Cytogenetic investigations and molecular analysis of the Y chromosome by the polymerase chain reaction amplification of sequence-tagged sites (STS-PCR) technique were performed in 126 patients ...affected by idiopathic oligo-azoospermia following accurate selection of cases. Seventeen patients evidenced an abnormal karyotype. Fourteen patients with a normal karyotype had microdeletions of the Y chromosome within interval 6. In azoospermic patients microdeletions were scattered along different subintervals, while in oligozoospermic patients they were clustered in subinterval 6E. The size of the deletion was not apparently related to the severity of the disease. These results suggest that cytogenetic analysis and the STS-PCR technique can detect a genetic cause of infertility in about one-quarter of patients with idiopathic oligo-azoospermia.
The differential diagnosis based on morphology between neuroblastoma (NB) and peripheral neuroepithelioma (PN) is difficult, since these tumors share many architectural and cytological features. In ...this study, a morphometrical approach to this diagnosis is made by using nuclear (shape factors) and tissue (volume density of nuclei and stroma) parameters. Quantitative morphological analysis adds important information, which when used with clinical and biochemical data facilitates the distinction. In the majority of cases, nuclei of PN are significantly less round than those of NB and their profile is much more irregular. The density of neoplastic nuclei is significantly higher in PN. However, in a certain number of cases, even the morphometrical study confirms how difficult it may be to differentiate these two neoplastic entities, since the values of parameters are largely overlapping. This suggests the existence of a "continuum" of changing features between NB and PN, which may substantiate the hypothesis of a common histogenesis.
The difficult differential diagnosis between the diffuse variants of cutaneous lymphoid hyperplasia (CLH; synonym; pseudolymphoma) and malignant follicular center cell lymphomas (FCCL) often requires ...a multidisciplinary approach. Eighteen CLH and 11 FCCL, diagnosed by conventional histology and immunophenotyping and subsequently examined with a polymerase chain reaction to show clonal immunoglobulin heavy-chain gene rearrangements, were subjected to a novel type of automated nuclear image analysis. Of all nuclear parameters tested in azure A-stained semithin sections, the mean nuclear profile area (TN) of lymphoid cells was the best criterion to distinguish between CLH and FCCL (p = 9 x 10(-6)). Additional distinctive features, in the order of decreasing significance, were the SD of TN; all chromatin textural parameters combined; and the light and the dark fractions of the central nuclear profile areas. Parameters related to the chromatin pattern were independent of nuclear profile size in FCCL, but not in CLH. Two lesions registered as CLH displayed the nuclear characteristics favoring this diagnosis, but showed B-cell monoclonality at the DNA level. In conclusion, computerized nuclear image analysis is a helpful additional diagnostic tool in the evaluation of diffuse CLH and cutaneous FCCL.
Type I and type III intestinal metaplasia in gastric mucosa have been examined using morphometric methods. Tissue (volume per cent gland, lumen, epithelium, goblet cell vacuoles) and nuclear ...parameters (area, with related standard deviation, and form factors) were used as indicators of gland crowding, nuclear-cytoplasmic ratio, nuclear atypia, and pleomorphism. In type III intestinal metaplasia, there is significantly (i) greater nuclear pleomorphism, (ii) a higher nuclear-cytoplasmic ratio, and (iii) smaller and less numerous goblet cell vacuoles in both the upper and the lower parts of the crypts. These two parameters have significantly higher values in the lower than in the upper part of individual crypts. No cell population with large pleomorphic nuclei characterized type III metaplasia, though there was more variation in nuclear size.
Previous studies have shown that oestrogen receptor content in breast cancer was correlated with qualitative and also, more strongly, with quantitative nuclear features in tissue sections. However, ...even with the better reproducible quantitative microscopical assessments, the variance in the correlation with oestrogen receptor was considerable. This might be due to the implicit problems of oestrogen receptor determination with the biochemical assay. Therefore, receptor content was studied using monoclonal antibodies in 50 consecutive invasive ductal breast cancers. Oestrogen receptor status was compared with qualitative features and with the mean and standard deviation of the nuclear area, morphometrically evaluated on immunostained and adjacent haematoxylin and eosin stained sections. In agreement with earlier observations, nearly all tumours with prominent elastosis were oestrogen receptor positive; but a minority of negative cases also showed elastosis. The correlation between the other qualitative features and receptor status was weak. A significant inverse correlation (P less than 0.001) existed between the receptor status and the mean and standard deviation of the nuclear area. Even with the highly reproducible morphometrical analysis, correlation between nuclear oestrogen receptor content and quantitative nuclear features was relatively weak. This might indicate that receptor status and nuclear morphometric features reflect different biological characteristics of breast cancers.
Abstract
Aim: HER2 is a key genetic driver in gastric cancer development. The anti-HER2 antibody, trastuzumab, in combination with standard chemotherapy,increases survival in HER2 positive gastric ...cancer. However it is still unclear why HER2 overexpressing tumours are initially and subsequently nonresponsive to trastuzumab treatment. Our aim is to investigate the possible mechanisms involved in trastuzumab resistance.
Methods: A panel of human gastric and oesophago-gastric cancer cell lines(NCI-N87, KATO III, OE 19) were screened for EGFR, HER2, HER3, MAPK, PI3K/Akt/mTOR and 4EBP-1 expressions by Western Blot analysis. Cells were treated with increasing concentrations of selective inhibitors such as trastuzumab (HER2 inhibitor), erlotinib (EGFR inhibitor), MEK inhibitor (BAY-86-9766) or PIK3CA/mTOR inhibitor (GDC-0980) as a single agent and/or in combination. MTT analyses were performed to evaluate the inhibitory effect of each compound on cell growth.
