Histology and histopathology Vol.32, nº3 (2017)
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- PublicationOpen AccessDock3-NMDA receptor interaction as a target for glaucoma therapy(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Kimura, Atsuko; Namekata, Kazuhiko; Guo, Xiaoli; Harada, Chikako; Harada, Takayukiy. Glaucoma is a neurodegenerative disease of the eye and it is one of the major causes of blindness. Glaucoma is usually associated with elevated intraocular pressure (IOP) and the current therapy focuses on reduction of IOP. However, neuroprotective strategies could also be beneficial for treatment of glaucoma because the pathology of the disease involves retinal ganglion cell (RGC) death and damage to the optic nerve. Dedicator of cytokinesis 3 (Dock3) is an atypical guanine exchange factor (GEF) that belongs to a family of Dock proteins, Dock1-11. Dock3 exerts neuroprotective effects on the retina and optic nerve, and studies revealed that some of the Dock3-mediated effects are GEF-independent. One of these mechanisms is that Dock3 directly binds to the GluN2B subunit of the Nmethyl-D-aspartate (NMDA) receptor. Upon stimulation by NMDA or optic nerve crush, overexpression of Dock3 promotes internalization and degradation of the NMDA receptor in the retina in vivo. It is suggested that this process is mediated by inhibition of Fyn, a Src family tyrosine kinase. Reduction in NMDA receptor expression results in decreased excitotoxic damage and oxidative stress, thereby promoting RGC survival. In this review, we discuss the therapeutic potential of neuroprotection for glaucoma and the effects of Dock3 on NMDA receptors. We also discuss apoptosis signalregulating kinase 1 (ASK1), a member of mitogenactivated protein kinase kinase kinase that is a key regulator of cellular responses to oxidative stress, as an innovative therapeutic target for glaucoma.
- PublicationOpen AccessAn optimized xylene-free protein extraction method adapted to formalin-fixed paraffin embedded tissue sections for western blot analysis(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Mansour, Anthony G.; Abou Khalil, Pamela; Bejjani, Noha; Chatila, Rajaa; Dagher Hamalian, Carole; Faour, Wissam H.Deparaffinization of formalin-fixed paraffin embedded (FFPE) tissues with xylene currently remains a major challenge to the biomedical community. We developed an efficient xylene-free protocol to isolate proteins from archived FFPE human tissue sections. A total of 79 different types of FFPE tissue sections of 8 µm thickness were obtained from various archived FFPE specimens. Deparaffinization was conducted by gently washing each section with around 1 ml of hot distilled water (≈80°C). The deparaffinized tissues were homogenized in lysis buffer, and the isolated proteins were quantified and efficiently resolved using western blot analysis for the presence of Protein kinase B (PKB/AKT) and β-actin. Moreover, a significant amount of proteins was successfully isolated with an average of 2.31 µg/µl. The migration pattern of AKT and β-actin obtained from the specimens was similar to the positive control obtained from protein lysates prepared from in vitro cultured MDA231 cancer cell lines. AKT was successfully identified in all specimens, and β-actin protein was resolved with an efficiency higher than 80%. The entire extraction procedure requires only 20 minutes. This newly developed technique is an efficient, safe, cost-effective, and rapid method to isolate proteins from FFPE tissue sections adequate for molecular analysis.
- PublicationOpen AccessNeutrophil extracellular traps in tissue pathology(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Nakazawa, Daigo; Kumar, Santhosh V.; Desai, Jyaysi; Anders, Hans JoachimNeutrophil extracellular traps (NETs) are innate immune systems against invading pathogens. NETs are characterized as released DNA mixed with cytoplasmic antimicrobial proteins such as myeloperoxidase, proteinase3 and neutrophil elastase. While NETs are thought to have an important role in host defense, recent work has suggested that NETs contribute to tissue injury in non-infectious disease states. Uncontrolled NET formation in autoimmune diseases, metabolic disorders, cancers and thrombotic diseases can exacerbate a disease or even be a major initiator of tissue injury. But spotting NETs in tissues is not easy. Here we review the available histopathological evidence on the presence of NETs in a variety of diseases. We discuss technical difficulties and potential sources of misinterpretation while trying to detect NETs in tissue samples
- PublicationOpen AccessGrading lung neuroendocrine tumors: Controversies in search of a solution(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Pelosi, Giuseppe; Pattini, Linda; Morana, Giovanni; Fabbri, Alessandra; Faccinetto, Alex; Fazio, Nicola; Valeri, Barbara; Sonzogni, AngelicaBackground. Pathological grading of tumors is a way to measure biological aggressiveness. In lung neuroendocrine tumors (NET), grading is tautologically included into the current 2015 WHO histologic classification. Little is known, however, about alternative grading systems in lung NET. Methods. Through an extensive search of the English literature on lung NET (updated to April 2016), the following key questions were addressed: a) current concepts of grading; b) clinicians’ requests for grading; c) functional parameters for grading; d) Ki-67 labeling index (LI) for grading; e) towards an effective pathology grading system. Results. There is some room for inconsistency in the histologic classification of lung NET, likely due to the varying attribution of defining criteria. Innovative diffusion-weighted imaging upon magnetic resonance or molecular analysis could help separate indolent from aggressive lung NET, thus integrating a grading approach other than histology. Troubles in the clinical handling of metastatic or individual tumors when relying on morphology alone support the development of a lungspecific grading system for the more accurate prediction of prognosis and planning therapy in individual patients. To integrate the 2015 WHO classification using innovative grading based on Ki-67 LI, mitotic count and necrosis, a new proposal is emerging where three categories of lung NET are identified, namely Lu-NET G1, Lu-NET G2 and Lu-NET G3, which would allow tumors with similar behavior and therapy to be better handled according to their own biological potential. Conclusion. A new formulation of lung NET grading could have clinical relevance for the individual handling of patients. Key words:
- PublicationOpen AccessHuman trophoblast cell during first trimester after IVF-ET differs from natural conceived pregnancy in development and function(Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Yang, Rui; Liu, Ying Ying; Zhao, Liang; Wang, Ying; Li, Rong; Liu, Ping; Ma, Cai Hong; Chen, Xin Na; Qiao, JieObjective. To explore the differences of the trophoblast cell function in first trimester between natural pregnancy and pregnancy after IVF-ET therapy. Methods. 102 cases with twin to singleton fetal reduction after IVF-ET treatment from July 2010 to August 2013 in Peking University Third Hospital were involved in analysis, and eight specimens were obtained from this group. 10 natural-pregnancy cases undergoing artificial abortion with unwanted pregnancy were recruited as control. Semi-quantitative immunohistochemical method was used to detect the expression of EGFR, Bcl-2, tubulin-α, metallothionein and AFP in villi in both groups. Results. Of the 102 cases, 14 cases (13.73%) were aborted. Preterm birth occurred in seven cases (7.86%). Low birth weight occurred in three patients (3.37%), and extremely low birth weight occurred in four cases (4.49%). The expression of EGFR, tubulin-α, Bcl-2, and metallothionein in the IVF-ET group was significantly lower than that in the control group (P<0.05). However, AFP expression was significantly higher in IVF-ET group than in control group (P<0.05). In IVF-ET group, the miscarriage case had weaker EGFR, tubulin-α, and metallothionein expression than full-term pregnancy; the early preterm labor case had weaker Bcl-2, tubulin-α, and metallothionein expression; and velamentous cord insertion case had weaker tubulin-α expression. Conclusions. The trophoblast cell function of IVF-ET group in first trimester is different from control group in proliferation, invasion, apoptosis and vascular development, and optimal pregnancy outcome depends on the self-healing balance of trophoblast cells.