Ankyrin-G induces nucleoporin Nup358 to associate with the axon initial segment of neurons.

RanBP2/Nup358 is a member of the large nucleoporin family constituting the nuclear pore complex (NPC). Depending on the cell type and the physiological state, Nup358 interacts with specific partner proteins and influences distinct mechanisms independent of its role in nucleocytoplasmic transport. Here, we provide evidence that Nup358 associates selectively with the axon initial segment (AIS) of mature neurons and mediated by the AIS scaffold ankyrin-G. The N-terminus of Nup358 is found to be sufficient for its localization at the AIS. Further, we show that Nup358 is expressed as two isoforms, one full-length and another shorter form of Nup358. These isoforms differ in their subcellular distribution in neurons and expression level during neuronal development. Overall, the present study highlights an unprecedented localization of Nup358 within the AIS and suggests its involvement in neuronal function

Overlap at the molecular and immunohistochemical levels between angioimmunoblastic T-cell lymphoma and a subgroup of peripheral T-cell lymphomas without specific morphological features

The overlap of morphology and immunophenotype between angioimmunoblastic T-cell lymphoma (AITL) and other nodal peripheral T-cell lymphomas (n-PTCLs) is a matter of current interest whose clinical relevance and pathogenic background have not been fully established. We studied a series of 98 n-PTCL samples (comprising 57 AITL and 41 PTCL-NOS) with five TFH antibodies (CD10, BCL-6, PD-1, CXCL13, ICOS), looked for mutations in five of the genes most frequently mutated in AITL (TET2, DNMT3A, IDH2, RHOA and PLCG1) using the Next-Generation-Sequencing Ion Torrent platform, and measured the correlations of these characteristics with morphology and clinical features. The percentage of mutations in the RHOA and TET2 genes was similar (23.5% of cases). PLCG1 was mutated in 14.3%, IDH2 in 11.2% and DNMT3A in 7.1% of cases, respectively. In the complete series, mutations in RHOA gene were associated with the presence of mutations in IDH2, TET2 and DNMT3A (p < 0.001, p = 0.043, and p = 0.029, respectively). Fourteen cases featured RHOA mutations without TET2 mutations. A close relationship was found between the presence of these mutations and a TFH-phenotype in AITL and PTCL-NOS patients. Interestingly, BCL-6 expression was the only TFH marker differentially expressed between AITL and PTCL-NOS cases. There were many fewer mutated cases than there were cases with a TFH phenotype. Overall, these data suggest alternative ways by which neoplastic T-cells overexpress these proteins. On the other hand, no clinical or survival differences were found between any of the recognized subgroups of patients with respect to their immunohistochemistry or mutational profile.This work was supported by grants from the Instituto de Salud Carlos III, from the Ministerio de Economía, Industria y Competitividad (RTICC RD06/0020/0107, RD12/0036/0060, PI 12/1682, PT13/0010/0007, PI16/ 01294, SAF2013-47416-R, CIBERONC-ISCIII, PIE15/ 0081, ISCIII-MINECO AES-FEDER (Plan Estatal I+D+I 2013–2016): PI14/00221, PIE14/0064, PIE15/0081 and PIE16/01294)) and the Asociación Española Contra el Cáncer, Spain. JG-R is a recipient of an iPFIS predoctoral fellowship (IFI14/00003) from ISCIII-MINECO-AESFEDER (Plan Estatal I+D+I 2013–2016). MSB was supported by a Miguel Servet contract (CP11/00018) from the ISCIII-MINECO-AES-FEDER (Plan Nacional I+D+I 2008–2011), and currently holds a Miguel Servet II contract (CPII16/00024), supported by ISCIII-MINECOAES- FEDER (Plan Estatal I+D+I 2013–2016) and the Fundación de Investigación Biomédica Puerta de Hierro.S

Mutational Profile and Pathological Features of a Case of Interleukin-10 and RGS1-Positive Spindle Cell Variant Diffuse Large B-Cell Lymphoma

