Immunoexpression of matrix metalloproteinases MMP-1, MMP-2, MMP-9 and MMP-14 in extragenital endometriosis and eutopic endometrium

Nicolae Testemitanu State University of Medicine and Pharmacy of the Republic of Moldova, Department of morphopathology, University of Medicine and Pharmacy, Craiova, Romania, Department of Pathology, Congresul consacrat aniversării a 75-a de la fondarea Universității de Stat de Medicină și Farmacie „Nicolae Testemițanu” din Republica Moldova, Ziua internațională a științei pentru pace și dezvoltareIntroduction: Endometriosis is a comon, benign, inflamatory, pathology, represented by the ectopic location of functional endometrial glands and stroma outside the uterine cavity [1, 2]. Matrix metalloproteinase (MMP) represents a large family zinc-dependent endopeptidases, involved in the degradation of the extracellular matrix in the process of endometrial cell implantation and are classified by their substrate specificity [7, 8]. Matrix metalloproteinases (MMPs) are essential in orchestrating proper physiological functioning of the endometrium; hence, alteration of MMP activities is considered as a critical factor for the development of endometriosis. MMPs are involved in the cellular event of epithelial-mesenchymal transition [10, 22]. Purpose: The aim of this study was to evaluate the immunohistochemical expression of matrixmetalloproteinases MMP1, MMP2, MMP9 and MMP14 in surgical excision specimens, collected from women with extragenital endometriosis compared to their expression in the normal endometrium. Material and methods: The patient group consist 42 women with endometriosis, age range of 21-63 years (median 40), diagnosed and surgically treated at Department of Surgery, Obstetrics and Gynecology from Gherghe Paladi Municipal Clinical Hospital, Sfantul Arhanghel Mihail Municipal Clinical Hospital, Chisinau, the Republic of Moldova and Emergency County Hospital, Craiova, Romania. Location included: the anterior abdominal wall after caesarean operation – 20, inguinal hernia – 7, umbilical hernia – 4, perineal region – 1, appendix – 4, colon – 5, and ileum – 1case. Results: The classical histopatological examination of the general expression of MMP-1, MMP-2, MMP-9 and MMP-14 revealed a diverse variability. For MMP-2, MMP-9, MMP-14 markers, the reaction positivity was variable not only from one case to another but also within each case, the latter being characterized by the identification of different areas within endometriosis. Immunohistochemical analysis has demonstrated the significant enhance of MMP-9 and MMP- 14 expressions in endometriosis and in endometrium. The distinctive feature of MMP-9 and MMP-14 expression in endometriosis was considerable increase of its activity precisely on the border of endometriotic lesion and the peritoneum. Conclusions: The MMP-9 and MMP-14 activity significant elevation, established on ectopic endometrium of women with endometriosis. Study of MMP-1, MMP-2, MMP-9 and MMP-14 activities in endometriotic lesions from women with endometriosis is perspective for further investigation in order to determine a possible role of matrix metalloproteinases in the development of invasiviness process in case of extragenital endometriosis

Immunoexpression of matrix metalloproteinases MMP-1, MMP-2, MMP-9 and MMP-14 in extragenital endometriosis and eutopic endometrium

Department of Morphopathology, Nicolae Testemitanu State University of Medicine and Pharmacy, Chisinau, the Republic of Moldova, Department of Pathology, University of Medicine and Pharmacy, Craiova, Romania, The 75th anniversary of Nicolae Testemitanu State University of Medicine and Pharmacy of the Republic of Moldova (1945-2020)Background: Matrix metalloproteinases are proteolytic enzymes responsible for the disorder of extracellular matrix modeling in endometriosis and their involvement in the invasion process. The aim of this study was to evaluate the immunohistochemical expression of matrix-metalloproteinases MMP-1, MMP-2, MMP-9 and MMP-14 in surgical excision specimens, collected from women with extragenital endometriosis compared to their expression in the normal endometrium. Material and methods: The study included 40 female patients diagnosed with extragenital endometriosis. The used methods consisted in processing the specimens by classical histological technique with paraffin inclusion and enzymatic immunohistochemical technique for the detection of metalloproteinases MMP-1, MMP-2, MMP-9 and MMP-14. Results: The expression of matrix metalloproteinases MMP-2, MMP-14 was significant in glandular cells from endometriotic lesions, while MMP-9 was evident in both stromal and glandular cells in these lesions. The expression MMP-1 was not present. Normal endometrial tissue showed high reactivity for MMP-14 and low reactivity for MMP-2 and MMP-9. Conclusions: This study reveals some aspects related to the morphological and clinical features of extragenital endometriosis with different locations and the correlation between the clinical evolution and some immunohistochemical markers with potential prognosis regarding the aggressiveness of such lesions

Fractal Analysis and the Diagnostic Usefulness of Silver Staining Nucleolar Organizer Regions in Prostate Adenocarcinoma