Results: In all cancer cells EGFR, HER2, MAPK, AKT and 4EBP1 were expressed at different degree. Although high HER2 expression levels, trastuzumab caused growth inhibition only in OE19 cancer cell line with no activity showed in KATO III and NCI-N87, suggesting a trastuzumab intrinsic resistance. The combination of MEK and PIK3CA/mTOR inhibitors in cancer cell lines was able to inhibit cell growth of NCI-N87, KATOIII and OE19 with an IC50 of 0.25 µM, 0.01 µM and 0.25 µM, respectively. The role of such therapeutic doublet in achieving cell growth inhibition was accompanied by significant reduction in the expression of AKT, pAKT, MAP, pMAP, 4EBP1 and p4EBP1 compared to control and trastuzumab treatment as revealed by Western Blot analysis.
Conclusions: The results of our in vitro study suggest that in HER2 trastuzumb resistant gastric cancer cell lines the double blockade of MAP and PI3K/Akt/mTOR pathways is able to induce significant growth inhibition suggesting a relevant crosstalk between the two pathways in this setting. On the basis of these preclinical data, new potential approaches could be explored in gastric cancer HER2 patients not responding to trastuzumab.
Disclosure: All authors have declared no conflicts of interest.
Abstract
Background: The receptor tyrosin kinase AXL was identified as a key regulator of CRCcells migration and invasion, targeting AXL represents a potential approach in CRC treatment. Our aim was ...to assess AXL and GAS6 expression on tumor samples derived form patients with CRC.
Methods: From a cohort of 123 CRC patients (from 2003 to 2011) specimens from paraffin blocks were obtained and tissue microarrays (TMA) were performed. AXL and GAS6 expression level were assessed by immunohistochemistry (IHC) and FISH. By using non parametric, univariate and multivariate analysis, AXL and GAS6 expression was correlated with survival and clinicopathologic features.
Results: AXL expression was negative in 49/123 (39,8%) cases and positive in 74/123 cases (60,2%); 25 (33,8%) defined as low expression(1-19% of stained cells) and 49 (66.2%) as high expression(≥20%). AXL staining mostly displayed membrane positivity, with a small number of cases with cytosolic staining and one nuclear staining.Tumoral stroma and non-tumoral adjacent tissues were negative, whereas 10% of cases had endothelial cells positivity. GAS6 expression was also assessed in a small cohort of 73 CRC specimens. GAS6 IHC expression was found in 26/73 (35,1%) defined as low (1-29%) expression, 47 (67,1%) defined as high (≥30%). GAS6 staining was iuxta-membrane, 10% of samples displayed stromal posivity, indicating that the ligand can also be provided by the tumor microenvironment. A significantly correlation between AXL and GAS6 expression level (Pearson correlation coefficient 0.6 with a p <0.0001) was found, suggesting the presence of autocrine stimulatory loops within the tumour itself . FISH analysis was performed on 49 tumour cases. AXL was amplified in 4 cases(8%) with a ratio ranging from 2.0 to 2.3. We further correlated AXL and GAS6 expression with clinicopathologic features. AXL showed no correlation with T, N, M, stage, grading, pathologic features. Only GAS6 was statistically significant with respect to lymph node involvement (P < .002) and clinical stage (P < .004).
Conclusion: AXLand GAS6 were frequently overexpressed in CRC tumor specimen with 8% of AXL gene amplification, to our knowledge this is a first report. However no AXL or GAS6 prognostic role derived from our data.
Disclosure: All authors have declared no conflicts of interest.
Cutaneous melanoma, stage I, from 35 survivors at 5 year follow-up and 16 non-survivors were studied. Mean nuclear area in the superficial layer was significantly larger than in the deep layer both ...in survivors and non-survivors, but the ratio between nuclear area in superficial and deep layers (so-called maturation index) did not differ between survivors and non-survivors. In comparison with the survivors, the mean nuclear area of non-survivors was significantly larger both in the superficial (51.1 microns2 vs 43.7 microns2, p less than 0.01) and deep (42.9 microns2 vs 36.4 microns2, p less than 0.05) layer. This points to a general increase in nuclear areas in metastasizing tumors. Furthermore, the coefficient of variation of nuclear area (standard deviation/mean) x 100 was not different between survivors and non-survivors, either in the superficial or in the deep layer. Inspection of histograms of areas of 1000-2000 nuclei per case in 20 random cases (10 survivors and 10 non-survivors) showed a homogeneous increase in nuclear area in non-survivors. None of the histograms revealed a cell clone with especially large nuclei. These data show that the increased mean nuclear area in non-survivors is due to a homogeneous increase of all nuclei throughout the tumor and not to a special cell clone with large nuclei within nuclei of otherwise normal size. The difference in mean nuclear area in superficial and deep layers indicates that careful selection of nuclei in either of these layers is essential to obtain reproducible and comparable results with interactive morphometry.
Morphometrical measurements of nuclear area and form factors are carried out by means of a semiautomated image analyzer on 90 cases of ductal breast cancers and on lymph node tumor deposits. The ...value of the mean nuclear area in lymph node metastases is significantly higher than in primary tumors regardless of the size of the tumor. Since the value of mean nuclear area is also significantly higher in primary tumors with lymph node involvement than in those without lymph node involvement it is possible to assume that primary tumors with large-nucleus areas are more liable to invade lymph nodes. No significant differences are noted as far as the form factors are concerned.
PDF
