Uptake of Clostridium botulinum C3 Exoenzyme into Intact HT22 and J774A.1 Cells

The Clostridium botulinum C3 exoenzyme selectively ADP-ribosylates low molecular weight GTP-binding proteins RhoA, B and C. This covalent modification inhibits Rho signaling activity, resulting in distinct actin cytoskeleton changes. Although C3 exoenzyme has no binding, the translocation domain assures that C3 enters cells and acts intracellularly. C3 uptake is thought to occur due to the high concentration of the C3 enzyme. However, recent work indicates that C3 is selectively endocytosed, suggesting a specific endocytotic pathway, which is not yet understood. In this study, we show that the C3 exoenzyme binds to cell surfaces and is internalized in a time-dependent manner. We show that the intermediate filament, vimentin, is involved in C3 uptake, as indicated by the inhibition of C3 internalization by acrylamide, a known vimentin disruption agent. Inhibition of C3 internalization was not observed by chemical inhibitors, like bafilomycin A, methyl-β-cyclodextrin, nocodazole or latrunculin B. Furthermore, the internalization of C3 exoenzyme was markedly inhibited in dynasore-treated HT22 cells. Our results indicate that C3 internalization depends on vimentin and does not depend strictly on both clathrin and caveolae.     

Immunohistochemical investigation in odontogenic myxoma

Three odontogenic myxomas are described immunohistochemically by a panel of poly- and monoclonal antibodies to characterize this tumor type. Three types of odontogenic myxoma cells were discriminated: spindle cells, stellate cells and hyaline cells. Neoplastic cells of myxomas were positively stained for transferrin, ferritin, alpha-1-antichymotrypsin (alpha 1-ACT), alpha-1-antitrypsin (alpha 1-AT), S-100 protein and vimentin; however, neuron specific enolase (NSE), S-100 alpha subunit, S-100 beta subunit, Factor VIII-related antigen (FVIII-AG) and cytokeratin (CK1) were negative. Spindle cells were positive for transferrin, ferritin, alpha 1-ACT, alpha 1-AT, S-100 protein and vimentin. Stellate cells were strongly positive for transferrin, alpha 1-AT, S-100 protein and vimentin. Hyaline cells reacted with alpha 1-ACT and alpha 1-AT. Myxomatous matrix showed negative reaction for all the antibodies used. These results have confirmed that odontogenic myxoma is a tumor of a dual fibroblastic-histiocytic origin.     

Immunohistochemical study of tubular epithelial cells and vascular endothelial cells in glomerulonephritis

Background and aims: In order to assess the role played by tubular epithelial cells (TEC) and interstitial vascular endothelial cells (VEC) in interstitial fibrogenesis in human glomerulonephritis, we studied the expression of markers of activated fibroblasts (α-smooth muscle actin (αSMA) and vimentin (Vim)) and of the transforming growth factor β (TGFβ), at the level of these cells. Methods: We studied retrospectively 41 renal biopsies from patients with primary and secondary glomerulonephritis [24 males, 17 females, mean age 45.5?±?12.9?years]. Immunohistochemistry using monoclonal antibodies (SMA, Vim, TGFβ) was assessed using a semiquantitative score, that was correlated with biological and histological data (quantified using a scoring system in order to assess active-inflammatory and chronic–sclerotic/fibrotic lesions). Results: The presence of SMA and Vim as markers of myofibroblasts was found in TECs and VECs. TEC Vim expression correlated with interstitial Vim expression (r?=?0.38; p?=?0.008), interstitial infiltrate (r?=?0.31; p?=?0.027), interstitial fibrosis (R?=?0.25; p?=?0.042), GFR (r?=??0.35; p?=?0.016), SMA (r?=??0.42; p?=?0.015), TGFβ (r?=?0.25; p?=?0.046), and hemoglobin (r?=??0.55; p?r?=??0.32; p?=?0.023) and interstitial fibrosis (r?=??0.34; p?=?0.017). Conclusion: Our study reflects the complexity of the involvement of VEC and mainly of TEC in fibrosis. The expression of mesenchymal markers at the tubular cell level (especially Vim) correlates with histological interstitial changes, with the decrease of renal function and more strongly with anemia.     

