal and malignant colorectal epithelia.

AIM: The authors assessed the influence of preoperative radiotherapy on autophagy process using a quantitative assessment of LC3 expression on both normal and tumoral colorectal tissues. MATERIALS AND METHODS: Normal and malignant tissue samples were taken from 50 patients that underwent surgery for colorectal adenocarcinoma of which 11 received preoperative radiotherapy. Tissue samples were included in paraffin and sections were immunomarked for LC3 expression. LC3 percentage was assessed with dedicated software on 10 randomly selected fields with 40× objective from both normal and malignant tissue samples of each patient. The resulting data were assessed and compared with a statistical apparatus. RESULTS: LC3 was overexpressed in tumoral tissue as compared with normal one. The LC3 percentage is different from person to person and the higher it is in normal epithelium, the higher will be in tumoral epithelium of the same person, regardless the irradiation. The LC3 expression levels are decreasing from tumoral non-irradiated epithelia to normal irradiated epithelia. LC3 expression in tumoral cells is granular, with particular perinuclear disposal and often "annular" pattern. CONCLUSIONS: The autophagy process has a basal level in the normal tissue, with interindividual variability. The autophagy process proved to be upregulated in the tumoral cells, with a particular morphological expression, namely the presence of cytoplasmic coarse granules disposed in an "annular" pattern. Preoperative radiotherapy is downregulating the autophagy process both in normal and tumoral tissue but to a lesser extent in the latter.

Paternity analysis in special fatherless cases without direct testing of alleged father.

The ability to establish the biological father, in which the father of a person is not available, named "reverse paternity determination", is based on the determination of STR alleles in mother and her child, other children and brothers of the alleged father, and deduction of genetic constitution of the father by the basis of genetic laws. Presented herein are two cases of reverse paternity determinations. Mother and parenthood DNA was isolated from 300 microl of fresh blood collected into tubes containing the EDTA, using Wizard Genomic DNA purification kit (Promega). DNA amplification was performed using GenePrint STR System (CTT, FFv, and Silver STR III multiplex) (Promega). Amplification results were read on 0.4 mm thick sequencing size polyacrilamide gel (4% for CTT and FFv, and 6% for Silver STR III). Gels were silver stained using Silver Sequence DNA Staining Reagents (Promega). Possible alleles of the alleged father for the loci CSF1PO, TPOX, TH01, F13A01, FESFPS, vWA, D16S539, D7S820, D13S317, LPL, F13B, and HPRTB were established by the selection from alleles determined in mother and parenthood, as to as obligate alleles with which the alleged father contributes to the genetic constitution of investigated child. Probability of paternity was calculated using population data on STR allele frequencies for Romanian population established by our laboratory. These are the first cases of litigious reverse paternity determinations in Romania.

VEGF-A and VEGF-B mRNA expression in gastro-oesophageal cancers.

INTRODUCTION: Angiogenesis is essential for the local growth, invasion and metastasis of the tumours. Vascular endothelial growth factors (VEGFs) play a crucial role in tumour angiogenesis. The aim of our study was to quantify the expression of several VEGF family molecules in human gastro-oesophageal cancers and to analyse possible correlations between genes expression and clinico-pathological features. MATERIALS AND METHODS: Gene expression was quantified in 43 gastro-oesophageal paired samples using qRT-PCR with TaqMan probes specific to VEGF-A, including soluble transcript variants and VEGF-B genes. RESULTS: VEGF-A, including the studied splice variants and VEGF-B mRNAs were expressed in both tumour and peritumour mucosa. The expression of VEGF-A and its isoforms was higher in tumour compared with paired peritumour mucosa, while no significant difference was observed in VEGF-B expression. VEGF-A expression tended to correlate with tumour invasion. CONCLUSION: VEGF-A has a tendency to over-express in gastro-oesophageal cancers, while VEGF-B does not seem involved in these tumours. Further studies are required to establish the utility of anti-VEGF-A therapy and to find biomarkers for pathogenesis or response to therapy in gastro-oesophageal tumours.

Expression of vascular endothelial growth factor and epidermal growth factor receptor in pancreatic ductal adenocarcinomas, neuroendocrine tumours and chronic pancreatitis.