Diffuse large B-cell lymphoma with spindle cell morphology is a rare variant. We present the case of a 74-year-old male who initially presented with a right supraclavicular (lymph) node enlargement. Histological analysis showed a proliferation of spindle-shaped cells with narrow cytoplasms. An immunohistochemical panel was used to exclude other tumors, such as melanoma, carcinoma, and sarcoma. The lymphoma was characterized by a cell-of-origin subtype of germinal center B-cell-like (GCB) based on Hans’ classifier (CD10-negative, BCL6-positive, and MUM1-negative); EBER negativity, and the absence of BCL2, BCL6, and MYC rearrangements. Mutational profiling using a custom panel of 168 genes associated with aggressive B-cell lymphomas confirmed mutations in ACTB, ARID1B, DUSP2, DTX1, HLA-B, PTEN, and TNFRSF14. Based on the LymphGen 1.0 classification tool, this case had an ST2 subtype prediction. The immune microenvironment was characterized by moderate infiltration of M2-like tumor-associated macrophages (TMAs) with positivity of CD163, CSF1R, CD85A (LILRB3), and PD-L1; moderate PD-1 positive T cells, and low FOXP3 regulatory T lymphocytes (Tregs). Immunohistochemical expression of PTX3 and TNFRSF14 was absent. Interestingly, the lymphoma cells were positive for HLA-DP-DR, IL-10, and RGS1, which are markers associated with poor prognosis in DLBCL. The patient was treated with R-CHOP therapy, and achieved a metabolically complete response

CD229 (Ly9) a Novel Biomarker for B-Cell Malignancies and Multiple Myeloma

CD229 is a cell-surface molecule predominantly expressed on lymphocytes. Its expression in B-cell malignancies is poorly known. We tested the presence of this immunoreceptor on a large number of malignancies and normal tissue using a new monoclonal antibody and tissue microarrays. Our data show that CD229 expression is restricted to hematopoietic cells. It was strongly expressed in myeloma and marginal-zone lymphomas. Because of the high expression on multiple myeloma cells, we also analyze the presence of soluble CD229 in the sera of these patients. We showed that serum levels of soluble CD229 in myeloma patients, at the time of diagnosis, could be useful as a prognostic biomarker. Altogether, our results indicate that CD229 represents not only a useful disease biomarker but also an attractive therapeutic target. CD229 (Ly9) homophilic receptor, which belongs to the SLAM family of cell-surface molecules, is predominantly expressed on B and T cells. It acts as a signaling molecule, regulating lymphocyte homoeostasis and activation. Studies of CD229 function indicate that this receptor functions as a regulator of the development of marginal-zone B cells and other innate-like T and B lymphocytes. The expression on leukemias and lymphomas remains poorly understood due to the lack of CD229 monoclonal antibodies (mAb) for immunohistochemistry application (IHC). In this study, we used a new mAb against the cytoplasmic region of CD229 to study the expression of CD229 on normal tissues and B-cell malignancies, including multiple myeloma (MM), using tissue microarrays. We showed CD229 to be restricted to hematopoietic cells. It was strongly expressed in all cases of MM and in most marginal-zone lymphomas (MZL). Moderate CD229 expression was also found in chronic lymphocyte leukemia (CLL), follicular (FL), classic mantle-cell (MCL) and diffuse large B-cell lymphoma. Given the high expression on myeloma cells, we also analyzed for the presence of soluble CD229 in the sera of these patients. Serum levels of soluble CD229 (sCD229) at the time of diagnosis in MM patients could be useful as a prognostic biomarker. In conclusion, our results indicate that CD229 represents not only a useful biomarker but also an attractive therapeutic target

Low Numbers of FOXP3 Positive Regulatory T Cells Are Present in all Developmental Stages of Human Atherosclerotic Lesions