Tissue-associated self-antigens containing exosomes: Role in allograft rejection

Exosomes are extracellular vesicles that express self-antigens (SAgs) and donor human leukocyte antigens. Tissue-specific exosomes can be detected in the circulation following lung, heart, kidney and islet cell transplantations. We collected serum samples from patients who had undergone lung (n?=?30), heart (n?=?8), or kidney (n?=?15) transplantations to isolate circulating exosomes. Exosome purity was analyzed by Western blot, using CD9 exosome-specific markers. Tissue-associated lung SAgs, collagen V (Col-V) and K-alpha 1 tubulin (Kα1T), heart SAgs, myosin and vimentin, and kidney SAgs, fibronectin and collagen IV (Col-IV), were identified using western blot. Lung transplant recipients diagnosed with bronchiolitis obliterans syndrome had exosomes with higher expression of Col-V (4.2-fold) and Kα1T (37.1-fold) than stable. Exosomes isolated from heart transplant recipients diagnosed with coronary artery vasculopathy had a 3.9-fold increase in myosin and a 4.7-fold increase in vimentin compared with stable. Further, Kidney transplant recipients diagnosed with transplant glomerulopathy had circulating exosomes with a 2-fold increased expression of fibronectin and 2.5-fold increase in Col-IV compared with stable. We conclude that circulating exosomes with tissue associated SAgs have the potential to be a noninvasive biomarker for allograft rejection.     

Clinicopathological features of low-grade malignant endolymphatic sac tumors

Background

Low-grade malignant endolymphatic sac tumor (ELST) is a rare neoplasm, occurring in the inner ear and invading the temporal bone. This study aims to investigate the clinicopathological features of low-grade malignant ELSTs.

Pituitary tumor transforming gene PTTG2 induces psoriasis by regulating vimentin and E-cadherin expression

Psoriasis is a common and intractable skin disease affecting the physical and mental health of patients. This study focused on the roles of pituitary tumor transforming gene 2 (PTTG2) in psoriasis. Using real-time quantitative PCR and western blot, the expression patterns of PTTG2 were compared in psoriatic epidermis cells and normal cells, from both mRNA levels and protein levels. Knockdown of PTTG2 by siRNA was conducted in HaCaT cells to investigate the changes in cell viability and migration in vitro. Expression changes of vimentin and E-cadherin were also detected in the transfected cells. Results showed PTTG2 was significantly overexpressed in the psoriatic epidermis cells (P < 0.05). The cell viability and migration were inhibited by the knockdown of PTTG2. Besides, knockdown of PTTG2 resulted in down-regulation of vimentin and up-regulation of E-cadherin, with significant differences compared to the siRNA control group (P < 0.05). This study indicated the involvement of PTTG2 in mediating epidermis cell viability and migration and in pathogenesis of psoriasis. PTTG2 might be a potential therapeutic target for psoriasis through inducing epithelial-to-mesenchymal transition (EMT) via regulating the expression of vimentin and E-cadherin.     

Nestin and vimentin colocalization affects the subcellular location of glucocorticoid receptor in cutaneous melanoma

Aims: Nestin (a neuronal stem cell/progenitor cell marker of central nervous system development), vimentin (which is ubiquitously expressed in mesenchymal cells), and the glucocorticoid receptor (GR, which is involved in the immune response, cell proliferation, and apoptosis) have been shown to interact in embryonic and undifferentiated tissues in modulating cell proliferation. The aim of this study was to analyse nestin, vimentin and GR expression in tumour tissue (melanoma), and their association with clinicopathological variables, to evaluate any effect on tumour progression. Methods and results: Immunohistochemistry, double‐label immunofluorescence and confocal laser scanning microscopy were performed on biopsy specimens of cutaneous melanoma from 81 patients. Fisher’s and Pearson’s tests showed a correlation between nestin, vimentin and subcellular GR location (P = 0.008). Their concomitant expression also correlated with Clark level and thickness (P = 0.02 and P = 0.029, respectively). Kaplan–Meier analysis revealed a poorer outcome for stage III and IV patients with associated expression of nestin, vimentin and cytoplasmic GR in tumour tissue (P = 0.02). Conclusions: These results suggest the presence in melanoma of growth mechanisms involving nestin, vimentin, and GR, similarly to that occurring in embryonic and undifferentiated cells, and may help in understanding tumour biology to provide a molecular basis for clinical therapies.