Immunocytochemical studies of aquaporin 4, Kir4.1, and α1-syntrophin in the astrocyte endfeet of mouse brain capillaries.

One of the most important physiological roles of brain astrocytes is the maintenance of extracellular K(+) concentration by adjusting the K(+) influx and K(+) efflux. The inwardly rectifying K(+) channel Kir4.1 has been identified as an important member of K(+) channels and is highly concentrated in glial endfeet membranes. Aquaporin (AQP) 4 is another abundantly expressed molecule in astrocyte endfeet membranes. We examined the ultrastructural localization of Kir4.1, AQP4, α1-syntrophin, and ß-spectrin molecules to understand the functional role(s) of Kir4.1 and AQP4. Immunogold electron microscopy of these molecules showed that the signals of these molecules were present along the plasma membranes of astrocyte endfeet. Double immunogold electron microscopy showed frequent co-localization in the combination of molecules of Kir4.1 and AQP4, Kir4.1 and α1-syntrophin, and AQP4 and α1-syntrophin, but not those of AQP4 and ß-spectrin. Our results support biochemical evidence that both Kir4.1 and AQP4 are associated with α1-syntrophin by way of postsynaptic density-95, Drosophila disc large protein, and the Zona occludens protein I protein-interaction domain. Co-localization of AQP4 and Kir4.1 may indicate that water flux mediated by AQP4 is associated with K(+) siphoning.

Immunohistochemical detection of dysbindin at the astroglial endfeet around the capillaries of mouse brain.

Dysbindin was first identified by the yeast two hybrid assay as a binding partner of dystrobrevin which is a cytoplasmic member of dystrophin glycoprotein complex. Immunolocalization of dystrobrevin in the astrocyte endfeet and endothelial cells in the rat cerebellum was reported. Therefore, we were interested in the expression and localization of dystrobrevin binding protein dysbindin in the mouse brain capillary wall and its surrounding astroglial endfeet. We examined whether the dysbindin expression is present in astroglial endfeet and/or capillary endothelial cells at light and electron microscopic levels. Using brain samples from five normal mice (C57BL/6ScSn), we prepared the anti-dysbindin antibody stained brain samples with immunoperoxidase method at light microscopic level and with immunogold method at ultrastructural level. Immunohistochemistry showed that dysbindin was located in the brain capillary at light microscopic level. Immunogold electron microscopy revealed that dysbindin signal was observed at the inside surface of plasma membrane of glial endfeet which surrounded the brain capillary endothelial cells and pericytes.

A novel missense mutation of the XBP1 gene in diffuse large B-cell lymphoma.

The differentiation of B cells into immunoglobulin-secreting plasma cells is controlled by two transcription factors, B lymphocyte-induced maturation protein 1 (BLIMP1) and X-box-binding protein 1 (XBP1). XBP1 is a positively acting transcription factor in the CREB/ATF family that is expressed at a high level in plasma cells, and Xbp1-deficient mice were devoid of plasma cells, demonstrating that XBP1 is crucial for plasmacytic differentiation. XBP1 acts downstream of BLIMP1 and regulates a variety of genes encoding endoplasmic reticulum-associated proteins. We have previously reported mutations in the PRDM1 gene (previously BLIMP1) in 2 of 15 cases of B-cell lymphoma. Here, we describe a novel mutation in the XBP1 gene in 1 of 5 cases of B-cell lymphoma. A single-base substitution was found in exon 1 (227G>A) of the XBP1 gene in a patient with diffuse large B-cell lymphoma, resulting in a somatic missense mutation (R76K). To date, no mutations in the XBP1 gene in B-cell lymphoma have been reported. Taken together with previous reports, the present results suggest that two key transcription factors for the plasmacytic differentiation, XBP1 and BLIMP1, are involved in the pathogenesis in diffuse large B-cell lymphoma.

Cytologic and immunocytochemical features of EBV negative primary effusion lymphoma: report on seven Japanese cases.

Primary effusion lymphoma (PEL) is very rare type of non-Hodgkin's lymphoma (NHL) usually confined to the body cavities such as the pleural space, pericardium, and peritoneum. PEL is a human herpes virus-8 (HHV-8)-associated lymphoma and commonly observed in human immunodeficiency virus (HIV)-infected patients. However, HIV-infected patients are extremely fewer in Japan in comparison with those in Western countries; PEL is usually not associated with HIV infection in Japan. This report presents seven Japanese cases of PEL. In situ hybridization revealed that the PEL cells were negative for EBV in all cases. An immunocytological analysis showed that only one case was positive for HHV-8, and PEL cells were positive for CD20 in all cases. MUM1 was positive, but CD10 and CD138 were negative in six cases. One case each was positive for CD30 and BCL-6. The phenotypic patterns of HIV-related is BCL6-/MUM1+/CD138+, thus, the phenotypic findings observed by immunocytochemistry in this study were somehow different from those reported in Western countries. However, the cytomorphological features of PEL cells showed large cell size, abundant basophilic cytoplasm, coarse chromatin, and occasional binucleated or multinucleated cells, similar to a large cell immunoblastic and anaplastic large cell lymphoma, indicating that the cytomorphological characteristics of PE cells in Giemsa and Papanicolaou stain were consistent with those reported abroad. The prognosis for PEL in these cases was poor, but the survival time was variable ranging from 1 month to 54 months, and was different from that of Western cases. No p16/CDKN2A expression was observed, and one case showed PEL cells with a BLIMP1 mutation.

Three cases of extranodal NK/T-cell lymphoma of the nasal type diagnosed by nasal brush cytology.

Extranodal natural killer (NK)/T-cell lymphoma of the nasal type is a rare type of malignant lymphoma that is most common in Asian countries. Here we describe cytomorphologic, immunocytochemical, and molecular cytochemical features of three cases of NK/T-cell lymphoma of the nasal type diagnosed by nasal brush cytology. Cytomorphologic findings common among the three cases included the presence of several cell types, including nasal cavity epithelial cells, histiocytes, phagocytic histiocytes, and lymphoid cells, within a necrotic background. Suspected lymphoma cells were medium to large lymphoid cells possessing light blue and abundant cytoplasm. A characteristic feature of these cells was the presence of the tongue-like projections of cytoplasm from one or both sides of the cells. We believe these intriguing cytologic findings are indicators of NK/T-cell lymphoma of the nasal type. Azurophilic granules were observed in all cases, ranging from extremely fine granules to large granular lymphocyte (LGL)-like granules. Immunocytochemical and molecular cytochemical analyses showed staining for natural killer cell antigen CD56 as well as cytotoxic granule-associated proteins granzyme B7 (GrB7) and T-cell-restricted intercellular antigen-1 (TIA-1). EBV (Epstein-Barr virus) encoded small RNAs (EBER) positivity was shown by in situ hybridization, and no rearrangement of the TCRgamma gene was observed. Comparison between cytobrush and cotton swab methodology showed that cytobrush resulted in more cell-rich specimens than did cotton swabs, suggesting that nasal brush cytology with cytobrush is most useful in the diagnosis of NK/T-cell lymphoma of the nasal type.