BACKGROUND: T cell mediated inflammation contributes to atherogenesis and the onset of acute cardiovascular disease. Effector T cell functions are under a tight control of a specialized T cell subset, regulatory T cells (Treg). At present, nothing is known about the in situ presence of Treg in human atherosclerotic tissue. In the present study we investigated the frequency of naturally occurring Treg cells in all developmental stages of human atherosclerotic lesions including complicated thrombosed plaques. METHODOLOGY: Normal arteries, early lesions (American Heart Association classification types I, II, and III), fibrosclerotic plaques (types Vb and Vc) and 'high risk' plaques (types IV, Va and VI) were obtained at surgery and autopsy. Serial sections were immunostained for markers specific for regulatory T cells (FOXP3 and GITR) and the frequency of these cells was expressed as a percentage of the total numbers of CD3+ T cells. Results were compared with Treg counts in biopsies of normal and inflammatory skin lesions (psoriasis, spongiotic dermatitis and lichen planus). PRINCIPLE FINDINGS: In normal vessel fragments T cells were virtually absent. Treg were present in the intima during all stages of plaque development (0.5-5%). Also in the adventitia of atherosclerotic vessels Treg were encountered, in similar low amounts. High risk lesions contained significantly increased numbers of Treg compared to early lesions (mean: 3.9 and 1.2%, respectively). The frequency of FOXP3+ cells in high risk lesions was also higher compared to stable lesions (1.7%), but this difference was not significant. The mean numbers of intimal FOXP3 positive cells in atherosclerotic lesions (2.4%) was much lower than those in normal (24.3%) or inflammatory skin lesions (28%). CONCLUSION: Low frequencies of Treg in all developmental stages of human plaque formation could explain the smoldering chronic inflammatory process that takes place throughout the longstanding course of atherosclerosis

Accumulation of CCR4+ CTLA-4hi FOXP3+CD25hi Regulatory T Cells in Colon Adenocarcinomas Correlate to Reduced Activation of Conventional T Cells

BACKGROUND: Colorectal cancer usually gives rise to a specific anti-tumor immune response, but for unknown reasons the resulting immunity is not able to clear the tumor. Recruitment of activated effector lymphocytes to the tumor is important for efficient anti-tumor responses, while the presence of regulatory T cells (Treg) down-modulate tumor-specific immunity. We therefore aimed to determine homing mechanisms and activation stage of Treg and effector T cell infiltrating colon tumors compared to cells from the unaffected mucosa in patients suffering from colon adenocarcinoma. METHODOLOGY/PRINCIPAL FINDINGS: Lymphocytes were isolated from unaffected and tumor mucosa from patients with colon adenocarcinoma, and flow cytometry, immunohistochemistry, and quantitative PCR was used to investigate the homing mechanisms and activation stage of infiltrating Treg and conventional lymphocytes. We detected significantly higher frequencies of CD25(high)FOXP3⁺CD127(low) putative Treg in tumors than unaffected mucosa, which had a complete demethylation in the FOXP3 promotor. Tumor-associated Treg had a high expression of CTLA-4, and some appeared to be antigen experienced effector/memory cells based on their expression of αEβ7 (CD103). There were also significantly fewer activated T cells and more CTLA-4⁺ conventional T cells susceptible to immune regulation in the tumor-associated mucosa. In contrast, CD8⁺granzyme B⁺ putative cytotoxic cells were efficiently recruited to the tumors. The frequencies of cells expressing α4β7 and the Th1 associated chemokine receptor CXCR3 were significantly decreased among CD4⁺ T cells in the tumor, while frequencies of CD4⁺CCR4⁺ lymphocytes were significantly increased. CONCLUSIONS/SIGNIFICANCE: This study shows that CCR4⁺CTLA4(hi) Treg accumulate in colon tumors, while the frequencies of activated conventional Th1 type T cells are decreased. The altered lymphocyte composition in colon tumors will probably diminish the ability of the immune system to effectively attack tumor cells, and reducing the Treg activity is an important challenge for future immunotherapy protocols