Pathological diagnosis of prostate adenocarcinoma often requires complementary methods. On prostate biopsy tissue from 39 patients including benign nodular hyperplasia (BNH), atypical adenomatous hyperplasia (AAH), and adenocarcinomas, we have performed combined histochemical-immunohistochemical stainings for argyrophilic nucleolar organizer regions (AgNORs) and glandular basal cells. After ascertaining the pathology, we have analyzed the number, roundness, area, and fractal dimension of individual AgNORs or of their skeleton-filtered maps. We have optimized here for the first time a combination of AgNOR morphological denominators that would reflect best the differences between these pathologies. The analysis of AgNORs’ roundness, averaged from large composite images, revealed clear-cut lower values in adenocarcinomas compared to benign and atypical lesions but with no differences between different Gleason scores. Fractal dimension (FD) of AgNOR silhouettes not only revealed significant lower values for global cancer images compared to AAH and BNH images, but was also able to differentiate between Gleason pattern 2 and Gleason patterns 3–5 adenocarcinomas. Plotting the frequency distribution of the FDs for different pathologies showed clear differences between all Gleason patterns and BNH. Together with existing morphological classifiers, AgNOR analysis might contribute to a faster and more reliable machine-assisted screening of prostatic adenocarcinoma, as an essential aid for pathologists

Antibody Elution Method for Multiple Immunohistochemistry on Primary Antibodies Raised in the Same Species and of the Same Subtype

Double or multiple antigen labeling in IHC classically relies on the existence of primary antibodies raised in different species or of different IgG isotypes to ensure the specific labeling with the secondary detection systems. However, suitable pairs of primary antibodies are not always available or the best choice (e.g., as diagnostic tools). During the last few years, several methods have been proposed to overcome this, but none of them offers the flexibility needed for reliable double or multiple enzymatic or fluorescent IHC. We present here a procedure that elutes the antibodies after a first round of immunolabeling, which, in combination with precipitation-based detection systems, allows multiple IHC rounds even for primary antibodies raised in the same species and IgG isotype. Compared with other proposed methods, this procedure ensures a reliable enzymatic or fluorescent staining without cross-reactivity and without loss of tissue antigenicity, thus offering a flexible tool for colocalization studies and pathological diagnosis. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials. (J Histochem Cytochem 57:567–575, 2009

Effects of Granulocyte-Colony Stimulating Factor After Stroke in Aged Rats

Popa-Wagner A, Stöcker K, Balseanu AT, et al. Effects of Granulocyte-Colony Stimulating Factor After Stroke in Aged Rats. Stroke. 2010;41(5):1027-1031.Background and Purpose—In aged humans, stroke is a major cause of disability for which no neuroprotective measures are available. Granulocyte-colony stimulating factor (G-CSF), a member of the cytokine family of growth factors, promotes brain neurogenesis and improves functional outcome after stroke in young animals. We tested the hypothesis that G-CSF provides a restorative therapeutic benefit in aged animals. Methods—Focal cerebral ischemia was produced by reversible occlusion of the right middle cerebral artery in 19- to 20-month-old male Sprague-Dawley rats. One hour after reperfusion, the aged rats were treated daily with 15 μg/kg G-CSF and for 15 days total. Rats were behaviorally tested and the brains removed for analysis at 28 days poststroke. Results—G-CSF treatment after stroke exerted a robust and sustained beneficial effect on survival rate and running function. Transient improvement after G-CSF treatment could be observed for coordinative motor function on the inclined plane test and for working memory in the radial-arm maze test. At the cellular level, G-CSF treatment increased the number of proliferating cells in the subventricular zone and dentate gyrus and also increased the number of newborn neurons in the subventricular zone ipsilateral to the lesion. Conclusions—These results suggest that G-CSF treatment in aged rats has a survival-enhancing capacity and a beneficial effect on functional outcome, most likely through supportive cellular processes such as neurogenesis

Inhibition of aquaporin-4 improves the outcome of ischaemic stroke and modulates brain paravascular drainage pathways

Aquaporin-4 (AQP4) is the most abundant water channel in the brain, and its inhibition before inducing focal ischemia, using the AQP4 inhibitor TGN-020, has been showed to reduce oedema in imaging studies. Here, we aimed to evaluate, for the first time, the histopathological effects of a single dose of TGN-020 administered after the occlusion of the medial cerebral artery (MCAO). On a rat model of non-reperfusion ischemia, we have assessed vascular densities, albumin extravasation, gliosis, and apoptosis at 3 and 7 days after MCAO. TGN-020 significantly reduced oedema, glial scar, albumin effusion, and apoptosis, at both 3 and 7 days after MCAO. The area of GFAP-positive gliotic rim decreased, and 3D fractal analysis of astrocytic processes revealed a less complex architecture, possibly indicating water accumulating in the cytoplasm. Evaluation of the blood vessels revealed thicker basement membranes colocalizing with exudated albumin in the treated animals, suggesting that inhibition of AQP4 blocks fluid flow towards the parenchyma in the paravascular drainage pathways of the interstitial fluid. These findings suggest that a single dose of an AQP4 inhibitor can reduce brain oedema, even if administered after the onset of ischemia, and AQP4 agonists/antagonists might be effective modulators of the paravascular drainage flow.</p