The relative expression levels of CD148 and CD180 on clonal B cells and CD148/CD180 median fluorescence intensity ratios are useful in the characterization of mature B cell lymphoid neoplasms infiltrating blood and bone marrow - Results from a single centre pilot study.

INTRODUCTION: The categorization of mature B cell neoplasms (MBN) infiltrating blood and bone marrow are met with difficulties. The inclusion of CD148 and CD180 in the routine flow cytometry/FCM panels has been suggested to refine the diagnosis. We studied the discriminatory ability of CD148 and CD180 median fluorescence intensity(MFI), CD148/CD180 ratio and their expression relative to T cells (CD148ab/T , CD180ab/T ), neutrophils (CD148ab/gr , CD180ab/gr ) and normal B cells (CD148ab/n , CD180ab/n ) in the differentiation of mature B cell neoplasms (MBN) especially non-chronic lymphocytic leukaemia (CLL). METHODOLOGY: The flow cytometric (FCM) expression of CD148 and CD180 was studied prospectively in 102 patients (non-CLL; n = 72); diagnosed by a comprehensive panel of immunophenotypic and cytogenetic studies. The MFI and ratios were statistically compared across MBNs by Mann-Whitney U test. Cut-off values, sensitivity and specificity were calculated for significant parameters by receiver operator characteristic curve. RESULTS: CD180MFI > 4.35 showed 100% sensitivity and 90.9% specificity for a diagnosis of marginal zone lymphoma (MZL) while, CD148/180 > 5.15 was 100% specific and 81.8% sensitive for lymphoplasmacytic lymphoma. CD148ab/T (>4.3; 100% specificity, 83.4% sensitivity) and CD148ab/gr (>1.1; 100% sensitivity, 90% sensitivity) were useful for differentiating blastoid-mantle cell lymphoma/MCL from diffuse large B cell lymphoma; while CD148MFI (≥20.25), CD148ab/T (>3.35) and CD148ab/gr (>0.95) showed >90% specificity and sensitivity for distinguishing MCL from CLL. Pairwise analysis also showed a good discriminant function of various parameters for distinguishing SMZL from other MBNs like FL, MCL as well as CLL. CONCLUSIONS: The current study shows an excellent utility of CD148MFI, CD180MFI, their ratio and relative expression levels in the subcategorization of immunophenotypically related MBNs.

Factors controlling permeability of the blood-brain barrier.

As the primary protective barrier for neurons in the brain, the blood-brain barrier (BBB) exists between the blood microcirculation system and the brain parenchyma. The normal BBB integrity is essential in protecting the brain from systemic toxins and maintaining the necessary level of nutrients and ions for neuronal function. This integrity is mediated by structural BBB components, such as tight junction proteins, integrins, annexins, and agrin, of a multicellular system including endothelial cells, astrocytes, pericytes, etc. BBB dysfunction is a significant contributor to the pathogeneses of a variety of brain disorders. Many signaling factors have been identified to be able to control BBB permeability through regulating the structural components. Among those signaling factors are inflammatory mediators, free radicals, vascular endothelial growth factor, matrix metalloproteinases, microRNAs, etc. In this review, we provide a summary of recent progress regarding these structural components and signaling factors, relating to their roles in various brain disorders. Attention is also devoted to recent research regarding impact of pharmacological agents such as isoflurane on BBB permeability and how iron ion passes across BBB. Hopefully, a better understanding of the factors controlling BBB permeability helps develop novel pharmacological interventions of BBB hyperpermeability under pathological conditions.

NOX4-driven ROS formation mediates PTP inactivation and cell transformation in FLT3ITD-positive AML cells.

