Computerized image cytometric analysis of: KI-67 and BCL-2 markers in: benign endometrial hyperplasia and endometrial carcinoma

The normal endometrium is a tissue subjected to rapid cycling proliferation and differentiation followed by breakdown under hormonal influences .Endometrial hyperplasia [EH], which is a frequent disorder seen in majority of endometrial biopsy samples, embraces a group of lesions with different biological potential with tendency to progress into endometrial adenocarcinoma [EC]. The reproducibility of the histopathologic diagnosis of endometrial hyperplasia is poor as regarding intra and inter observer errors as well as the over and under diagnosis of malignant changes. Strict criteria, good sampling and use of additional ancillary molecular techniques are expected to improve the sensitivity and accuracy of this diagnosis. To evaluate the immnohistochemical expression of Ki-67 and Bcl-2 in various types of endometrial hyperplasia versus endometrioid variant of endometrial adenocarcinoma. 2-To correlate the immunohistochemical expression with grade and stage of EC and 3-To evaluate the prognostic value of morphometric D-score in prediction of endometrial hyperplasia- cancer progression. Paraffin embedded-tissue specimens obtained from 57 female patients were studied. Cases were categorized into the following groups: Control group [5 cases of normal proliferative endometrium], fifteen cases of various degrees of hyperplastic lesions including 10 cases of [EH] and 5 cases of intra epithelial neoplasia [EIN]. Fifteen cases of combined lesions [EH+EIN+grade 1EC] and lastly 22 cases of various grades of endometrioid variant of EC. All 57 studied cases were subjected to:Immunohistochemical staining for Ki-67 and Bcl-2 markers. The percent of positivity was calculated using computerized image cell analysis. In addition computerized morphometric analysis for D-score calculation were performed on standard diagnostic hematoxylin and eosin [H and E] stained sections of all cases of endometrial hyperplasia. Normal proliferative endometrim [5 cases, 100%] expressed high score of both Ki-67 and Bcl-2 markers. Also, in the hyperplastic group; the peak immunoreactivity for Ki-67 was seen in the EIN subgroup [5/5,100%]. In contrast, Ki-67 immunoreactivity was seen in only 80% of EH subgroup. However, the difference was statistically insignificant [p>0.05]. KI index was higher in carcinoma compared to EH. The difference between both groups was statistically significant [p=0.001]. Bcl-2 immunopositivity was seen in three cases of EIN [60%] and eight cases of EH cases [80%].Endometrial carcinoma showed Bcl-2 expression in [11 out of 22 cases50%]. Bcl-2 was expressed in the majority of cases of EH [8 cases out of 10 cases 80%] and showed uniform glandular positivity, but it was diminished in EIN subgroup [2 out of 5 cases, 60%]. Ki-67 was expressed in the majority of endometrial adenocarcinoma [20/22 cases, 90.9%]. Bcl-2 expression was decreased as the grade increased but this relation was statistically insignificant [p=>0.05]. Ki-67 and Bcl-2 scores were found to be negatively correlated in the same histologic group and this inverse relation was statistically significant [p=0.00]. Finally, D-score calculation revealed that: D-score value was more than 1 in cases of EH and less than 1 in EIN cases and the cut off value was 2.33. Normal proliferative endometrium expressed high levels of both Ki-67 and Bcl-2. It was shown also, that Ki-67 was sequentially increased from EH through EIN to EC. In contrast, Bcl-2 positivity was decreased significantly in cases of EC mainly the high grades. Such inverse relation between both markers [Ki-67and Bcl-2] was proved to be statistically significant. The imbalance between proliferation and apoptosis may be an important factor in the development of different endometrial lesions; benign and malignant. Precancerous endometrial lesions [EIN] could be identified, by using morphometric analysis for D-score calculation based on combination of architecture and cytological features with consideration to volume percentage stroma parameter

use of ELISA and immunohistochemistry techniques for detection of toxoplasma gondii antigen in tissues of experimentally infected mice

The capability of double antibody sandwich enzyme-linkad immunosorbent assay [ELISA] for detecting antigens of Toxoplasma gondii [T. gondii] in different mice tissue specimens was evaluated in comparison to the immunohistochemistry [IHC] technique. Results proved that tissue antigens were detectable in liver, kidney and mesenteric lymph node [LN] specimens by both methods from the second day of infection, with statistically significant increase in its amount in all organs throughout the period of the study. Using ELISA technique, the highest antigen level was recorded on the second day [0.120 +/- 0.0015] and the fourth day [0.147 +/- 0.0034] of infection in LN specimens, while, the liver showed the highest antigen level at the sixth day post infection [PI][0.165 +/- 0.0066]. On the other hand, using the IHC technique, the highest number of tachyzoites was recorded in LN sections in all studied durations, the second, the fourth and the sixth days PI [1.1 +/- 0.875, 1.6 +/- 1.173 and 3.1 +/- 1.370 respectively]. Thus, sandwich ELISA technique might offer a valuable aid for rapid diagnosis of acute toxoplasmosis in human tissues, and it has proved to be more accurate than IHC technique, since its results was coincided with the pathogenesis of the disease.

Correlation between extent of tumor vascularization, prognosis and hematogenous metastases in gastric carcinoma using vascular endothelial growth factor [VEGF] and Von-Willebrand factor

Angiogenesis has a crucial importance for tumor growth and development of metastasis. This study aimed to evaluate angiogenesis. Microvessel density [MVD] was counted by staining endothelial cells immunohistochemically using anti von Willebrand vWF [factor VIII] antibody as well as vascular endothelial growth factor [VEGF], a known endothelial proliferative and mitogenic marker. Thirty prospective cases [24 cases of adenocarcinoma and 6 of diffuse infiltrating tumors] were enrolled in this study. Then, four cases were excluded from the study because they had inoperable tumors. Thus, a total of 26 patients was studied histologically, immunohistochemically and followed up for a period ranging from 6-30 months [median 12 months]. The immunohistochemical results of these biomarkers were correlated with standard prognostic factors [tumor grade, stage and lymph node metastasis] as well as overall survival period. It was found that MVD was significantly increased in deep advanced tumors and in the presence of nodal metastasis. However, VEGF positivity was demonstrated in only 46% of the cases and was not correlated with the clinicopathologic parameters. The MVD for cases that developed hematogenous metastasis was significantly higher than those having non metastatic tumors. Also, MVD was correlated with a relatively higher survival rates in favor of the hypovascular group