OBJECTIVE: Angiogenesis is a crucial event for pancreatic carcinogenesis, and it also plays an important role in chronic pancreatitis. The aim of our study was to evaluate the mRNA expression of vascular endothelial growth factor (VEGF) and epidermal growth factor receptor (EGFR) in chronic inflammatory or malignant pancreatic pathology in order to elucidate the differences in expression patterns and potential clinical implications. METHODS: Thirty-five patients who had undergone endoscopic ultrasonography followed by endoscipic ultrasound-guided fine needle aspiration (EUS-FNA) of focal pancreatic masses were included in the study. VEGF and EGFR mRNA expression levels in the samples collected by EUS-FNA were analyzed using quantitative real-time polymerase chain reaction (PCR). RESULTS: VEGF expression was detected in all chronic pancreatitis and adenocarcinoma samples and in only 62.5% of pancreatic neuroendocrine tumors. EGFR expression was detected in only 40% of the chronic pancreatitis cases, 76.9% of adenocarcinomas and in 50% of pancreatic neuroendocrine tumors. Both VEGF and EGFR mRNA levels were significantly higher in pancreatic ductal adenocarcinoma than those in normal tissue. VEGF expression inversely correlated with pancreatic ductal adenocarcinoma size, while EGFR expression was related to local invasiveness of adenocarcinoma. CONCLUSION: Both VEGF and EGFR mRNA expression in EUS-FNA samples may be used as a diagnostic marker associated with invasiveness in patients with pancreatic adenocarcinoma.

MMR gene expression pattern in sporadic colorectal cancer.

BACKGROUND AND AIMS: Colorectal carcinoma is the second leading cause of death by cancer in Europe as its incidence increases with life span. Continuing research to detect new highly sensitive and specific noninvasive biomarkers is essential. The aim of this study was to compare 9 mismatching repair (MMR) genes activation levels in normal, polyp and malignant tissues in order to detect a MMR gene expression pattern in sporadic colorectal malignant pathology. METHODS: MMR mRNA levels were evaluated in tumor-normal tissue paired samples and polyps collected from 29 patients undergoing standard surgical procedures with curative intention. Real-Time quantitative Reverse Transcription PCR (qRT PCR) with TaqMan probes specific to ANKRD17, EXO1, MLH1, MLH3, MSH2, MSH3, MSH4, MSH5, MSH6 gene transcripts were used. RESULTS: The general tendency observed was a lower mRNA level of MMR genes in tumor samples compared with the normal tissue, with the exception of EXO1 gene. The number of patients that showed a higher expression of MMR genes in normal tissue was significantly greater than the number of patients that showed a higher expression inside the tumor (p=0.0024). ANKRD17 mRNA levels were higher in normal tissue than in tumor for 16 cases, by contrast with only 6 cases of higher mRNA levels in tumor. CONCLUSIONS: ANKRD17 mRNA appears to be the most sensitive target and may have a potential value as an additional marker for the existing multitarget assay panel for colorectal cancer detection.

Noninvasive detection through REMS-PCR technique of K-ras mutations in stool DNA of patients with colorectal cancer.

BACKGROUND AND AIMS: Tumor exfoliated cells that shed into stool are attractive targets for molecular screening and early detection of colon malignancies. Many studies have suggested that the detection of activated ras may have diagnostic or prognostic importance. The aim of this study was to establish the suitability for use in diagnostic laboratories of the noninvasive screening test of K-ras mutation determination in the stool and its routine prognostic value in colorectal cancer. METHODS: Paired stool and tissue specimens obtained after polypectomy and colorectal biopsy from 28 patients diagnosed solely by histopathological findings with primary colorectal carcinoma, were prospectively studied for K-ras codon 12 mutations by restriction endonuclease-mediated selective (REMS)-PCR. RESULTS: DNA was obtained in 28 of tissue samples (100%) and 26 of stool samples (92.8%). K-ras codon 12 mutation was seen in 14 (50.0%) paired stool and tissue samples. Mutation detection was possible in 1000-fold excess of wild-type sequence. These results may be important in the design of genetic screening programs, determination of prognosis, early detection and treatment for patients with colon malignancy. CONCLUSIONS: The sensitivity and specificity of K-ras determination on stool-derived DNA in patients with colorectal carcinoma, support the opportunity of a large-scale trial to validate its use as a screening test. REMS- PCR is not labor intensive, but a sensitive, rapid, and robust assay for the detection of point mutations, and was introduced by us in a routine diagnostic laboratory.