Activating mutations of FMS-like tyrosine kinase 3 (FLT3), notably internal tandem duplications (ITDs), are associated with a grave prognosis in acute myeloid leukemia (AML). Transforming FLT3ITD signal transduction causes formation of reactive oxygen species (ROS) and inactivation of the protein-tyrosine phosphatase (PTP) DEP-1/PTPRJ, a negative regulator of FLT3 signaling. Here we addressed the underlying mechanisms and biological consequences. NADPH oxidase 4 (NOX4) messenger RNA and protein expression was found to be elevated in FLT3ITD-positive cells and to depend on FLT3ITD signaling and STAT5-mediated activation of the NOX4 promoter. NOX4 knockdown reduced ROS levels, restored DEP-1 PTP activity and attenuated FLT3ITD-driven transformation. Moreover, Nox4 knockout (Nox4(-/-)) murine hematopoietic progenitor cells were refractory to FLT3ITD-mediated transformation in vitro. Development of a myeloproliferative-like disease (MPD) caused by FLT3ITD-transformed 32D cells in C3H/HeJ mice, and of a leukemia-like disease in mice transplanted with MLL-AF9/ FLT3ITD-transformed murine hematopoietic stem cells were strongly attenuated by NOX4 downregulation. NOX4-targeting compounds were found to counteract proliferation of FLT3ITD-positive AML blasts and MPD development in mice. These findings reveal a previously unrecognized mechanism of oncoprotein-driven PTP oxidation, and suggest that interference with FLT3ITD-STAT5-NOX4-mediated overproduction of ROS and PTP inactivation may have therapeutic potential in a subset of AML.

Podocyte injury in pure membranous and proliferative lupus nephritis: distinct underlying mechanisms of proteinuria?

Proteinuria is a major feature of lupus nephritis (LN) and reflects podocyte injury. Analysis of podocyte biomarkers was performed attempting to identify if podocyte phenotype is distinct in pure membranous and proliferative LN. Expression of synaptopodin, Wilms tumor protein 1 (WT1), glomerular epithelial protein 1 (GLEPP1) and nephrin was evaluated in 52 LN biopsies by immunohistochemistry. Preserved synaptopodin expression was observed in only 10 (19.2%) of all biopsies while 42 (80.8%) had reduced expression. Both groups had comparable proteinuria at the time of biopsy (p = 0.22); however, in the mean follow-up of four years there was a tendency toward lower mean levels of proteinuria in patients with preserved synaptopodin staining (0.26±0.23 vs. 0.84±0.90 g/24 h, p = 0.05) compared with those with diminished expression. Thirty-nine (75%) biopsies were classified as proliferative and 13 (25%) as pure membranous. Comparison of podocyte biomarkers demonstrated a predominance of preserved staining of synaptopodin (69.2%), WT1 (69.2%), GLEPP1 (53.9%) and nephrin (60%) in the pure membranous group whereas only <10% of the proliferative showed preserved expression. Our data suggest that in proliferative forms there seems to occur structural podocyte damage, whereas in the pure membranous the predominant preserved pattern suggests a dysfunctional podocyte lesion that may account for the better long-term prognosis of proteinuria outcome.

An unbiased screen identifies DEP-1 tumor suppressor as a phosphatase controlling EGFR endocytosis.

BACKGROUND: The epidermal growth factor (EGF) stimulates rapid tyrosine phosphorylation of the EGF receptor (EGFR). This event precedes signaling from both the plasma membrane and from endosomes, and it is essential for recruitment of a ubiquitin ligase, CBL, that sorts activated receptors to endosomes and degradation. Because hyperphosphorylation of EGFR is involved in oncogenic pathways, we performed an unbiased screen of small interfering RNA (siRNA) oligonucleotides targeting all human tyrosine phosphatases. RESULTS: We report the identification of PTPRK and PTPRJ (density-enhanced phosphatase-1 [DEP-1]) as EGFR-targeting phosphatases. DEP-1 is a tumor suppressor that dephosphorylates and thereby stabilizes EGFR by hampering its ability to associate with the CBL-GRB2 ubiquitin ligase complex. DEP-1 silencing enhanced tyrosine phosphorylation of endosomal EGFRs and, accordingly, increased cell proliferation. In line with functional interactions, EGFR and DEP-1 form physical associations, and EGFR phosphorylates a substrate-trapping mutant of DEP-1. Interestingly, the interactions of DEP-1 and EGFR are followed by physical segregation: whereas EGFR undergoes endocytosis, DEP-1 remains confined to the cell surface. CONCLUSIONS: EGFR and DEP-1 physically interact at the cell surface and maintain bidirectional enzyme-substrate interactions, which are relevant to their respective oncogenic and tumor-suppressive functions. These observations highlight the emerging roles of vesicular trafficking in malignant processes.